Sidekick No More: Neural Translation Control by p70 ribosomal S6 kinase 1
Abstract and Keywords
The p70 ribosomal S6 kinase 1 (S6K1) plays a critical role in stimulus-dependent anabolism in the body and has been the focus of intense study for diseases such as diabetes, cardiac myopathies and cancers, rather than neurological disorders. This chapter evaluates the evidence for S6K1 driven protein synthesis in the brain, its role in mediating learning and memory, dysfunction in disease, and other outstanding questions. S6K1 has been studied for several decades, almost entirely in the context of being the major mediator of translational control downstream of the mechanistic target of rapamycin complex 1 (mTORC1). Interestingly, recent work in neural systems is slowly unravelling the varied roles of S6K1 as a signal aggregator, not limited to mTORC1, to affect neural translation, thus making its involvement in brain function more nuanced than earlier thought.
A Historical Timeline of S6K1 Research in Neurobiology
Ribosomal S6 kinase B1 (gene ID: 6198, EC 188.8.131.52), or p70 ribosomal S6 kinase 1 (S6K1), is a well-known serine-threonine kinase belonging to the AGC kinase superfamily (Pearce, Kommander, & Alessi, 2010). It is a member of a family of related kinases that share the nominal ability to phosphorylate S6 ribosomal protein but have differing roles in cell biology. The S6 kinase A group is commonly referred to as the p90 RSK or p90rsk kinases. The kinases of the group have four isoforms, containing an extra kinase domain, and are almost exclusively activated by mitogen-activated kinases and mainly influence transcription (Frödin & Gammeltoft, 1999). The S6 kinase B group includes two members, namely S6K1 and S6K2, which can also be referred to as p70rsk. The S6K1 and 2 split likely resulted from a gene duplication and translocation event, placing the two isoforms on different chromosomes with highly a conserved kinase domain but with divergent sequences at the N- and C-termini. This high sequence conservation at the business area of the kinases is often cited as a reason for studying S6K1 more than S6K2. However, there is a growing body of compelling evidence, stemming largely from cancer studies, that show that S6K1 and S6K2 have different cellular localization, interactomes, and disease involvement (see Pardo & Seckl, 2013 for an in-depth review). While there is an inherent distinction in the roles played by the two S6Ks, individual knock down of S6K1 or S6K2 induce compensatory upregulation of the other kinase (Nardella et al., 2011), while deletion of both causes perinatal lethality in mice (Pende et al., 2004). Due to the presence of two nuclear localization sequences, S6K2 is largely thought to regulate transcription and cell fate determination programs. There are probably a handful of studies on the exclusive role of S6K2 in the brain. Therefore, this review will discuss studies on the involvement of S6K1 in brain function in health and disease.
Contrary to the popular notion of S6K1 being an acolyte of the mechanistic target of rapamycin complex 1 (mTORC1) and protein kinase B (Akt), its existence was indicated, prior to the formal discovery mTOR (as early as 1980s), by multiple studies pertaining to growth factor signaling (Roberts & Morelos, 1982; Wettenhall, Chesterman, Walker, & Morgan, 1983; Novak-Hofer & Thomas, 1984). The kinase was subsequently cloned from rat liver extracts in 1990 (Banerjee et al., 1990; Kozma et al., 1990). In the interim, studies in neuroscience indicate that S6K1 was implicated in insulin and fibroblast growth factor signaling in astrocytes (Pierre, Toru-Delbauffe, Gavaret, Pomerance, & Jacquemin, 1986; Gavaret et al., 1989) as the only bonafide kinase to phosphorylate S6 protein that is a constituent of the 40S ribosomal unit. The first report involving S6K1 in neurons identified this kinase to be important in insulin signaling in cultured fetal neurons as well as distinct from protein kinase-C and cAMP-dependent protein kinase (Heidenreich & Toledo, 1989). In an oft-cited paper, Jefferies et al. (1997) provided a critical input that S6K1 activity was rapamycin-sensitive and hence activated by mTOR, which was then corroborated across multiple cell lines and model systems. Another finding of this study was that S6K1/2 controls translation of 5′ TOP mRNA, which subsequently has been disproved by evidence from cells that lack S6K1 and S6K2 but still show robust 5′TOP mRNA translation.
In the1990s, tremendous strides were made in parallel in the fields of molecular mechanisms of learning and memory (see Kandel, 2001; Malenka & Nicoll, 1999), identifying the translation control components and their sequence of action for proper translation (see Costa-Mattioli, Sossin, Klann, & Sonenberg, 2009; Kozak, 1992) in non-neuronal systems. Pioneering work was also done in uncovering the role of translation in learning and memory (Kang & Schuman, 1996) implicating signaling cascades that eventually were shown to require S6K1 activation. Therefore, by early 2000s, S6K1 was accepted widely as an influencer in synaptic protein synthesis. With the development of the S6K1, S6K2 knock-out mouse (Shima et al., 1998; Pende et al., 2004), researchers gained a tool to directly examine the role of S6K1 or S6K1 or both at a systems level. This facilitated a spate of studies that dissected the role of this kinase in many neurobiological phenomena that I discuss later in the chapter. This genetic deletion has also been used to dissect the role of S6K1 in multiple disease models of autism, Parkinson’s disease, Alzheimer’s disease, and schizophrenia (Bhattacharya et al., 2012; Caccamo et al., 2015; Bowling et al., 2014; Liu & Lu, 2010). Another major enabler to S6K1 research arrived in 2010 with Pfizer introducing the first small molecule inhibitor, PF 4708671 (Pearce et al., 2010b). This was followed by a handful of other reports of new blockers identified by using rational drug design approaches (discussed later in the chapter). These reagents offer methods to dissect S6K1 function in a larger array of contexts and may yield candidates for translation into clinical arenas.
As compared to the massive body of work that has focused on mTORC1 and ERK in neurobiology, S6K1 remains understudied and is largely considered one of the conduits of these critical hub kinases. Of the approximately 500 papers published involving S6K1 actions in the brain, only a handful dissect the role of this kinase alone. In contrast, there are more than 5000 papers on S6K1/2 that deal with longevity, caloric balance, muscle maintenance and memory etc., all outside the nervous system. It is perhaps time we started to closely examine the role of S6K1 in neural maintenance and function. This review is focused mainly on dissecting S6K1 mediated control of neural translation, its effects on learning, memory, behavior and its dysregulation in disease. In doing so, the review will attempt to highlight confounds, current limitations and possible modes of investigation for the future.
S6K1 Signaling and Synaptic Plasticity
Like most AGC kinases, resting S6K1 has a bilobed fold arrangement in which the N- and C-termini interact with each other, hiding the catalytic domain within (Sunami et al., 2010). The beginning of the C-terminal harbors an activation T-loop, which includes the serine threonine residues that are targets of upstream kinases and mediate the conformation change that renders the kinase domain and active site accessible. Hence S6K1 requires a carefully orchestrated set of activating signals that lead to phosphorylation of its multiple serine-threonine residues (see Magnuson, Ekim, & Fingar, 2012 for extensive explanation). This allows S6K1 to serve as a signal aggregator of many signals and stimuli initiated by a plethora of cell surface receptors. In neuronal cells, S6K1 activation has been primarily studied downstream of Trk-B- BDNF, Insulin-like growth factor (IGF1/2), metabotropic glutamate 1/5 receptor (mGluR5), and dopamine D1/2 receptors (Becker, Ibrahim, Cui, Lee, & Yee, 2011; Bhattacharya et al., 2012; Bowling et al., 2014; Zhou, Lin, Zheng, Sutton, & Wang, 2010).
Neurotrophin and growth factor signaling stimulation canonically activates phosphatidyl inositol 3 kinase (PI-3K) and protein kinase B (or Akt) and PDK1. As receptor tyrosine kinases, growth factor signaling also activates the Ras-Raf pathway that induces mitogen-activated kinase (MAK kinase) signaling. Akt inhibits the TSC1-2 complex that then relieves the brake on mTORC1, causing phosphorylation of Thr 389, which strongly cooperates with the Thr229 site phosphorylation mediated by PDK1 (Magnuson et al., 2012). MAP kinase activation of S6K1 has been mainly studied in context of extracellular—signal regulated kinase (ERK1/2) which phosphorylates Ser 421 on S6K1 (Lehman, Calvo, & Gomez-Cambronero, 2003). ERK mediated phosphorylation may occur through influencing TSC1-2 activity differently as reported by Winter, Jefferson, & Kimball (2011). Apart from mTORC1 and ERK1/2, S6K1 may also be activated by Rho-GTP-cdc 42 signaling and atypical protein kinase C isoforms λ and ζ, though the precise phosphorylation sites have yet to be explored in great detail (Fang et al., 2003; Chuo and Blenis, 1996; Romanelli, Martin, Toker, & Blenis, 1999). S6K1 is also reported to be activated at Thr 371 by glycogen synthase kinase 3β (GSK-3 β) facilitated by mTORC1 (Shin, Wolgamott, Yub, Blenis, & Yoona, 2011). S6K1 activity is regulated by dephosphorylation by PP2A in which the kinase forms a complex (Hahn, Miranda, Francis, Vendrell, Zorzano, & Teleman., 2010). Acetylation by p300/CBP (cAMP responsive element binding protein binding protein) and PCAF (p300/CBP associated factor) at the C-terminal domain which antagonizes Thr 389 phosphorylation and kinase activity has been reported to occur in response to Sirtuin signaling (Hong, Zhao, Lombard, Fingar, & Inoki, 2014). S6K1 also undergoes ubiquitination via ROC1 action in response to mitogenic activity, which may serve as a negative feedback to S6K1 signaling (Wang et al., 2008). These latter mechanisms have not been investigated in any detail in a neurobiological context.
Once active, S6K1 phosphorylates an array of targets which channel pro-anabolic signals downstream to translation, mRNA quality control, transcription, ribosome biogenesis leading to cell growth, motility, differentiation, synaptic plasticity and changes in neuronal morphology. Relevant to mRNA processing and translation are eukaryotic initiation factor 4B (eIF4B), eukaryotic elongation factor 2 (eEF2) via suppressing eEF2 kinase, PDCD4, SKAR and the traditional ribosomal protein S6 (Magnuson et al., 2012). S6K1 was also purported to phosphorylate Fragile X Mental Retardation Protein (FMRP), a key translation regulatory protein (Narayanan et al., 2008). It was believed to be the only known kinase for FMRP before work by Bartley, O’Keefe, & Bordey (2014) disproved this assertion. In this study using two knockout of Tsc1, the authors elevated mTORC1-S6K1 signaling, which did not affect Ser499 phosphorylation of FMRP. They also showed that this phosphorylation remains intact in S6K1 KO mice. Concurrent phosphorylation of eIF4B (activating) and PDCD4 (inhibiting) increase the helicase activity of eIF4A, which then promotes translation initiation. At the same time, phosphorylation of eEF2K by S6K1 inactivates it, which then relieves the dampening of eEF2 which in its dephosphorylated form mediates translation of ribosome of the loaded mRNA. SKAR serves as a conduit to recruiting S6K1 to exon junction complexes of newly processed mRNA for serving as a quality controller for pioneer round of translation (Ma, Yoon, Richardson, Julich, & Blenis, 2008). Ribosomal S6 protein is phosphorylated by S6K1, but is also a target of RSK sister family of kinases. Feedback signaling of S6K1 attenuates growth factor signaling by way of IRS-1, mTORC1 by acting on Raptor and GSK-3 and on mTORC2 by phosphorylating Rictor (Harrington et al., 2004; Julien, Carriere, Moreau, & Roux, 2010).
The role of mTORC1 and ERK 1/2 in early and long-term plasticity has been exhaustively studied (see Graber, McCamphill, & Sossin, 2013; Kelleher, Govindarajan, & Tonegawa, 2004). This was largely made possible by the availability of specific small molecule inhibitors (like rapamycin and SL-327) that are blood-brain-barrier permeable. In such conditions of mTORC1 and ERK blockade, S6K1 has been mostly used as a readout examining levels of phosphorylated Thr 389 or 421 with the assumption of almost linear signal transmission. This has led to a skewed informational database on S6K1 activity, which suggests its involvement in many synaptic plasticity paradigms ipso facto mTOR and ERK activation. Only recently have investigators commenced using S6K1 specific blocker PF-4708671, so most previous work has been with the S6K1 KO mouse. Antion, Hou, Wong, Hoeffer, & Klann, (2008a) investigated traditional long-term potentiation (LTP) paradigms in this S6K1 KO mice to find that the deletion of S6K1 did not impair L-LTP evoked by 4 high frequency trains stimuli (4x HFS), 2x HFS and theta-burst stimulation. Intriguingly, the authors found that S6K1 deletion impaired E-LTP, which is believed to be protein-synthesis independent. This may indirectly be contributed by the abrogation of negative feedback to IRS-1, leading to a pervasive increase of Akt phosphorylation. Recently, abrogation of S6K1 target- Serine 366 site on eEF2K in Aplysia has shown that it impairs long-term facilitation (LTF, McCamphill, Ferguson, & Sossin, 2017).
The protein synthesis dependence of LTD in the hippocampus is well documented (Huber, Kayser, & Bear, 2000; Graber et al., 2013). While there exists much debate on whether translation initiation or elongation is the greater contributor to LTD, evidence for the contribution of S6K1 to the process is conflicting, at best. Reports show that rapamycin and W13 (inhibiting CamKII) incubation of WT hippocampal slices does abrogate LTD and decreased S6K1 activation (Sethna et al., 2016). Additionally, mice with δRGD deletion of TSC2 (Chevere-Torres et al., 2012) showed no appreciable S6K1 activation but had enhanced ERK1/2 signaling, yet impaired LTD. However. induction remains intact with an increase in S6rp phosphorylation and translation of elongation factor 1 alpha in S6K1 KO LTD (eIF1a; Antion et al., 2008a). Subsequently, S6K1 knock down was shown to have no effect on manifestation of LTD induced by mGluR5 activation while also rescuing exaggerated LTD in Fmr 1 KO model of Fragile X syndrome (Bhattacharya et al., 2012). Therefore, it appears that in conditions where S6K1 is developmentally ablated, LTD expression is possibly routed through sister kinases like S6K2, Rsk etc. In conditions where S6K1 exists and is acutely perturbed by targeting signaling partners, LTD and perhaps LTP is impaired. This may arise from S6K1 being present in multiple signaling complexes that change according to stimulus and localization. An interesting example of such dynamics is provided by Bernard et al. (2013), where early life seizure induced enhancement in mGluR-LTD is S6K1 and protein synthesis dependent. Activated S6K1 levels in this model are not enhanced immediately after the insult, but accumulate slowly, showing a significant increase by postnatal day 60. S6K1 was found to be in a dynamic complex with FMRP and protein-phosphatase 2A (PP2A) which changed localization from cytosolic to synaptic compartments on the induction of LTD.
Though S6K1 is ubiquitously expressed, important in a variety of tissues, the available commercial antibodies do not work well in immunohistochemical experiments and have largely been used for western blot analyses. In the mouse brain, in situ hybridization using antisense probe to S6K1 reveals high expression in the hippocampal formation and sparse expression across the rest of the areas (http://mouse.brain-map.org/gene/show/48349; Lein et al., 2007), which is perplexing given notable levels of S6K1 detected in striatum, cortex, and cerebellum. Within a neuron again, epitope tagged S6K1 have been expressed that show localization in the cell body and neurites (Cammalleri et al., 2003). A very recent study shows that application of PF-4708671 can reduce S6 ribosomal protein phosphorylation across the post synaptic cell (Pirbhoy, Farris, Steward, 2017). Hence data of direct S6K1 visualization is limited and is an important area of research given the extensive nature of decentralized function in neuron and glia (Jung, Gkogkas, Sonenberg & Holt, 2014). In terms of translation, and in response to a specific stimulus, the amount of somatic and dendritic protein synthesis that is controlled by localized pools of S6K1 is still very limited. An associated question is, do more somatically localized S6K1 bear greater contribution to transcription over translation? Finally, it is unclear if only certain neurons are recruited in response to a specific stimuli (like in fear conditioning) and hence tracking S6K1 activity at whole tissue level would again dilute effects. In summary, though much is purported to be known about S6K1 signaling and synaptic plasticity, many confounds and contradictions still persist.
S6K1, Translation and Structural Plasticity of Neurons
On the heels of the finding that mTORC1 and ERK1/2 underwrite many critical aspects of LTP and LTD came reports of their implication in structural plasticity of spines and dendritic arbors (Wu, Deisseroth, & Tsein, 2001, Jaworski & Sheng, 2006). Subsequently, several studies linked local and general protein synthesis to spine changes and axon regeneration (Piper et al., 2006; Verma et al., 2005). In 2005, Jaworski, Spangler, Seeburg, Hoogenraad, & Sheng demonstrated how over activation of Akt, or depression of mTORC1 and S6K1 modulated dendritic arbors and neuron morphology in cultured cells. Jointly, S6K1 had already been implicated in cell size and longevity (Pende et al., 2004; Selman et al., 2009). A sufficient number of studies show clear implication of S6K1 in structural morphology in neurons, some independent of the involvement of protein synthesis.
Evidence of a translation-driven role for S6K1 in structural plasticity is manifold. Just overexpressing a constitutively active (T389E) species of S6K1 can increase neuronal complexity (Dwyer, Maldonado-Aviles, Lepack, DiLeone, & Duman, 2015). S6K1 inhibition by either genetic deletion or pharmacological inhibition, normalized both enhanced protein synthesis and increased filopodial spines in the mouse model of fragile X syndrome (FXS; Bhattacharya et al., 2012; Bhattacharya et al., 2016). Surprisingly, in S6K1 KO, no appreciable size difference was seen in neurons or in spine density from littermate wild-type (Bhattacharya et al., 2012). But this was only one brain area and done at one specific age. In the case of wild-type CA1 neurons though, S6K1 inhibition did cause an increase in spine density (Bhattacharya et al., 2016). Bowling et al. (2014), demonstrated the Haloperidol, a typical antipsychotic, increased protein synthesis in striatal neurons and dendritic morphology that was substantially reduced upon expression of an shRNA against S6K1. This was also found to decrease Haloperidol-induced increases in translation. A proteomic survey done in this study showed proteins that regulated cytoskeletal rearrangements and scaffold proteins that hold up spines are maximally activated upon Haloperidol stimulation, implicating a role of S6K1 in this process. In Angelman Syndrome model mice, harboring a deletion of UBE3A proteosomal unit, hippocampal slices treated with PF-4708671 (specific S6K1 inhibitor) normalized actin polymerization, LTP, similar to rapamycin (Sun et al., 2016) in indicating that normalizing proteostasis leads to changes in spines as well. Ancillary to these spine related effects, a report by Yang et al., (2014) demonstrated that S6K1 and 4E-binding protein (4E-BP) play different roles in axon regeneration in the murine optic nerve crush model. S6K1 activation in this model promotes regeneration while a complimentary study (Gong et al., 2015) showed that activating 4E-BP or repressing S6K1-GSK3β impedes the process by interfering with mTORC1 signaling and translation. Finally, S6K1 dependent translation of myelin basic protein was reported by Michel, Zhao, Karl, Lewis, Fyffe-Maricich (2015), shown to be more ERK-dependent than mTORC1. These data are summarized in Figure 1 showing protein synthesis dependent interactions in violet arrows.
In contrast, there also exist studies that showcase S6K1’s role in structural plasticity independent of translation control (shown in Figure 1 with green arrows). As early as 1998, Burnett et al., using a yeast-two hybrid screen had identified Neurabin (Neural tissue specific binding protein) as a binding partner for S6K1. Neurabin was subsequently found to be a neuronal spinophilin, which interact with last five amino acids of S6K1 and increases its phosphotransferase activity. Subsequently, Buchsbaum, Connolly, & Feig, 2003, verified that spinophilin binding brings S6K1 in proximity of Rac, activating this pathway that is critical to changes in spine architecture. S6K1 has been shown to phosphorylate Rictor in the mTORC2 complex (known to regulate actin remodeling) in its feedback to Akt in non-neuronal systems (Julien et al., 2010). Contradicting this is the report that in the Rictor knock out mice there are spine changes with no concomitant change in S6K1 activation levels (Huang et al., 2013), leaving the room for further scrutiny of this signaling loop in the future. Further support for an mTOR independent pathway to S6K1 that impinges in spinogenesis was provided by reports like Lai, Liang, Fei, Huang, & Ip (2015) where BDNF- induced spine changes were shown to involve cyclin dependent kinase 5 (Cdk 5) which phosphorylated S6K1 at Ser 411, leaving the Thr 389 site untouched. In this pathway S6rp seemed to be the downstream S6K1 target rather than eEF2K. It remains to be verified how S6rp causes no changes in ribosomal processivity in this case. A very recent report (Al-Ali et al., 2017) of spinal injury in mice reported that S6K1 inhibition promoted regeneration and is a potential “druggable” target for spinal injury conditions. The authors suggest that PF-4708671 application or siRNA knock down of S6K1 in cultured hippocampal cells increased neurite outgrowth. This blockade of S6K1 activates the feedback loop of S6K1 to PI3K which in turn activates mTORC1 and 2, though it abrogates any translation dependent effects downstream of S6K1.
Taken together, it is clear that S6K1 intimately influences neuronal architecture and structural plasticity. Given the neuro-centric thrust of most studies, it is likely that the same effect should show up in other neural cells as well. S6K1 can modulate this process both by translation and translation-independent modes, though the downstream effects through both mechanism remain to be worked out.
S6K1 in Behavior
Behavioral studies have provided the keenest insight into signaling distinctly mediated by S6K1. In this section, behavioral results of modulating S6K1 in wild type animals will be discussed, while the following section addresses the effects of S6K1 in rodent disease models. A key but less highlighted feature of most studies discussed here: the effect of genetic versus pharmacological removal of S6K1 is not always concurrent.
The first behavioral study contrasted the effects of knocking out S6K1 versus S6K2 on mouse behavior (Antion et al., 2008b). Ablation of S6K1 affected early onset of fear memory, conditioned taste aversion (CTA) and impaired acquisition of platform location in Morris Water Maze (MWM) test of spatial memory. Another important phenotype was hypoactivity in S6K1 KO mice, which has been employed later to titrate optimum dosing of S6K1 inhibitors (Huynh, Santini, & Klann, 2014; and Bhattacharya et al., 2016). In contrast, deletion of S6K2 KO reported decreased contextual fear memory, a reduction in latent inhibition of CTA, and intact spatial learning in MWM (Antion et al., 2008b). An important aspect to note here is the mixed background of knock out mice that have been speculated to contribute to the behavioral phenotypes observed. Another set of behavioral test battery was carried out in Bhattacharya et al., (2012) to test the efficacy of genetically deleting S6K1 to rescue phenotypes of fragile X syndrome (FXS). In this study, background was a mix of 129/SvJ and C57/Bl6, with a greater contribution of the latter due to more frequent backcrosses. S6K1 KO alone continued to remain hypoactive with a diminished performance in the rotarod test as well. S6K1 KO mice showed indifferent preference to social approach test (mice vs. object), with a distinct preference to the novel over the familiar mice in the novelty phase of the test. The mice also showed no impairment in behavioral flexibility as tested by the Y-maze, but buried more marbles in the eponymous name which may correlate to increased preservative/anxiogenic behavior. Interestingly, S6K1 abrogation in FXS model mice rescued a wide range of phenotypes, to be discussed further. In 2016, pharmacologic blockade of S6K1 in WT mice using two small molecule inhibitors showed no effect on social preference while slightly increasing reversal times in the Y-maze, though both showed enhanced marble burying (Bhattacharya et al., 2016).
A separate approach to model depression and the implication of S6K1 therein was adopted by Dwyer et al., (2015) using viral microinjection in the prefrontal cortex (PFC). Expression of constitutively active form of S6K1 (T389A δ CT) had an anti-depressant effect on rats, with increased locomotion, decreased times in forced swim test and abrogated the decreased sucrose preference after chronic stress. Suppression S6K1 signaling by expressing a kinase inactive form (K100R) showed opposite effects and blocked the anti-depressant effects of ketamine. This study established a strong connection between the state of S6K1 activity and depressive phenotypes. It remains to be validated by a pharmacologic approach.
S6K1 has also been implicated to mediate gustatory learning (Belelovsky, Kaphzan, Elkobi, & Rosenblum, 2009) using the CTA paradigm. However, S6K1 does not contribute to feeding behavior directly, though it regulates glucose homeostasis. Shown in a report using a conditionally driven S6K1 in AgrP and POMC neurons of hypothalamus, the deletion of kinase changed intrinsic excitability of these neurons (Smith et al., 2015). This effect of S6K1 is thought to be intimately related to its role in regulating metabolism.
The role of S6K1 in fear memory has been better dissected as fallout of interest in the contribution of protein synthesis in various phases of this learning paradigm. S6K1 signaling also seems to be region-specific in fear, a point that is not well appreciated. For instance, Gafford, Parsons, & Helmstetter (2013) measured phospho-S6K1 levels 1, 10 and 36 days after contextual fear conditioning in the dorsal hippocampus (DH) and anterior cingulate cortex (ACC). 1 day after phospho-S6K1 levels were elevated in the DH with an increase only after 36 days in the ACC, alluding to the temporally spaced translational phases of memory. S6K1 seems to mediate not memory acquisition, but rather reconsolidation and extinction. Hyunh et al. (2014) examined the dependence of cued fear memory retrieval/reconsolidation on S6K1 activity. They reported that the effect of rapamycin inhibition on retrieval could only be recapitulated if both eIF4E and S6K1 were blocked simultaneously. S6K1 did underwrite the persistence of reconsolidated memory 10 days later. This seemed to impinge via ERK signaling to Ser 421 site S6K1. In a follow-up paper in 2017, the same team showed that this ERK-S6K1 signaling is key to within-session extinction learning in the BLA and involves phosphorylation of AMPA receptor sub unit GluA1 (Huynh et al., 2017).
Overall, current data suggests that in most behaviors that have previously been shown to require mTORC1 activation, there exists a reliance on S6K1 signaling. What is implicit in the reports is the region specificity of this effect, though no concerted effort has been devoted in closely examining this issue. There is also significant divergence in behavioral phenotypes when S6K1 is genetically ablated versus shorter time suppression by blockers or viral expression of mutant kinase. It is now possible to undertake circuit limited experiments where S6K1 would be perturbed in some cells and intact in others to see how the kinase modulates behavioral outcomes at this level.
S6K1 in Neurological Disease
There is considerable ongoing effort in studying the role of S6K1 in etiology of diabetes, obesity, liver function and cancer (Um, D’Alessio, & Thomas, 2004; Savinska et al., 2004; Ben-Hur et al., 2013; Tavares et al., 2015). Alongside, the Akt-ERK-mTOR signaling in neurology has implications in neurodevelopmental, neurodegenerative and traumatic injury conditions. S6K1 plays a substantial role in the pathology of disorders caused by this signaling nexus but also in implicated in the pathology of certain conditions in a stand-alone capacity.
Given the central role that mTOR plays in anabolism and development, dysregulated mTORC1 signaling in neurodevelopment disorders (NDD) is expected. The best studied model in this subset of conditions is FXS. S6K1 was shown to be elevated as a part of dysregulated mTORC1 signaling in the Fmr1 KO mice hippocampus (Sharma et al., 2010) and, in patient-derived fibroblasts, lymphocytes and post-mortem brain tissue (Kumari et al., 2014; Hoeffer et al., 2012). Therefore S6K1 seemed to be intimately associated with the eccentric protein synthesis seen in FXS and a potential target for intervention. Genetic deletion of S6K1 in the background of Fmr1 loss (Bhattacharya et al., 2012) resulted in restored proteostasis, mTORC1 signaling, synthesis of FMRP target proteins relevant to synaptic plasticity, DHPG-evoked LTD and abnormal filopodial spines. In terms of behavioral phenotypes, S6K1 ablation rescued behavioral inflexibility and inappropriate social interaction, but not marble burying repetitive behavior. Given S6K1 KO are small (Shima et al., 1998), rescue of weight gain was an expected fallout. Additionally, macroorchidism in FXS has been shown to be caused by abnormal proliferation of Sertoli cells (Slegtenhorst-Eegdeman et al., 1998)—a process known to require S6K1 activity downstream of follicle stimulating hormone (FSH; Lécureuil et al. 2005). Therefore, rescue of macroorchidism by S6K1 genetic deletion was again expected. A key issue unresolved in this study was whether the effects were mediated by S6K1 inhibition on translation initiation or elongation.
However, the effects of pharmacologically targeting S6K1 in FXS, in neurodevelopmentally complete adult mice, were published recently (Bhattacharya et al., 2016). Head-to-head comparison of two S6K1 blockers, PF-4708671 and FS-115 was done in Fmr1 and wild-type littermates. FS-115 is more brain-penetrant and remains in the CNS for a longer time than PF-4708671. While both agents reduced translation in hippocampal and cortical brain preparations, they preferentially affected eEF2 and S6 phosphorylation rather than eIF4B, implying an elongation bias in FXS. Additionally, increased availability of FS115 in the brain of treated FXS mice normalized marble burying behavior, in part due to continued S6K1 blockade or effects on other AGC kinases. PF-4708671 was only able to correct macro-orchidism, while FS-115 impacted both phenotypes arguing for a CNS-driven control of weight and metabolism in FXS. Both agents successfully countered abnormal social interactions and behavioral inflexibility, with higher dosing acutely inducing a hypo-locomotor effect. Through these studies, it is apparent that S6K1 mediated protein synthesis plays a key role in FXS phenotypes, but requires more pre-clinical validation before it can be considered for clinical trials. For example, the effect of short-term S6K1 inhibition on LTD remains to be clarified as does studying S6K1 depression in other brain areas like the amygdala that underpins the anxiety phenotypes in seen with this syndrome.
An allied effect of S6K1 in NDD was reported recently by Huang, Chen, & Page (2016) in a germline, heterozygous Pten knock down model. The authors found a transient spike in mTORC1-S6K1 activity at post-natal day 14 (P14) in Pten +/– mice pups that caused increased axonal branching and connectivity between the medial PFC to the basolateral amygdala (BLA). This hyper-connectivity was coincident with impaired social behavioral and attendant anxiety in these mice. Interestingly, S6K1 blockade between P4 and P14 using PF-4708671 was successful in normalizing this phenotype, but a similar dose regime in adulthood did not have any effect. The authors also report that a similar spike in mTORC1 activity is also seen in young FXS mice, which argues for a conserved etiology of regional alterations in signaling and connections in syndromic models of ASD. Additionally, Angelman syndrome is reported to have imbalances in proteostasis, which are addressable by manipulating S6K1 signaling. Sun et al (2016) reported that improved LTP and spine maintenance could be achieved by administering PF-4708671, rapamycin and mTORC2 activator A-443654. S6K1 inhibition can counteract the phenotypes of STRADA deletion that cause rare neurodevelopmental disorders called PMSE (polyhydramnios, megacephaly, symptomatic epilepsy; Parker et al., 2013). STRADA, or STE20-related kinase adaptor alpha, is a pseudo-kinase that binds LKB1, a kinase upstream of AMP-activated kinases, and monitors energy homeostasis, cell size and senescence in cells.
In contrast, the relationship in aging between protein synthesis and S6K1 signaling appear to be complex. While it is well established that protein synthesis declines in the aging brain (Schimanski & Barnes, 2010), equally strong evidence exists for the beneficial effects of suppressing S6K1 signaling to improve longevity across multiple models (Selman et al., 2009; Kennedy & Kaeberlein, 2009). At the outset, these two phenomena seem contradictory, and the defining quality could well be the timing of the intervention. That S6K1 signaling should be lower in the aging brain can be reconciled in the light of increased oxidative stress, autophagy and eIF2alpha stress signaling (Keller et al., 2004; Ma et al., 2013; Massaad & Klann, 2011). However, S6K1 hyperactivity contributes to increased insulin resistance in neurodegenerative conditions (Cholerton Baker, Craft 2013), which can trigger aging. In support of the latter model, Caccamo et al., (2015) presented work that shows that reducing S6K1 expression improved spatial memory and synaptic deficits in the 3xAD mouse model for Alzheimer’s disease by suppressing the translation of Tau and BACE1. This study solely focuses the effect of having reduced S6K1 expression a priori before onset of AD on disease pathology. Though not implicating any upstream signaling effects, the authors squarely demonstrate that the effect involves translation control and reinforces the fact that suppressing the translation of specific pathologic proteins in AD is of therapeutic value. However, if intervention is to be attempted after the advent of amyloid β in the system, Zare, Motamedi, Digaleh, Khodagholi, and Maghsoudi (2015) suggest that activating S6K1 can help prevent cell death. Similarly, suppression of S6K1 signaling leads to neuronal death in Parkinson disease models (Xu et al., 2014). In the current state, it is unclear what the biggest contributor to S6K1 signaling is in a degenerating brain and if it is entirely neuronal. A plausible hypothesis is that S6K1 signaling changes in response to peripheral metabolism and at some point uncouples from mTORC1 in aging brain. It is clear that much work will be devoted to this solving this conundrum in the future.
Finally, efforts of all pre-clinical studies converge upon a search for a translatable therapeutic approach. Most therapeutic efforts in the industry are geared to controlling S6K1 in the context of obesity, cancer and diabetes. The key breakthroughs have come with the advent of a slew of small molecule inhibitors, from Pfizer (PF-4708671; Pearce et al., 2010), Axon MedChem (LY2584702; Tolcher et al., 2014), Sentinel Oncology (FS-115; Bhattacharya et al., 2016), and a battery of candidates from rational drug design screens (Qin et al., 2015). Currently. there are no clinical trials ongoing targeting S6K1 for any disease. S6K1 can also be inhibited by multi-specificity AGC kinase inhibitors like H89 2HCl and AT 13148 (Xi et al., 2016; Liu et al., 2016). Efforts to target S6K1 for neurological disorders using these molecules are at a relatively early stage, with the key challenges being to find blood-brain-barrier-permeable agents and titrating the ideal dosage for increased effectiveness while minimizing side effects such as decreased locomotion.
Outstanding Questions, Experimental Limitations, and Looking Forward
Dedicated research by multiple groups across the world is slowly bringing S6K1 out of the shadow of mTORC1 and ERK1/2 signaling. That S6K1 is a strong regulatory member of cellular signaling is well accepted outside the central nervous system, but this notion has been slow to spread for neurobiology, in part due to the absence of reliable small molecule inhibitors. However, some serious experimental limitations still exist in the field that require acknowledgement prior to addressing them. Additionally, there seems to be a paucity of clinical information about S6K1 that will impede future translational work. Next I attempt to collate the many pitfalls that plague holistic S6K1 research now, which must be investigated, interpreted and overcome in the future.
Better Reagents Fuel Better Research
Neuronal translation is equal parts somatic and decentralized in processes (dendritic and axonal). While this notion is strongly entrenched in neurobiology, another form of translational localization is becoming more apparent. For neurons to work effectively, astroglial and oligodendritic cells need to also perform their functions effectively. Our view of translation has been constrained to only consider the neuronal aspects; it is time to embrace that it is very likely that there are orchestrated changes in translation across neurons and glial cells to respond to any stimulus. This becomes relevant when one considers that S6K1 is ubiquitously expressed in all cells and likely will regulate translation differently downstream of cell-specific or cell autonomous cues. This should be an easily achievable goal with immunohistochemistry.
However, in reality, experimenters routinely note the intractability of most commercially available phospho-S6K1 antibodies for use in brain sections. Therefore, usually phosphorylated S6 ribosomal protein is used as a proxy, which is influenced by activation status of Rsk kinases and actually provides a measure of the level of translational activation. A quick look at the product sheets of these antibodies shows that almost all validation is done in non-neuronal cells and hardly any in tissue lysates or fixed section. Probing S6K1 KO mouse brain lysates with these antibodies is currently the best way to determine specificity, which becomes tedious for researchers without access to such control samples. Since a large number of these primary antibodies are rabbit polyclonals, lot to lot variations is a major concern. This may be alleviated with more monoclonal and recombinant choices appearing in the market. A reliable and consistent antibody source for S6K1 that works in multiple applications will address a slew of questions about relative localization of activated and non-activated forms of this kinase in polarized neural cells.
There is also a lack of genetic reagents to perturb S6K1 function that are readily deployable for neuroscience. Conditional knockdowns of S6K1 exist, but have been used only sparingly for neurobiological studies (Smith et al., 2015). It is unclear how much upregulation of S6K2 occurs in global deletion of S6K1, since no reliable antibody exists for this isoform. Bowling et al., employed an effective shRNA for S6K1 in neuronal cultures which paradoxically hasn’t been used often. With the advent of CRISPR/Cas9 technology, it should become easier to develop guides that knockdown S6K1 in targeted fashion. AAV2 constructs with S6K1 are available (Dwyer et al., 2015), and we should see more studies with these reagents being published in the future, however conditional Tet On/Off systems have yet to be made.
Natural Variation to Discover Phenotype Stratification
The Val633Met variation in BDNF is perhaps one of the most intensely studied examples of a genetic change in neuropsychiatry. Usually naturally occurring variants that are not immediately disease causing but prodromal or correlate with early onset or symptom severity have been less studied in neurobiology as compared to cancer. The translation control pathway is usually under tremendous purifying pressure to maintain them error-free and seldom come up in large genetic studies. That said, duplications and truncations that enhance S6K1 activity have been found in breast cancers and other solid tumors. Additionally, ExAC (http://exac.broadinstitute.org) and other mutation aggregator websites provide a list of common, rare and ultra-rare S6K1 exonic variants that exist in humans across the world. These are likely to cause subtle changes in enzyme activity that first need to be cataloged and then cross-referenced in existing exome databases for their presence or absence in specific patient populations. In the light of small molecules being developed for S6K1, knowing how these would react with variant S6K1 proteins will be critically important in determining candidate stratification and treatment outcome measures in trials feature S6K1 inhibition. With increased cognizance given to the role that altered metabolomics plays in the expression of neurological disease, studying S6K1 variants gathers greater significance.
Steady State versus Activity-Driven Translation
In almost all studies of S6K1, and, to an extent, translation control in neural systems, no distinction is made between steady-state and stimulus-driven states. This may be in unconscious bias since any brain function is a combinatorial outcome of these two phases, but this becomes important when considering pharmacological interventions. For example, unpublished data from multiple sources allude to that fact that perturbing S6K1 activity in slices incubated in ACSF alone with no stimulation will not affect translation as profoundly as rapamycin would. However, the same perturbation in the wake of agents that induce chemical LTP/LTD have a more dramatic effect. The reduced role of S6K1 in steady state translation is also alluded to by the relatively benign effects of complete deletion in knock out animals. It is very likely that S6K1 is actively involved in stimulus-driven state changes in neural systems, which is exemplified by its critical role in glial transformation (Nakamura, Garcia, & Pieper, 2008). More refined experiments are required to clearly delineate this mechanism.
What, Where, When and How Much?
It is well documented that behavior and related neurobiological phenomena are related in many instances to translation by S6K1. This translation control not only has a temporal order, but likely happens in the different cells making up the tri-partite synapse at different rates, producing different proteins. Thus far, most studies have focused on the “when,” and future studies must look to answer the “where,” “what” and “how much.” The will become critical as it is increasingly being noted that different brain areas and circuits may regulate their proteomic flux differently in the face of the same stimulus. Hence a system administration of a drug (like rapamycin or PF-4708671) will impact all areas equally and lead to off-target results in neural circuits that don’t increase translation in response to a signal. Knowing in what translation case S6K1 is downstream of mTORC1 and in which S6K1 acts independently will help sort out which ways to deliver the drug, what drug combinations will work etc. Neurobiology as a whole is shifting to acknowledge that region and sub-type specificity in circuits underlie much of what the brain expresses. It is time that the proteomic and molecular map is also put in place to track these changes and help inform future drug development for disease conditions.
A. B. is supported by funds from the Department of Biotechnology, Government of India and the FRAXA Research Foundation. I apologize for not including the work of all authors in the interest of presenting a focused view of the existing literature and highlighting areas that require greater attention in S6K1 biology.
Al-Ali, H., Ding, Y., Slepak, T., Wu, W., Sun, Y., Martinez, Y., … Bixby, J. L. (2017). The mTOR Substrate S6 Kinase 1 (S6K1) is a negative regulator of axon regeneration and a potential drug target for central nervous system injury. Journal of Neuroscience, 37(30), 7079–7095.Find this resource:
Antion, M. D., Hou, L., Wong, H., Hoeffer, C. A., & Klann, E. (2008a). mGluR-dependent long-term depression is associated with increased phosphorylation of S6 and synthesis of elongation factor 1A but remains expressed in S6K-deficient mice. Molecular Cellular Biology, 28(9), 2996–3007.Find this resource:
Antion, M. D., Merhav, M., Hoeffer, C. A., Reis, G., Kozma, S. C., Thomas, G., … Klann, E. (2008b). Removal of S6K1 and S6K2 leads to divergent alterations in learning, memory, and synaptic plasticity. Learning & Memory, 15(1), 29–38.Find this resource:
Banerjee, P., Ahmad, M. F., Grove, J. R., Kozlosky, C., Price, D. J., & Avruch, J. (1990). Molecular structure of a major insulin/mitogen-activated 70-kDa S6 protein kinase. Proceedings of National Academy Science of USA, 87(21), 8550–8554.Find this resource:
Bartley, C. M., O’Keefe, R. A., & Bordey, A. (2014). FMRP S499 Is Phosphorylated Independent of mTORC1-S6K1 activity. PLoS ONE, 9(5), e96956.Find this resource:
Becker, M. A., Ibrahim, Y. H., Cui, X., Lee, A. V., & Yee, D. (2011). The IGF pathway regulates erα through a S6k1-dependent mechanism in breast cancer cells. Molecular Endocrinology, 25(3), 516–528.Find this resource:
Belelovsky, K., Kaphzan, H., Elkobi, A., & Rosenblum, K. (2009). Biphasic activation of the mTOR pathway in the gustatory cortex is correlated with and necessary for taste learning. Journal of Neuroscience, 29(23), 7424–7431.Find this resource:
Ben-Hur, V., Denichenko, P., Siegfried, Z., Maimon, A., Krainer, A., Davidson, B., & Karni, R. (2013). S6K1 alternative splicing modulates its oncogenic activity and regulates mTORC1. Cell Reports, 3(1), 103–115.Find this resource:
Bernard, P. B., Castano, A. M., O'Leary, H., Simpson, K., Browning, M. D., & Benke, T. A. (2013). Phosphorylation of FMRP and alterations of FMRP complex underlie enhanced mLTD in adult rats triggered by early life seizures. Neurobiology of Disease, 59, 1–17.Find this resource:
Bhattacharya, A., Kaphzan, H., Alvarez-Dieppa, A. C., Murphy, J. P., Pierre, P., & Klann E. (2012). Genetic removal of p70 S6 kinase 1 corrects molecular, synaptic, and behavioral phenotypes in fragile X syndrome mice. Neuron, 76(2), 325–337.Find this resource:
Bhattacharya, A., Mamcarz, M., Mullins, C., Choudhury, A., Boyle, R. G., Smith, D. G., … Klann, E. (2016). Targeting translation control with p70 S6 kinase 1 inhibitors to reverse phenotypes in fragile x syndrome mice. Neuropsychopharmacology, 41(8), 1991–2000.Find this resource:
Bowling, H., Zhang, G., Bhattacharya, A., Pérez-Cuesta, L. M., Deinhardt, K., Hoeffer, C. A., … Chao M. V. (2014). Antipsychotics activate mTORC1-dependent translation to enhance neuronal morphological complexity. Science Signaling, 7(308), ra4.Find this resource:
Buchsbaum, R. J., Connolly, B. A., & Feig, L. A. (2003). Regulation of p70 S6 kinase by complex formation between the Rac guanine nucleotide exchange factor (Rac-GEF) Tiam1 and the scaffold spinophilin. Journal of Biological Chemistry, 278(21), 18833–18841.Find this resource:
Burnett, P. E., Blackshaw, S., Lai, M. M., Qureshi, I. A, Burnett, A. F., Sabatini, D. M., & Snyder, S. H. (1998). Neurabin is a synaptic protein linking p70 S6 kinase and the neuronal cytoskeleton. Proceedings of National Academy of Science of USA, 95(14), 8351–8356.Find this resource:
Caccamo, A., Branca, C., Talboom, J. S., Shaw, D. M., Turner, D., Ma, L., … Oddo, S. (2015). Reducing ribosomal protein s6 kinase 1 expression improves spatial memory and synaptic plasticity in a mouse model of Alzheimer's disease. Journal of Neuroscience, 35(41), 14042–14056.Find this resource:
Cammalleri, M., Lütjens, R., Berton, F., King, A. R., Simpson, C., Francesconi, W., & Sanna, P. P. (2003). Time-restricted role for dendritic activation of the mTOR-p70S6K pathway in the induction of late-phase long-term potentiation in the CA1. Proceedings of National Academy of Science of USA, 100(24), 14368–14373.Find this resource:
Chévere-Torres, I., Kaphzan, H., Bhattacharya, A., Kang, A., Maki, J. M., Gambello, M. J., … Klann, E. (2012). Metabotropic glutamate receptor-dependent long-term depression is impaired due to elevated ERK signaling in the ΔRG mouse model of tuberous sclerosis complex. Neurobiology of Disease, 45(3), 1101–1110.Find this resource:
Cholerton, B., Baker, L. D., & Craft, S. (2013). Insulin, cognition, and dementia. European Journal of Pharmacology, 719(1–3), 170–179.Find this resource:
Chou, M. M., & Blenis, J. (1996). The 70Kd S6 kinase complexes with and is activated by the Rho family G proteins Cdc42 and Rac1. Cell, 85, 573–583.Find this resource:
Costa-Mattioli, M., Sossin, W. S., Klann, E., & Sonenberg, N. (2009). Translational control of long-lasting synaptic plasticity and memory. Neuron, 61(1), 10–26.Find this resource:
Dwyer, J. M., Maldonado-Avilés, J. G., Lepack, A. E., DiLeone, R. J., & Duman, R. S. (2015). Ribosomal protein S6 kinase 1 signaling in prefrontal cortex controls depressive behavior. Proceedings of National Academy of Science of USA, 112(19), 6188–6193.Find this resource:
Fang, Y., Park, I. H., Wu, A. L., Du, G., Huang, P., Frohman, M. A., … Chen, J. (2003). PLD1 regulates mTOR signaling and mediates Cdc42 activation of S6K1. Journal of Biological Chemistry, 13(23), 2037–2044.Find this resource:
Frödin, M., & Gammeltoft, S. (1999). Role and regulation of 90 kDa ribosomal S6 kinase (RSK) in signal transduction. Molecular and Cellular Endocrinology, 151(1–2), 65–77.Find this resource:
Gafford, G. M., Parsons, R. G., & Helmstetter, F. J. (2013). Memory accuracy predicts hippocampal mTOR pathway activation following retrieval of contextual fear memory. Hippocampus, 23(9), 842–847.Find this resource:
Gavaret, J. M., Matricon, C., Pomerance, M., Jacquemin, C., Toru-Delbauffe, D., & Pierre, M. (1989). Activation of S6 kinase in astroglial cells by FGFa and FGFb. Brain Research Dev Brain Research, 45(1), 77–82.Find this resource:
Gong, X., Zhang, L., Huang, T., Lin, T. V., Miyares, L., Wen, J., Hsieh, L., & Bordey, A. (2015). Activating the translational repressor 4E-BP or reducing S6K-GSK3β activity prevents accelerated axon growth induced by hyperactive mTOR in vivo. Human Molecular Genetics, 24(20), 5746–5758.Find this resource:
Graber, T. E., McCamphill, P. K., & Sossin, W. S. (2013). A recollection of mTOR signaling in learning and memory. Learning & Memory, 20(10), 518–530.Find this resource:
Hahn, K., Miranda, M., Francis, V. A., Vendrell, J., Zorzano, A., & Teleman, A. A. (2010). PP2A regulatory subunit PP2A-B' counteracts S6K phosphorylation. Cell Metabolism, 11(5), 438–444.Find this resource:
Harrington, L. S., Findlay, G. M., Gray, A., Tolkacheva, T., Wigfield, S., Rebholz, H., … Lamb R. F. (2004). The TSC1–2 tumor suppressor controls insulin-PI3K signaling via regulation of IRS proteins. Journal of Cell Biology, 166(2), 213–223.Find this resource:
Heidenreich, K. A., & Toledo, S. P. (1989). Insulin receptors mediate growth effects in cultured fetal neurons. II. Activation of a protein kinase that phosphorylates ribosomal protein S6. Endocrinology, 125(3), 1458–1463.Find this resource:
Hoeffer, C. A., Sanchez, E., Hagerman, R. J., Mu, Y., Nguyen, D. V., Wong, H., … Tassone, F. (2012). Altered mTOR signaling and enhanced CYFIP2 expression levels in subjects with fragile X syndrome. Genes Brain Behavior, 11(3), 332–341.Find this resource:
Hong, S., Zhao, B., Lombard, D. B., Fingar, D. C., & Inoki, K. (2014). Cross-talk between sirtuin and mammalian target of rapamycin complex 1 (mTORC1) signaling in the regulation of S6 kinase 1 (S6K1) phosphorylation. Journal of Biological Chemistry, 289(19), 13132–13141. doi: 10.1074/jbc.M113.520734.Find this resource:
Huang, W., Zhu, P. J., Zhang, S., Zhou, H., Stoica, L., Galiano, M., … Costa-Mattioli, M. (2013). mTORC2 controls actin polymerization required for consolidation of long-term memory. Nature Neuroscience, 16(4), 441–448.Find this resource:
Huang, W. C., Chen, Y., & Page, D. T. (2016). Hyperconnectivity of prefrontal cortex to amygdala projections in a mouse model of macrocephaly/autism syndrome. Nature Communications, 7, 13421.Find this resource:
Huber, K. M., Kayser, M. S., & Bear, M. F. (2000). Role for rapid dendritic protein synthesis in hippocampal mGluR-dependent long-term depression. Science, 288(5469), 1254–1257.Find this resource:
Huynh, T. N., Santini, E., & Klann, E. (2014). Requirement of mammalian target of rapamycin complex 1 downstream effectors in cued fear memory reconsolidation and its persistence. Journal of Neuroscience, 34(27), 9034–9039.Find this resource:
Huynh, T. N., Santini, E., Mojica, E., Fink, A. E., Hall, B. S., Fetcho, R. N., … Klann, E. (2017). Activation of a novel p70 S6 kinase 1-dependent intracellular cascade in the basolateral nucleus of the amygdala is required for the acquisition of extinction memory. Molecular Psychiatry. doi: 10.1038/mp.2017.99.Find this resource:
Jaworski, J., & Sheng, M. (2006). The growing role of mTOR in neuronal development and plasticity. Molecular Neurobiology, 34(3), 205–219.Find this resource:
Jaworski, J., Spangler, S., Seeburg, D. P., Hoogenraad, C. C., & Sheng, M.(2005). Control of dendritic arborization by the phosphoinositide-3'-kinase-Akt-mammalian target of rapamycin pathway. Journal of Neuroscience, 25(49), 11300–11312.Find this resource:
Jefferies, H. B., Fumagalli, S., Dennis, P. B., Reinhard, C., Pearson, R. B., & Thomas, G. (1997). Rapamycin suppresses 5'TOP mRNA translation through inhibition of p70s6k. EMBO Journal, 16(12), 3693–3704.Find this resource:
Julien, L. A., Carriere, A., Moreau, J., & Roux, P. P. (2010). mTORC1-activated S6K1 phosphorylates Rictor on threonine 1135 and regulates mTORC2 signaling. Molecular & Cellular Biology, 30(4), 908–921.Find this resource:
Jung, H., Gkogkas, C. G., Sonenberg, N., & Holt, C. E. (2014). Remote control of gene function by local translation. Cell, 157(1), 26–40.Find this resource:
Kandel, E. R. (2001). The molecular biology of memory storage: a dialogue between genes and synapses. Science, 294(5544), 1030–1038.Find this resource:
Kang, H., & Schuman, E. M. (1996). A requirement for local protein synthesis in neurotrophin-induced hippocampal synaptic plasticity. Science, 273(5280), 1402–1406.Find this resource:
Kelleher, R. J. 3rd, Govindarajan, A., & Tonegawa, S. (2004). Translational regulatory mechanisms in persistent forms of synaptic plasticity. Neuron, 44(1), 59–73.Find this resource:
Keller, J. N., Dimayuga, E., Chen, Q., Thorpe, J., Gee, J., & Ding, Q. (2004). Autophagy, proteasomes, lipofuscin, and oxidative stress in the aging brain. International Journal of Biochemistry Cell Biology, 36(12), 2376–2391.Find this resource:
Kennedy, B. K., & Kaeberlein, M. (2009). Hot topics in aging research: Protein translation. Aging Cell, 8(6), 617–623.Find this resource:
Kozak, M. (1992). Regulation of translation in eukaryotic systems. Annual Review of Cell Biology, 8, 197–225.Find this resource:
Kozma, S. C., Ferrari, S., Bassand, P., Siegmann, M., Totty, N., & Thomas, G. (1990). Cloning of the mitogen-activated S6 kinase from rat liver reveals an enzyme of the second messenger subfamily. Proceedings of National Academy of Science of USA, 87(19), 7365–7369.Find this resource:
Kumari, D., Bhattacharya, A., Nadel, J., Moulton, K., Zeak, N. M., Glicksman, A., … Usdin, K. (2014). Identification of fragile X syndrome specific molecular markers in human fibroblasts: A useful model to test the efficacy of therapeutic drugs. Human Mutation, 35(12), 1485–1494.Find this resource:
Lai, K. O., Liang, Z., Fei, E., Huang, H., & Ip, N. Y. (2015). Cyclin-dependent kinase 5 (Cdk5)-dependent phosphorylation of p70 ribosomal s6 kinase 1 (S6K) is required for dendritic spine morphogenesis. Journal of Biological Chemistry, 290(23), 14637–14646.Find this resource:
Lécureuil, C., Tesseraud, S., Kara, E., Martinat, N., Sow, A., Fontaine, I., …Crépieux, P. (2005). Follicle-stimulating hormone activates p70 ribosomal protein S6 kinase by protein kinase A-mediated dephosphorylation of Thr 421/Ser 424 in primary Sertoli cells. Molecular Endocrinology, 19, 1812–1820.Find this resource:
Lehman, J. A., Calvo, V., & Gomez-Cambronero, J. (2003). Mechanism of ribosomal p70S6 kinase activation by granulocyte macrophage colony-stimulating factor in neutrophils: Cooperation of a MEK-related, THR421/SER424 kinase and a rapamycin-sensitive, m-TOR-related THR389 kinase. Journal of Biological Chemistry, 278(30), 28130–28138.Find this resource:
Lein, E. S., Hawrylycz, M. J., Ao, N., Ayres, M., Bensinger, A., Bernard, A., … Jones, A. R. (2007). Genome-wide atlas of gene expression in the adult mouse brain. Nature, 445, 168–176.Find this resource:
Liu, S., & Lu, B. (2010). Reduction of protein translation and activation of autophagy protect against PINK1 pathogenesis in Drosophila melanogaster. PLoS Genetics, 6(12), e1001237.Find this resource:
Liu, X., Müller, F., Wayne, A. S., & Pastan, I. (2016). Protein kinase inhibitor H89 enhances the activity of pseudomonas exotoxin a-based immunotoxins. Molecular Cancer & Therapeutics, 5, 1053–1062.Find this resource:
Ma, X. M., Yoon, S. O, Richardson, C. J., Jülich, K., & Blenis, J. (2008). SKAR links pre-mRNA splicing to mTOR/S6K1-mediated enhanced translation efficiency of spliced mRNAs. Cell, 133(2), 303–313.Find this resource:
Ma, T., Trinh, M. A., Wexler, A., Bourbon, C., Gatti, E., Pierre, P., Cavener, D. R., Klann, E. (2013). Suppression of eIF2α kinases alleviates Alzheimer's disease-related plasticity and memory deficits. Nature Neuroscience, 16(9), 1299–1305.Find this resource:
Magnuson, B., Ekim, B., & Fingar, D. C. (2012). Regulation and function of ribosomal protein S6 kinase (S6K) within mTOR signalling networks. Biochemical Journal, 441(1), 1–21.Find this resource:
Malenka, R. C., & Nicoll, R. A. (1999). Long-term potentiation—a decade of progress? Science, 285(5435), 1870–1874.Find this resource:
Massaad, C. A., & Klann, E. (2011). Reactive oxygen species in the regulation of synaptic plasticity and memory. Antioxidant and Redox Signal, 14(10), 2013–2054.Find this resource:
McCamphill, P. K., Ferguson, L., & Sossin, W. S. (2017). A decrease in eukaryotic elongation factor 2 phosphorylation is required for local translation of sensorin and long-term facilitation in Aplysia. Journal of Neurochemistry, 142(2), 246–259.Find this resource:
Michel, K., Zhao, T., Karl, M., Lewis, K., & Fyffe-Maricich, S. L. (2015). Translational control of myelin basic protein expression by ERK2 MAP kinase regulates timely remyelination in the adult brain. Journal of Neuroscience, 35(20), 7850–7865.Find this resource:
Nakamura, J. L., Garcia, E., & Pieper, R. O. (2008). S6K1 plays a key role in glial transformation. Cancer Research 68(16), 6516–6523.Find this resource:
Narayanan, U., Nalavadi, V., Nakamoto, M., Thomas, G., Ceman, S., Bassell, G. J., & Warren S. T. (2008). S6K1 phosphorylates and regulates fragile X mental retardation protein (FMRP) with the neuronal protein synthesis-dependent mammalian target of rapamycin (mTOR) signaling cascade. Journal of Biological Chemistry, 283(27), 18478–18482.Find this resource:
Nardella, C., Lunardi, A., Fedele, G., Clohessy, J. G., Alimonti, A., Kozma, S. C., … Pandolfi, P. P. (2011). Differential expression of S6K2 dictates tissue-specific requirement for S6K1 in mediating aberrant mTORC1 signaling and tumorigenesis. Cancer Research, 71(10), 3669–3675.Find this resource:
Novak-Hofer, I., & Thomas, G. (1984). An activated S6 kinase in extracts from serum- and epidermal growth factor-stimulated Swiss 3T3 cells. Journal of Biological Chemistry, 259(9), 5995–6000.Find this resource:
Pardo, O. E., & Seckl, M. J. (2013). S6K2: The neglected S6 kinase family member. Frontiers of Oncology, 3, 191.Find this resource:
Parker, W. E., Orlova, K. A., Parker, W. H., Birnbaum, J. F., Krymskaya, V. P., Goncharov, D. A., … Crino, P. B. (2013). Rapamycin prevents seizures after depletion of STRADA in a rare neurodevelopmental disorder. Science Translational Medicine, 5(182), 182ra53.Find this resource:
Pearce, L. R., Alton, G. R., Richter, D. T., Kath, J. C., Lingardo, L., Chapman, J., … Alessi D. R. (2010b). Characterization of PF-4708671, a novel and highly specific inhibitor of p70 ribosomal S6 kinase (S6K1). Biochemical Journal, 431(2), 245–255.Find this resource:
Pearce, L. R., Komander, D., Alessi, D. R. (2010). The nuts and bolts of AGC protein kinases. Nature Reviews Molecular Cell Biology, 11, 9–22. doi: 10.1038/nrm2822.Find this resource:
Pende, M., Um, S. H., Mieulet, V., Sticker, M., Goss, V. L., Mestan, J., … Thomas, G. (2004). S6K1(-/-)/S6K2(-/-) mice exhibit perinatal lethality and rapamycin-sensitive 5'-terminal oligopyrimidine mRNA translation and reveal a mitogen-activated protein kinase-dependent S6 kinase pathway. Molecular & Cellular Biology, 24(8), 3112–3124.Find this resource:
Pierre, M., Toru-Delbauffe, D., Gavaret, J. M., Pomerance, M., & Jacquemin, C. (1986). Activation of S6 kinase activity in astrocytes by insulin, somatomedin C and TPA. FEBS Letters, 206(1), 162–166.Find this resource:
Piper, M., Anderson, R., Dwivedy, A., Weinl, C., van Horck, F., Leung, K. M., … Holt, C. (2006). Signaling mechanisms underlying Slit2-induced collapse of Xenopus retinal growth cones. Neuron, 49(2), 215–228.Find this resource:
Pirbhoy, P. S., Farris, S., & Steward, O. (2017). Synaptically driven phosphorylation of ribosomal protein S6 is differentially regulated at active synapses versus dendrites and cell bodies by MAPK and PI3K/mTOR signaling pathways. Learning & Memory, 24(8), 341–357.Find this resource:
Qin, J., Rajaratnam, R., Feng, L., Salami, J., Barber-Rotenberg, J. S., Domsic, J., … Marmorstein, R. (2015). Development of organometallic S6K1 inhibitors. Journal of Medicinal Chemistry, 58(1), 305–314.Find this resource:
Roberts, S., & Morelos, B. S. (1982). Inhibition of cerebral protein kinase activity and cyclic AMP-dependent ribosomal-protein phosphorylation in experimental hyperphenylalaninaemia. Biochemical Journal, 202(2), 343–351.Find this resource:
Romanelli, A., Martin, K. A., Toker, A., & Blenis, J. (1999). p70 S6K is regulated by protein kinase C eta and participants in a phosphoinositide 3 kinase-regulated signaling complex. Molecular & Cellular Biology, 19, 2921–2928.Find this resource:
Savinska, L. O., Lyzogubov, V. V., Usenko, V. S., Ovcharenko, G. V., Gorbenko, O. N., Rodnin, M. V., … Filonenko, V. V. (2004). Immunohistochemical analysis of S6K1 and S6K2 expression in human breast tumors. Eksp Onkol, 26(1), 24–30.Find this resource:
Schimanski, L. A., & Barnes, C. A. (2010). Neural protein synthesis during aging: Effects on plasticity and memory. Frontiers in Aging Neuroscience, 2, article 26. doi: 10.3389/fnagi.2010.00026. eCollection 2010.Find this resource:
Selman, C., Tullet, J. M., Wieser, D., Irvine, E., Lingard, S. J., Choudhury, A. I., … Withers, D. J. (2009). Ribosomal protein S6 kinase 1 signaling regulates mammalian life span. Science, 326(5949), 140–144.Find this resource:
Sethna, F., Zhang, M., Kaphzan, H., Klann, E., Autio, D., Cox, C. L., & Wang, H. (2016). Calmodulin activity regulates group I metabotropic glutamate receptor-mediated signal transduction and synaptic depression. Journal of Neuroscience Research, 94(5), 401–408.Find this resource:
Sharma, A., Hoeffer, C. A., Takayasu, Y., Miyawaki, T., McBride, S. M., Klann, E., & Zukin, R. S. (2010). Dysregulation of mTOR signaling in fragile X syndrome. Journal of Neuroscience, 30(2), 694–702.Find this resource:
Shima, H., Pende, M., Chen, Y., Fumagalli, S., Thomas, G., & Kozma, S. C. (1998). Disruption of the p70(s6k)/p85(s6k) gene reveals a small mouse phenotype and a new functional S6 kinase. EMBO Journal, 17(22), 6649–6659.Find this resource:
Shin, S., Wolgamott, L., Yub, Y., Blenis, J., & Yoona, S. O. (2011). Glycogen synthase kinase (GSK)-3 promotes p70 ribosomal protein S6 kinase (p70S6K) activity and cell proliferation. Proceedings of National Academy of Sciences of USA, 108(47), E1204–E1213.Find this resource:
Slegtenhorst-Eegdeman, K. E., de Rooij, D. G., Verhoef-Post, M., van de Kant, H. J., Bakker, C. E., Oostra, B. A., … Themmen, A. P. (1998). Macroorchidism in FMR1 knockout mice is caused by increased Sertoli cell proliferation during testicular development. Endocrinology, 139(1), 156–162.Find this resource:
Smith, M. A., Katsouri, L., Irvine, E. E., Hankir, M. K., Pedroni, S. M., Voshol, P. J., … Withers, D. J. (2015). Ribosomal S6K1 in POMC and AgRP neurons regulates glucose homeostasis but not feeding behavior in mice. Cell Reports, 11(3), 335–343.Find this resource:
Sun, J., Liu, Y., Tran, J., O'Neal, P., Baudry, M., & Bi, X. (2016). mTORC1-S6K1 inhibition or mTORC2 activation improves hippocampal synaptic plasticity and learning in Angelman syndrome mice. Cell Molecular Life Science, 73(22), 4303–4314.Find this resource:
Sunami, T., Byrne, N., Diehl, R. E., Funabashi, K., Hall, D. L., Ikuta, M., … Sharma, S. (2010). Structural basis of human p70 ribosomal S6 kinase-1 regulation by activation loop phosphorylation. Journal of Biological Chemistry, 285(7), 4587–4594.Find this resource:
Tavares, M. R., Pavan, I. C., Amaral, C. L., Meneguello, L., Luchessi, A. D., & Simabuco, F. M. (2015). The S6K protein family in health and disease. Life Science, 131, 1–10.Find this resource:
Tolcher, A., Goldman, J., Patnaik, A., Papadopoulos, K. P., Westwood, P., Kelly, C. S., … Rosen, L. S. (2014). A phase I trial of LY2584702 tosylate, a p70 S6 kinase inhibitor, in patients with advanced solid tumours. European Journal of Cancer, 50(5), 867–875.Find this resource:
Um, S. H., D’Alessio, D., Thomas, G. (2004). Nutrient overload, insulin resistance, and the ribosomal protein S6 Kinase 1, S6K1. Cell Metabolism, 6, 393–402.Find this resource:
Verma, P., Chierzi, S., Codd, A. M., Campbell, D. S., Meyer, R. L., Holt, C. E., & Fawcett, J. W. (2005). Axonal protein synthesis and degradation are necessary for efficient growth cone regeneration. Journal of Neuroscience, 25(2), 331–342.Find this resource:
Wang, M. L., Panasyuk, G., Gwalter, J., Nemazanyy, I., Fenton, T., Filonenko, V., & Gout, I. (2008). Regulation of ribosomal protein S6 kinases by ubiquitination. Biochemical Biophysical Research Communications, 369, 382–387.Find this resource:
Wettenhall, R. E., Chesterman, C. N., Walker, T., & Morgan, F. J. (1983). Phosphorylation sites for ribosomal S6 protein kinases in mouse 3T3 fibroblasts stimulated with platelet-derived growth factor. FEBS Letters, 162(1), 171–176.Find this resource:
Winter, J. N., Jefferson, L. S., & Kimball, S. R. (2011). ERK and Akt signaling pathways function through parallel mechanisms to promote mTORC1 signaling. American Journal of Physiology, 300(5), C1172–C1180.Find this resource:
Wu, G. Y., Deisseroth, K., & Tsien, R. W. (2001). Spaced stimuli stabilize MAPK pathway activation and its effects on dendritic morphology. Nature Neuroscience, 4(2), 151–158.Find this resource:
Xi, Y., Niu, J., Shen, Y., Li, D., Peng, X., & Wu, X. (2016). AT13148, a first-in-class multi-AGC kinase inhibitor, potently inhibits gastric cancer cells both in vitro and in vivo. Biochemical Biophysical Research Communications, 478(1), 330–336.Find this resource:
Xu, Y., Liu, C., Chen, S., Ye, Y., Guo, M., Ren, Q., … Chen, L. (2014). Activation of AMPK and inactivation of Akt result in suppression of mTOR-mediated S6K1 and 4E-BP1 pathways leading to neuronal cell death in in vitro models of Parkinson's disease. Cell Signaling, 26(8), 1680–1689.Find this resource:
Yang, L., Miao, L., Liang, F., Huang, H., Teng, X., Li, S., … Hu, Y. (2014). The mTORC1 effectors S6K1 and 4E-BP play different roles in CNS axon regeneration. Nature Communications, 5, 5416. doi: 10.1038/ncomms6416.Find this resource:
Zare, N., Motamedi, F., Digaleh, H., Khodagholi, F., & Maghsoudi, N. (2015). Collaboration of geldanamycin-activated P70S6K and Hsp70 against beta-amyloid-induced hippocampal apoptosis: An approach to long-term memory and learning. Cell Stress Chaperones, 20(2), 309–319.Find this resource:
Zhou, X., Lin, D. S, Zheng, F., Sutton, M. A., & Wang, H. (2010). Intracellular calcium and calmodulin link brain-derived neurotrophic factor to p70S6 kinase phosphorylation and dendritic protein synthesis. Journal of Neuroscience Research, 88(7), 1420–1432.Find this resource: