90-kDa ribosomal protein S6 kinase 5; Mitogen- and stress-activated protein kinase 1; MSPK1; Nuclear mitogen- and stress-activated protein kinase-1; Ribosomal protein S6 kinase, 90kD, polypeptide 5; Ribosomal protein S6 kinase, polypeptide 5; RLPK; RLSK; Rps6ka5; Rsk-like (RSKL); Rsk-like protein kinase (RLPK)
Mitogen- and stress-activated protein kinase (Msk) 1 and Msk2 are nuclear serine/threonine kinases that are widely expressed in tissues including heart, brain, placenta, lung, liver, kidney, and pancreas, with the highest levels observed in brain, muscle, and placenta (Deak et al. 1998). They belong to a group of structurally related kinases called the AGC kinase (cAMP-dependent, cGMP-dependent, and protein kinase C) family. This group includes p70 ribosomal S6 kinase (S6 K), protein kinase B (PKB; also called Akt), protein kinase C-related kinase (Prk), p90 ribosomal S6 kinase (Rsk), and serum- and glucocorticoid-inducible kinase (Sgk) (Pearce et al. 2010).
Msk1 was identified through its homology to the N-terminal Rsk domain (Deak et al. 1998). Similarly to Rsk, Msk1 contains two kinase domains, connected with a linker region, in a single polypeptide (Deak et al. 1998). The N-terminal kinase domain is related to the AGC family of protein kinases (containing PKA, PKG, and PKC families), whereas the C-terminal kinase domain is related to the calmodulin-activated protein kinase family (Pearce et al. 2010). The C-terminal tail fulfills at least three functions: it contains a nuclear localization sequence and a mitogen-activated protein (MAP) kinase docking sequence, and can also act as an auto-inhibitory sequence (McCoy et al. 2007). Mutation of either the C- or N-terminal kinase domains in Msk1 is sufficient to block the phosphorylation of its substrates (Deak et al. 1998).
Because it can be activated through two different pathways (extracellular signal-regulated kinases 1 and 2 (ERK1/2) and p38 MAP kinase), Msk1 is able to integrate signals from growth factors, pro-inflammatory cytokines (tumor necrosis factor-α ( TNF-α)) and cellular stress (ultraviolet (UV)-irradiation, hydrogen peroxide). Msk1 is predominantly found in the nucleus and it contains a nuclear localization sequence in the C-terminal region, but a portion is localized to the cytosol of mouse fibroblasts and HEK 293 cells (Deak et al. 1998).
Regulation of Msk1 Activity
In cells, Msk1 is activated via a complex series of phosphorylation and autophosphorylation reactions downstream of ERK1/2 or p38 α-MAP kinase. Many stimuli have been shown to activate Msk1 in cell cultures, including UV irradiation, anisomycin, nerve growth factor (NGF), TNF-α, interleukin (IL)-1, lysophosphatidic acid, endothelin-1, α1-adrenergic stimulation, arsenic trioxide, and oxidative stress. In addition, exercise has been shown to induce Msk1 activation in animal skeletal muscle and light activates Msk1 in the suprachiasmatic nucleus in the brain (extensively reviewed in Arthur 2008; Vermeulen et al. 2009; Lazou and Markou 2010; Duda and Frodin 2014).
cAMP-responsive element (CRE) binding protein ( CREB) was the first Msk1 substrate to be identified (Deak et al. 1998). Since then, several studies using a variety of experimental settings, in vitro and in vivo, have shown phosphorylation of CREB (S133) and activation of its transcriptional activity by Msk1 in response to growth factors, phorbol esters, pro-inflammatory cytokines, G protein-coupled receptor agonists, or cellular stresses (for review see Vermeulen et al. 2009; Lazou and Markou 2010; Naqvi and Arthur 2014). Phosphorylation of CREB by Msk1 leads to transcription of several immediate early genes that have CRE in their promoter including c-fos, junB, Nur77, Mkp1, MUC5AC, Cox-2, IL-1β, ANF, and c-jun. Msk1 also phosphorylates activating transcription factor 1 (ATF1) and activating transcription factor 2 (ATF2).
Another transcription factor that is a substrate for Msk1 is the nuclear factor (NF)-κB (Vermeulen et al. 2003; Vermeulen et al. 2009; Reber and Haegeman 2014). In unstimulated cells, NF-κB is found in the cytoplasm where it is kept inactive by binding to the inhibitory protein IκB. Stimulation with various agonists leads to the release of NF-κB and its translocation to the nucleus where further posttranslational modifications regulate its activity. Msk1 phosphorylates the p65/RelA subunit of NF-κB at S276 in response to TNF-α in fibroblasts, and in response to oxidative stress in skeletal myoblasts.
STAT3 (signal transducer and activator of transcription 3) and ER81 (E26 transformation-specific (ETS)-related protein 81; also called Ets transcript variant 1, Etv1) are also targeted by Msk1. Phosphorylation of STAT3 at S727 is mediated through Msk1 in JB6 cells in response to UVA irradiation and in erythroid cells in response to erythropoietin (Wierenga et al. 2003). The activity of the transcription factor ER81, which is involved in oncogenesis and breast tumor formation, is regulated by Msk1 via direct phosphorylation at S191 and S216. However, it should be noted that ER81 phosphorylation has not been validated in vivo.
Increasing evidence indicates an important role of Msk1 in remodeling chromatin structure and the epigenetic regulation of gene expression. Using H89 as an inhibitor of Msk1 activity, it has been shown that Msk1 phosphorylates histone H3 at S10 and S28 in response to phorbol esters (12- O-tetradecanoylphorbol-13-acetate (TPA)), epidermal growth factor (EGF), or anisomycin (Thomson et al. 1999). Studies using various types of cells from genetically modified animals provided further evidence for the role of Msk1 in H3 phosphorylation (for a review see Brami-Cherrier et al. 2009; Lazou and Markou 2010; Drobic et al. 2014). However, the mechanism by which Msk1 is recruited to chromatin and directed to phosphorylated S10 or S28 in H3 is not known. It is not likely that Msk1 is preloaded onto immediate early gene promoters before induction, as it has been demonstrated that Msk1 is associated with the IL-6 promoter after cells are stimulated (Vermeulen et al. 2003). Furthermore, E-26-like protein 1 (Elk-1) is required for the recruitment of ERK and Msk1 to the promoter of immediate early genes c-fos and egr-1 (Zhang et al. 2008). On the other hand, using an in vitro system, it was recently shown that the recruitment of Msk1 and the subsequent phosphorylation of H3 on the c-fos chromatin requires CREB and, to a lesser extent, ATF1. Phosphorylation of CREB at S133 is essential for this process (Shimada et al. 2010).
In addition to H3, Msk1 also phosphorylates other chromatin-associated proteins such as nonhistone chromosomal protein HMGN1 (also called HMG-14) as well as histone H2A (Thomson et al. 1999; Brami-Cherrier et al. 2009).
Msk1 has been implicated in the regulation of translational control by phosphorylating eukaryotic translation initiation factor 4E–BP1. 4E–BP1 binds to eIF4E in resting cells preventing formation of eIF-4 F complex, which is essential for cap-dependent initiation of translation. UVB irradiation promotes phosphorylation of 4E–BP1 by Msk1 leading to the dissociation from eIF4E and the release of translational block after UVB irradiation (Liu et al. 2002).
Cell Death-Related Proteins
The Bcl-2 family member Bad has been also reported as Msk1 substrate. Bad is a proapoptotic protein and its phosphorylation promotes cell survival in many cell types. UVB irradiation promotes phosphorylation of Bad at S112 through Jnk1, Rsk2, and Msk1 (She et al. 2002). Furthermore, Msk1 mediates phosphorylation of neuronal Bad following Ca2+ influx (Clark et al. 2007).
Msk1 phosphorylates cPLA2 at S727 in vitro and is also implicated in the translocation of cPLA2 to the membrane (Lazou and Markou 2010). However, it remains to be investigated whether cPLA2 is a real target of MSK1 in vivo. In mouse epidermal JB6 cells, the expression of Msk1 C-terminal kinase-dead mutant inhibited UVB-induced phosphorylation and activation of Akt, implicating Msk1 as the kinase responsible (Nomura et al. 2001). Another protein, E3 ubiquitin ligase Tripartite motif containing 7 (TRIM7) was recently reported to be directly phosphorylated and activated by MSK1 in response to Ras–Raf–MEK–ERK pathway signaling (Chakraborty et al. 2015). Furthermore, MSK1 phosphorylates β-catenin and regulates its nuclear translocation and transcriptional activity (Wu et al. 2016).
Physiological Roles of Msk1
Inflammation and Immunity
As mentioned above, Msk1 contributes to the transcriptional activation of NF-κB and CREB, two transcription factors with important roles in the regulation of inflammatory genes (Arthur 2008; Vermeulen et al. 2009). Msk1 plays a critical role in limiting inflammation in innate immunity inducing the transcription of the MAP kinase phosphatase Dusp1 and the anti-inflammatory cytokines IL-10 and IL-1ra through TLR (toll-like receptor) signaling (Ananieva et al. 2008; Elcombe et al. 2013). Furthermore, Msk1 controls the transcription of COX-2 in response to TLR signaling. The role of Msk1 in the adaptive immune responses is less characterized. (Arthur and Elcombe 2014; Reyskens and Arthur 2016).
There are controversial reports regarding the role of Msk1 in cell death. Activation of Msk1 by glutamatergic neurotransmission results in enhanced neuronal injury (Hughes et al. 2003). Furthermore, in human lymphoma B cells, the sequential activation of p38-MAP kinase and Msk1 leads to the Mn2+-dependent activation of caspase-8 and cell death (El Mchichi et al. 2007). Although, expression of a dominant-negative mutant of Msk1 in these cells inhibits caspase-8 activation, the underlying mechanisms are not yet known. On the other hand, increasing evidence suggests that Msk1 represents a prosurvival pathway. Decreased expression of Msk1 enhances arsenic trioxide-induced apoptosis of leukemic cells in vitro and in vivo (Kannan-Thulasiraman et al. 2006). Furthermore, Msk1 knockdown reduces Bad phosphorylation and enhances Noxa and Bim expression, leading to enhanced TGFβ-induced caspase-3 activity and cell death (van der Heide et al. 2011). In agreement with this latter study, Msk1 promotes cell survival by phosphorylating Bad in mouse epidermal cells (She et al. 2002). In addition, lack of MSK1/2 enhances the susceptibility of erythrocytes to undergo suicidal erythrocyte death or eryptosis following pathophysiological cell stressors (Lang et al. 2015). Recently, it was shown that activation of Msk1 in cardiac myocytes increases resistance against oxidative stress-induced cell death (Mellidis et al. 2014). Based on the above reports, the role of Msk1 in the regulation of cell death pathways may be dependent on cell-type specific factors that are as yet undefined.
Cell Growth and Carcinogenesis
Msk1 activity has been implicated in cellular transformation. Using Msk1 dominant-negative mutants or small interfering RNA (siRNA) against Msk1, it was shown that Msk1 is required for TPA-induced or EGF-induced cellular transformation of JB6 Cl41 cells. This effect is mediated by phosphorylation of histone H3 at S10 as well as AP-1 activation (Kim et al. 2008). Msk1 is also a positive regulator of epithelial cell proliferation (Schiller et al. 2006). Although it has been reported that in a murine system of chemical carcinogenesis, tumor development was significantly reduced in Msk1/Msk2 knockout mice compared with wild-type ones (Chang et al. 2011), the role of Msk1 in cancer development is not yet fully clarified. Furthermore, in another cell system, pharmacological inhibition of Msk1 inhibits hypertrophic cell growth of cardiac myocytes (Markou et al. 2009).
Several lines of evidence support the notion that Msk1 plays crucial roles in several fundamental processes in the mammalian CNS, ranging from neuronal plasticity in memory formation to behavioral responses to drugs of abuse (Frenguelli and Correa 2014). It has been shown that Msk1 contributes to chromatin remodeling in striatal neurons and hippocampus resulting in long-term synaptic plasticity and memory formation (Brami-Cherrier et al. 2007). Furthermore, using Msk1-knockout mice, it was demonstrated that Msk1 plays an important role in cocaine-induced transcriptional events and behavioral alterations. In these mice, phosphorylation of CREB and histone H3 as well as induction of c- fos and dynorphin in response to cocaine was prevented. In addition, a selective impairment of locomotor sensitization was observed ((Brami-Cherrier et al. 2005, Brami-Cherrier et al. 2009). Several reports have also suggested roles for Msk1 in neurodegenerative diseases. Msk1 deficiency was shown to be responsible for the transcriptional dysregulation and striatal neurodegeneration in a mouse model of Huntington’s disease, whereas decreased MSK1 expression was observed in the caudate nucleus from the striatum of Huntington’s patients (Roze et al. 2008; Brami-Cherrier et al. 2009; Martin et al. 2011).
Msk1 is a serine/threonine kinase that is activated in response to both growth factor and cellular stress stimuli. Msk1 belongs to a family of protein kinases that contain two protein kinase domains: an N-terminal kinase domain related to the AGC kinase family, and a C-terminal kinase domain related to the CaMK family, and it is activated via a complex series of phosphorylation and autophosphorylation reactions. Msk1 is predominantly located in the nucleus and has been shown to modulate gene expression by phosphorylating various substrates, including transcription factors and chromatin-associated proteins. Among the substrates that have been identified are CREB, ATF1, STAT3, the p65/RelA subunit of NF-κB and the immediate early genes c-fos, junB, c-jun, and Nurr77. In addition to transcription factors, Msk1 has been shown to phosphorylate the nucleosomal proteins histone H3 and nonhistone chromosomal protein HMG-14, the E3 ubiquitin ligase Trim7 as well as the eukaryotic translation initiation factor 4E–BP1 and the proapoptotic protein Bad. It seems that Msk1 plays a role in integrating the effects of diverse extracellular signals and it remains to be determined how it affects its downstream targets leading to the observed signal- and cell-type-dependent specificity. The full panoply of Msk1 physiological roles has not been elucidated yet, but substantial evidence implicates Msk1 in the regulation of cytokine production and the inflammatory response, as well as neuronal synaptic plasticity. In addition, recent work suggests that Msk1 can also be functionally involved in tumor initiation, cell growth, and cell death regulation. This evidence along with the growing number of newly identified Msk1 substrates makes this kinase a suitable target for therapeutic manipulation of several diseases.
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