C-terminal domain small phosphatase 1 and MAP kinase reciprocally control REST stability and neuronal differentiation

Abstract

The repressor element 1 (RE1) silencing transcription factor (REST) in stem cells represses hundreds of genes essential to neuronal function. During neurogenesis, REST is degraded in neural progenitors to promote subsequent elaboration of a mature neuronal phenotype. Prior studies indicate that part of the degradation mechanism involves phosphorylation of two sites in the C terminus of REST that require activity of beta-transducin repeat containing E3 ubiquitin protein ligase, βTrCP. We identify a proline-directed phosphorylation motif, at serines 861/864 upstream of these sites, which is a substrate for the peptidylprolyl cis/trans isomerase, Pin1, as well as the ERK1/2 kinases. Mutation at S861/864 stabilizes REST, as does inhibition of Pin1 activity. Interestingly, we find that C-terminal domain small phosphatase 1 (CTDSP1), which is recruited by REST to neuronal genes, is present in REST immunocomplexes, dephosphorylates S861/864, and stabilizes REST. Expression of a REST peptide containing S861/864 in neural progenitors inhibits terminal neuronal differentiation. Together with previous work indicating that both REST and CTDSP1 are expressed to high levels in stem cells and down-regulated during neurogenesis, our results suggest that CTDSP1 activity stabilizes REST in stem cells and that ERK-dependent phosphorylation combined with Pin1 activity promotes REST degradation in neural progenitors.T he repressor element 1 (RE1) silencing transcription factor (REST) is a transcriptional repressor that suppresses neuronal gene expression in nonneural cells, such as fibroblasts, as well as in neural progenitors (1-3). Its targets represent genes required for the terminally differentiated neuronal cell phenotype, including genes encoding voltage and ligand-dependent ion channels, receptors, growth factors, and axonal-guidance proteins (4-7). Thus, during neurogenesis, REST is progressively down-regulated to allow elaboration of the mature neuronal phenotype (3). Nonetheless, precisely how REST itself is regulated still remains an open question. Relatively little is known about either its transcriptional or posttranscriptional regulation (3,8,9). In contrast, several studies have focused on posttranslational regulation of REST (3, 10, 11), but the identity of the signaling molecules involved has received little attention.In neural progenitors and human embryonic kidney (HEK) cells, rapid REST turnover is mediated by targeting to a proteasomal pathway (3, 10, 11). REST degradation during neuronal differentiation in culture requires interaction with beta-transducin repeat containing E3 ubiquitin protein ligase (βTrCP) for targeting to the proteasome (11). βTrCP was also required for cell-cycle-dependent degradation of REST in HEK cells (10). Two adjacent phosphorylated peptides in the C-terminal domain of REST were identified as βTrCP substrates in these studies, and function as degrons. One kinase responsible for the phosphorylation and degron activity in hippocampus is casein kinase 1, CK1 (12), but whether it function...

REST, a master transcriptional regulator in neurodegenerative disease

“…CK1 phosphorylates REST at serine residues within its degron motifs, enabling the E3 ligase β-TrCP to recognize and bind REST (the substrate) through phospho-degron motifs [15•]. A recent study by Mandel and colleagues identified a proline-rich sequence upstream of the phospho-degron motifs which, when phosphorylated by ERK1/2, facilitates loss of REST at the end stage of neural differentiation [60]. …”

Dephosphorylating eukaryotic RNA polymerase II

“…An additional mode of action has been recently proposed for Scps role in gene silencing. REST degradation and association with REST-complex cofactors is regulated by phosphorylation at S861 and S864 [130]. Phosphorylation of these residues increases REST affinity to E3 ubiquitin ligase SCF β-TRCP and reduces it's affinity to CoREST [130], a protein that binds the C-terminus of REST and helps recruit additional silencing factors [131].…”

“…REST degradation and association with REST-complex cofactors is regulated by phosphorylation at S861 and S864 [130]. Phosphorylation of these residues increases REST affinity to E3 ubiquitin ligase SCF β-TRCP and reduces it's affinity to CoREST [130], a protein that binds the C-terminus of REST and helps recruit additional silencing factors [131]. Scps directly dephosphorylate REST at S861/S864 both in vitro and in vivo , and can abolish these negative effects on REST function [130].…”

“…Phosphorylation of these residues increases REST affinity to E3 ubiquitin ligase SCF β-TRCP and reduces it's affinity to CoREST [130], a protein that binds the C-terminus of REST and helps recruit additional silencing factors [131]. Scps directly dephosphorylate REST at S861/S864 both in vitro and in vivo , and can abolish these negative effects on REST function [130]. By preventing ubiquitin-mediated degradation of REST and promoting its assembly with CoREST Scps stabilize the silencing complex and prevent neuronal gene expression [130].…”

“…Scps directly dephosphorylate REST at S861/S864 both in vitro and in vivo , and can abolish these negative effects on REST function [130]. By preventing ubiquitin-mediated degradation of REST and promoting its assembly with CoREST Scps stabilize the silencing complex and prevent neuronal gene expression [130]. These two models are not mutually exclusive and may coexist.…”