Specific Silencing of the REST Target Genes in Insulin-Secreting Cells Uncovers Their Participation in Beta Cell Survival

Abstract

The absence of the transcriptional repressor RE-1 Silencing Transcription Factor (REST) in insulin-secreting beta cells is a major cue for the specific expression of a large number of genes. These REST target genes were largely ascribed to a function of neurotransmission in a neuronal context, whereas their role in pancreatic beta cells has been poorly explored. To identify their functional significance, we have generated transgenic mice expressing REST in beta cells (RIP-REST mice), and previously discovered that REST target genes are essential to insulin exocytosis. Herein we characterized a novel line of RIP-REST mice featuring diabetes. In diabetic RIP-REST mice, high levels of REST were associated with postnatal beta cell apoptosis, which resulted in gradual beta cell loss and sustained hyperglycemia in adults. Moreover, adenoviral REST transduction in INS-1E cells led to increased cell death under control conditions, and sensitized cells to death induced by cytokines. Screening for REST target genes identified several anti-apoptotic genes bearing the binding motif RE-1 that were downregulated upon REST expression in INS-1E cells, including Gjd2, Mapk8ip1, Irs2, Ptprn, and Cdk5r2. Decreased levels of Cdk5r2 in beta cells of RIP-REST mice further confirmed that it is controlled by REST, in vivo. Using siRNA-mediated knock-down in INS-1E cells, we showed that Cdk5r2 protects beta cells against cytokines and palmitate-induced apoptosis. Together, these data document that a set of REST target genes, including Cdk5r2, is important for beta cell survival.

REST represses a subset of the pancreatic endocrine differentiation program

“…This led to a decrease in the number of endocrine-committed progenitors by E14.5 and ultimately reduced the numbers of glucagon + and insulin + cells in E18.5 pancreas. Eventually, adult mice developed diabetes, an observation that we already made in RIP-REST mice overexpressing REST specifically in beta cells (Martin et al, 2012). Diabetes in Pdx1-tTA/TetO-REST mice is most likely caused by an impaired function, rather than being caused by the moderate reduction in insulin-producing cells.…”

“…By analyzing a new RIP-REST founder line, we further showed that higher levels of REST trans-gene expression drastically compromised beta cell survival, leading to diabetes (Martin et al, 2012). Here, we report the same impairment in glucose homeostasis with another transgenic system driving REST expression in adult mature beta cells (under the control of Pdx1 regulatory region) (Fig.…”

“…Taking into account that pancreatic beta cells share many similarities with neurons, including the fact that REST is also disallowed in these endocrine cells (Atouf et al, 1997; Martin et al, 2008, 2012; Thiel and Schuit, 2008), we initiated this study to evaluate the role of REST in endocrine differentiation. We report that REST is expressed in pancreas progenitors and that its expression decreases in endocrine progenitors and endocrine cells.…”

“…Western blots were performed as previously described (Martin et al, 2012). Specific protein levels were revealed with polyclonal rabbit antibodies against human REST (Martin et al, 2003), or with polyclonal rabbit antibodies against murine REST (07-579, Merck Millipore).…”

“…Indeed, we have demonstrated that Rest ectopic expression in pancreatic insulin-producing cells impairs their function and survival by specifically down-regulating the expression of important exocytotic members as well as pro-survival genes (Martin et al, 2008, 2012). As specified for a subset of other genes (Pullen et al, 2010; Quintens et al, 2008), REST is thus “disallowed” in beta cells, as it is in neurons (Atouf et al, 1997).…”