Regulation of neuronal traits by a novel transcriptional complex.

The transcriptional repressor, REST, helps restrict neuronal traits to neurons by blocking their expression in nonneuronal cells. To examine the repercussions of REST expression in neurons, we generated a neuronal cell line that expresses REST conditionally. REST expression inhibited differentiation by nerve growth factor, suppressing both sodium current and neurite growth. A novel corepressor complex, CoREST/HDAC2, was shown to be required for REST repression. In the presence of REST, the CoREST/HDAC2 complex occupied the native Nav1.2 sodium channel gene in chromatin. In neuronal cells that lack REST and express sodium channels, the corepressor complex was not present on the gene. Collectively, these studies define a novel HDAC complex that is recruited by the C-terminal repressor domain of REST to actively repress genes essential to the neuronal phenotype.

The co-repressor mSin3A is a functional component of the REST-CoREST repressor complex.

The repressor REST/NRSF restricts expression of a large set of genes to neurons by suppressing their expression in non-neural tissues. We find that REST repression involves two distinct repressor proteins. One of these, CoREST, interacts with the COOH-terminal repressor domain of REST (Andres, M. E., Burger, C., Peral-Rubio, M. J., Battaglioli, E., Anderson, M. E., Grimes, J., Dallmanm J., Ballas, N. , and Mandel, G. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 9873-9878). Here we show that the co-repressor mSin3A also interacts with REST. The REST-mSin3A association involves the NH(2)-terminal repressor domain of REST and the paired amphipathic helix 2 domain of mSin3A. REST forms complexes with endogenous mSin3A in mammalian cells, and both mSin3A and CoREST interact with REST in intact mammalian cells. REST repression is blocked in yeast lacking Sin3 and rescued in its presence. In mammalian cells, repression by REST is reduced when binding to mSin3A is inhibited. In mouse embryos, the distribution of mSin3A and REST transcripts is largely coincident. The pattern of CoREST gene expression is more restricted, suggesting that mSin3A is required constitutively for REST repression, whereas CoREST is recruited for more specialized repressor functions.

Expression of neuronal traits in pancreatic beta cells. Implication of neuron-restrictive silencing factor/repressor element silencing transcription factor, a neuron-restrictive silencer.

Pancreatic beta cells (insulin-producing cells) and neuronal cells share a large number of similarities. Here, we investigate whether the same mechanisms could control the expression of neuronal genes in both neurons and insulin-producing cells. For that purpose, we tested the role of the transcriptional repressor neuron-restrictive silencing factor/repressor element silencing transciption factor (NRSF/REST) in the expression of a battery of neuronal genes in insulin-producing cells. NRSF/REST is a negative regulator of the neuronal fate. It is known to silence neuronal-specific genes in non-neuronal cells. We demonstrate that, as in the case of the neuronal pheochromocytoma cell line PC12, mRNA coding for NRSF/REST is absent from the insulinoma cell line INS-1 and from three other insulin- and glucagon-producing cell lines. NRSF/REST activity is also absent from insulin-producing cell lines. Transient expression of REST in insulin-producing cell lines is sufficient to silence a reporter gene containing a NRSF/REST binding site, demonstrating the role of NRSF/REST in the expression of neuronal markers in insulin-producing cells. Finally, by searching for the expression of NRSF/REST-regulated genes in insulin-producing cells, we increased the list of the genes expressed in both neurons and insulin-producing cells.

Dexamethasone regulates the expression of neuronal properties of a rat insulinoma cell line.

Insulin producing cells of the pancreas (beta cells) and neuronal cells share a large number of similarities. For example, different molecules, thought to be specific of neuronal cells, are expressed by beta cells. The factors regulating the expression of these molecules in beta cells are poorly understood. In the present work, we have studied the effect of dexamethasone, a synthetic glucocorticoid, on the expression of three different neuronal traits expressed by INS-1 cells, a highly differentiated beta cell line. We demonstrate that dexamethasone treatment decreases the steady state levels of mRNAs coding for both the low- and the high-affinity NGF receptors and of mRNA coding for NF-H, an intermediate neurofilament specific of neurons. This effect was time-dependent, the decrease being detectable after 4-8 h treatment. The decrease in NGF receptors mRNAs steady state levels was paralleled by a decrease in the number of NGF binding sites as demonstrated after Scatchard analysis. We further focused on the mechanisms by which dexamethasone affects the expression of the low affinity NGF receptor. The effect is countered by the glucocorticoid antagonist RU486, indicating that it is mediated by the glucocorticoid receptor. Finally, the decrease in the low-affinity nerve growth factor receptor mRNA steady state level after dexamethasone treatment is not due to mRNA destabilization but can be rather explained through a change in gene transcription.

Tight hormonal control of PrP gene expression in endocrine pancreatic cells.

In this work, we have studied whether PrP, a protein found predominantly on the surface of neurons, could also exist in pancreatic endocrine cells. For this purpose, we have compared the expression of PrP mRNAs in insulin- (beta) and glucagon- (alpha) producing cells to that of neuronal and nonneuronal cells. We show that PrP mRNAs are expressed in all the beta and alpha cells tested at levels similar to those observed in neuronal cells. We also show that the expression of PrP is tightly regulated by hormones. Indeed, recombinant human growth hormone and dexamethasone, 2 factors implicated in beta cell maturation, increase PrP mRNA steady state level. PrP can thus be added to the list of neuronal factors expressed in the endocrine pancreas. Moreover, beta cells could represent a new model to study the control of PrP expression.

Growth hormone and prolactin regulate the expression of nerve growth factor receptors in INS-1 cells.

We have previously demonstrated that beta-cells express both p75NGF-R and Trk-A, the low and high affinity nerve growth factor (NGF) receptors, respectively. In the current study, we provide evidences that in the beta-cell line INS-1, the expression of these receptors is tightly controlled by GH and PRL, two hormones implicated in beta-cell development and function. Within 24 h of treatment of INS-1 cells with human (h) GH, the numbers of low and high affinity NGF-binding sites, calculated after Scatchard analysis, increase 3- and 2.5-fold, respectively. The increase in the concentration of the high affinity NGF-binding sites is paralleled by an increase in Trk-A protein without any change at the mRNA steady state level, suggesting a translational/posttranslational effect. On the other hand, the increase in low affinity binding sites is paralleled by an increase in the p75NGF-R mRNA steady state level. The effect requires at least 8 h of treatment, and a dose of 50 ng/ml hGH is sufficient to induce an increase in the p75NGF-R mRNA steady state level. The effect of hGH can be mimicked in the same time- and dose-dependent manner by rat PRL and bovine GH, suggesting that the expression of NGF receptors can be transduced by both the somatogenic and lactogenic pathways. Finally, the increase in the p75NGF-R mRNA steady state level after PRL treatment is not due to mRNA stabilization, suggesting a transcriptional control, and requires concurrent protein synthesis. GH and PRL could thus be important regulators of the sensitivity of beta-cells to NGF.