α-Internexin and the neurofilament triplet proteins (NF-L, NF-M, and NF-H) co-assemble into intermediate filament networks in neurons. We have found that the RE1 silencing transcription factor (REST) plays a contributory role in the neuron-specific expression of the α-internexin, NF-H and NF-M genes. Chromatin immunoprecipitation and transient transfection experiments performed with catecholaminergic neuronal Cath a.-differentiated (CAD) cells and non-neuronal NIH3T3 cells demonstrated that REST repressed transcription of these genes in NIH3T3 cells by binding and recruiting mSin3A, CoREST, histone deacetylase (HDAC) 1 and MeCP2 to the RE1 sites in the intron-1 of α-internexin and the 5′ flanking regions of NF-H and NF-M. No repression effect of the RE1 sites was observed in CAD cells, which express these neuronal genes but not REST. Treatment of NIH3T3 cells with trichostatin A, a HDAC inhibitor, relieved the REST-mediated repression and induced ectopic activation of α-internexin, NF-H and NF-M. The trichostatin A treatment did not affect the levels of REST occupancy but caused coordinated changes in acetylation and methylation of histones around the RE1 sites of these genes in NIH3T3 cells consistent with a transition from transcriptional repression to transcriptional activation. Thus, REST regulates expression of these neuronal genes, partly by a HDAC-dependent epigenetic mechanism.