Reduced expression of the neuron restrictive silencer factor permits transcription of glycine receptor α1 subunit in small-cell lung cancer cells

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Abstract
Small-cell lung cancer (SCLC) cells express various markers of neuronal differentiation associated with deficient activity of the neuron-restricted silencer factor (NRSF). Here, we characterize mechanisms by which NRSF target genes are upregulated in SCLC and their functional consequences for cell survival. Since the glycine receptor (GlyR) α1 subunit gene, GLRA1, contains a sequence motif for NRSF binding (NRSE) within its 5′ UTR, it served as a cellular surrogate marker for NRSF activity. Expression of GLRA1 in nontransformed cells is largely restricted to cells in the spinal cord, retina and brain stem. In experiments described here, we detected GLRA1 transcripts in three out of four SCLC-derived cell lines and in three of five biopsy samples obtained from SCLCs. In contrast, no GLRA1 transcripts were found in 10 nonmalignant nor 15 non-small-cell lung cancer biopsies. Consistent with this observation, NRSF-mediated suppression of an expression construct harbouring the NRSE of the GLRA1 (GLRA1 NRSE) gene was impaired in three of four ‘classic’ SCLC cell lines, whereas exogenous overexpression of NRSF in NRSF-deficient SCLC cell lines reconstituted silencing of the reporter plasmid. The level of NRSF transcripts as well as the level of specifically bound NRSF to the NRSE correlated with the level of GLRA1 transcripts in SCLC cell lines. Splice variants encoding truncated NRSF proteins and expressed in some SCLC did not antagonize the repression of NRSE-containing genes. Most interestingly, reconstitution of NRSF expression induced apoptosis in SCLC cells, suggesting that inhibition of NRSF activity is a crucial step in the carcinogenesis of a subgroup of SCLC.