3p21.3 Tumor Suppressor Gene H37/Luca15/RBM5 Inhibits Growth of Human Lung Cancer Cells through Cell Cycle Arrest and Apoptosis

Abstract

Deletion at chromosome 3p21.3 is the earliest and the most frequently observed genetic alteration in lung cancer, suggesting that the region contains tumor suppressor gene(s) (TSG). Identification of those genes may lead to the development both of biomarkers to identify high-risk individuals and novel therapeutics. Previously, we cloned the H37/Luca15/RBM5 gene from 3p21.3 and showed its TSG characteristics. To investigate the physiologic function of H37 in the lung and its mechanism of tumor suppression, we have stably transfected H37 into A549 non-small cell lung cancer cells. A549/H37 cells show significant growth inhibition compared with the vector controls by in vitro and in vivo cell proliferation assays. Using this lung cancer cell model, we have found that the molecular mechanism of H37 tumor suppression involves both cell cycle (G 1 ) arrest and apoptosis. To further define H37's function in cell cycle/apoptotic pathways, we investigated differential expression profiles of various cell cycle and apoptosis regulatory proteins using Western blot analysis. Both cyclin A and phophorylated RB levels were decreased in H37-transfected cells, whereas expression of Bax protein was increased. Mitochondrial regulation of apoptosis further downstream of Bax was investigated, showing change in the mitochondrial membrane potential, cytochrome c release into the cytosol, and enhanced caspase-9 and caspase-3 activities. We also report that H37 may mediate apoptosis in a p53-independent manner, and Bax knockdown by small interfering RNA suggests Bax plays a functional role downstream of H37. Lastly, we proposed a tumor suppression model of H37 as a post-transcriptional regulator for cell cycle/apoptotic-related proteins.

Up-regulation of the proapoptotic caspase 2 splicing isoform by a candidate tumor suppressor, RBM5

“…RBM5 expression is repressed in 70-80% of lung cancers (25). RBM5 overexpression causes cell cycle arrest, apoptosis, and inhibition of tumor growth (25)(26)(27)29), sensitizing cells to apoptosis induced by death receptor ligands, FAS, TNF-␣, and TRAIL (31,32). However, the mechanism for this sensitization was not known.…”

Alternative pre-mRNA splicing regulation in cancer: pathways and programs unhinged

“…It was soon realized that this encoded an inhibitory RNA complementary to the 39 untranslated region (UTR) of the RBM5 mRNA, and that full-length, sense-oriented RBM5 actually potentiates apoptosis initiated by Fas (Rintala-Maki and Sutherland 2004). In addition to its role in apoptosis, expression of RBM5 has been found to inhibit proliferation when transfected into several different cell lines (Edamatsu et al 2000;Oh et al 2006). RBM5 also appears to stimulate p53 transcription and promote higher levels of p53 transcripts through an unknown mechanism (Kobayashi et al 2010).…”

The two single nucleotide polymorphisms in the H37/RBM5 tumour suppressor gene at 3p21.3 correlated with different subtypes of non-small cell lung cancers

“…The growing literatures on H37 strongly suggest its involvement in apoptosis and cell cycle regulation, with all results converging on a role for H37 as a TSG. Using the lung cancer cell model, we showed that H37 clearly inhibits tumour growth both in vitro and in vivo with antitumour mechanisms involving cell cycle (G1) arrest and apoptosis [11]. H37 also mediates growth inhibition by eliciting reduced expression of cyclin A and pRB as well as by increased expression of pro-apoptotic protein Bax [11].…”

“…Using the lung cancer cell model, we showed that H37 clearly inhibits tumour growth both in vitro and in vivo with antitumour mechanisms involving cell cycle (G1) arrest and apoptosis [11]. H37 also mediates growth inhibition by eliciting reduced expression of cyclin A and pRB as well as by increased expression of pro-apoptotic protein Bax [11]. Consistently, H37 triggers mitochondrial apoptotic pathways downstream of Bax, in a p53-independent manner, which include breakdown in the mitochondrial membrane potential, cytochrome c release into cytosol, and enhanced caspase-9 and caspase-3 activities [11].…”

“…H37 also mediates growth inhibition by eliciting reduced expression of cyclin A and pRB as well as by increased expression of pro-apoptotic protein Bax [11]. Consistently, H37 triggers mitochondrial apoptotic pathways downstream of Bax, in a p53-independent manner, which include breakdown in the mitochondrial membrane potential, cytochrome c release into cytosol, and enhanced caspase-9 and caspase-3 activities [11]. Further, as compared with adjacent normal tissue, our findings show reduced expression in primary lung cancers of H37 transcript and protein in 9 of 11 (82%) and 46 of 62 (73%) samples, respectively [12], although the H37/3p21.3 LOH genomic status for these samples was unknown.…”