Region q14 of human chromosome 13 is frequently deleted in hematopoietic malignancies, including chronic lymphocytic leukemia (CLL) and multiple myeloma (MM). Region commonly deleted in those tumors encompasses parts of four adjacent genes, termed DLEU1, DLEU2 (both ORFIess), RFP2 and KCNRG. As no mutations were found in those genes in CLL samples, haploinsufficiency mechanism of gene inactivation was suggested. KCNRG encodes protein with a high homology to tetramerization domain of voltage-gated K+ channels (Kv channels). This protein may interfere with the normal assembly of the K+ channel proteins, binding to their tetramerization domain and causing the suppression of Kv channels. Using the patch-clamp technique, we determined that KCNRG indeed suppresses Kv channel activity in the malignant human prostate cells (Ivanov et al., 2003). It has been shown that in LNCaP prostate cells and human lymphocytes the rate of mitoses decreases when Kv channels are blocked, implying that K+ channels are involved in cell proliferation. Here we propose a thorough study of influence of KCNRG gene on white blood cell proliferation and apoptosis intended to discover a clues to MM and CLL pathogenesis. A level of KCNRG expression in HL-60, RPMI-8226 and LNCaP cal line will be evaluated. We will induce an overexpression of KNCRG gene in LNCaP, HL-60 and RPMI-8226 cell lines and will quantify its effects on cell morphology, proliferation and apoptosis. An assessment of differentiation of control and KCNRG-overexpressing HL-60 cells will be performed. An assessment of invasiveness of HL-60 control and KCNRG-overexpressing HL-60 cells will be performed. Also we will perform microarray profiling of wild-type LNCaP, HL-60 and RPMI-8226 cell lines before and after transfection with KCNRG-overexpressing construct. Common gene changes revealed by activation of KCNRG will be confirmed in Real-Time PCR experiments. ? ? ?
