A Rho-GTPase Effector, Rhotekin-2 (RTKN2) Is Associated with BMP8b and IL-16 Cytokine Expression and Increased Sensitivity to Apoptosis in Lymphocytes.

Rho GTPases are molecular switches controlling a broad range of cellular processes in lymphocytes including activation, immune response and gene transcription. They rapidly cycle between a guanosine diphosphate (GDP)-bound and guanosine triphosphate (GTP)-bound state, and the GTP-bound state is the active conformation that initiates downstream signaling pathways via effector proteins. We identified a novel Rho effector, RTKN2, that is highly expressed in lymphocytes, particulary freshly isolated CD4 T-cells; and is switched off in activated T-cells. When the protein was over-expressed in an unrelated cell line, it conferred resistance to apoptosis, and this resistance was maintained using the media alone from the over-expressing cells. In the current study we investigated genes relating to high or low expression of RTKN2 in lymphocytes. In human samples, RTKN2 expression was down regulated in PHA activated T-cells and it correlated with the expression of Zap-70, a Rho-guanine nucleotide exchange factor (GEF). We then utilized a real time cytokine array (Superarray Bioscience Corporation) to compare high versus low RTKN2 expression levels in the same population of lymphocytes. The real time array linked high levels of RTKN2 with cytokines BMP8b, (a TGF-beta family protein) and IL-16 (a modulator of T cell activation, and an inhibitor of HIV replication relationship), and this observation was confirmed in a series of T-cell samples. Further, we studied the effect of down regulation of RTKN2 in primary lymphocytes, using short hairpin (sh)RNA plasmid that also expressed GFP. Transfected GFP-positive cells had lower survival than cells transfected with scrambled (sh)RNA, and also demonstrated increased sensitivity to the the induction of apoptosis by oxysterols. These findings indicate that RTKN2 plays a role in survival of T-lymphocytes and this may be mediated through the secretion of cytokines.