Abstract 1392: G Protein Coupled Receptor Kinase 6 Couples Stromal Cell Derived Factor-1α-CXCR4 Signaling to ERK/AKT Activation in Rat Cardiac Stem Cells
Stromal cell derived factor-1α (SDF1) through the interaction with it receptor CXCR4 plays an important role in regulating the localization and function of hematopoietic stem cells. Recently, we have described SDF1-CXCR4 signaling in the heart and found it to regulate myocardial survival through the activation of ERK and AKT. CXCR4 is known to couple to the heterotrimeric G-Protein, Gi. The G-protein receptor kinase 6 (GRK6), may serve to uncouple the receptor from downstream signaling or to facilitate signaling via phosphorylation- dependent and independent mechanisms. To further understand cardiac SDF1-CXCR4 signaling and function we used rat clonal cKit+ cardiac stem cells (rCSC) to assess the role of GRK6 in CXCR4 responses to SDF1. To specifically target GRK6 actions in SDF1-CXCR4 signaling and function in rCSCs we used siRNA to knockdown GRK6 in vitro. In untreated rCSCs, GRK6 specific siRNA significantly knocked down the expression of GRK6 by 50 – 60% which lead to significant decreases in levels of P-AKT, and P-ERK (about 70 - 90% respectively, P± 0.05) and increased levels of P-JNK, P-p38, and Caveolin1 in untreated rCSC. The treatment of rCSC with 25 nM SDF1 significantly increased GRK6 expression and this increase was attenuated by GRK6 specific siRNA. AKT, ERK, JNK, and p38 phosphorylation were significantly increased 5 min after rCSC treatment with 25nM SDF1 (0.5–3 fold increase, P<0.05) and GRK6 siRNA significantly attenuated SDF1activation (0.5 to 2 fold decrease, P<0.05). Our results indicate that in rCSC, GRK6 is necessary for SDF1-CXCR4 dependent ERK and AKT signaling Thus in rCSCs GRK6 serves predominantly in G-protein independent SDF1-CXCR4 signaling possibly through caveolar signaling complexes. In conclusion, in rCSCs SDF1-CXCR4 interaction activates downstream ERK and AKT activation via a GRK6 dependent mechanism. These findings will be important for the understanding of SDF1-CXCR4 regulation of rCSC function.