Structural perspective of ARHI mediated inhibition of STAT3 signaling: an insight into the inactive to active transition of ARHI and its interaction with STAT3 and importinβ.

ARHI, a putative tumor suppressor protein with unique 32 amino acid extension in the N-terminal region, differs from oncogenes Ras and Rap, negatively regulates STAT3 signaling and inhibits the migration of ovarian cancer cells. ARHI associates directly with STAT3, also forms complex with importinβ, and prevents formation of RanGTPase-importinβ complex, which is essential for transporting STAT3 into the nucleus. Hence, the structural aspects pertaining to ARHI mediated inhibition of STAT3 translocation can provide hints on the regulation of STAT3 signaling mechanism. Accordingly, in the present study, the structure of ARHI was predicted and its transition from inactive to active state studied using MD simulations and free energy landscape analysis. The transition of ARHI is marked by the movement of switch I region towards γ-phosphate of GTP, in addition, the hydrophobic interaction between N-terminal helix and switch II region of ARHI accounts for its low intrinsic GTPase activity. Further, the protein-protein interaction studies reveal that the residues of N-terminal helix, effector domain, P-loop and G box motif of ARHI actively form polar and non-polar interaction with NTD of STAT3 and make them compact thereby rendering STAT3 inaccessible for Ran-importinβ mediated translocation. On the other hand, ARHI competes with RanGTPase and interacts with importinβ via basic-acidic patch interaction, which leads to inhibition of STAT3 translocation. The interacting residues involved for this structural mechanism would be instrumental in designing inhibitors for STAT3, which mimics ARHI thereby leading to the suppression of cancer cell growth.