G protein α₁₂ (Gα₁₂) is a negative regulator of kidney injury molecule-1-mediated efferocytosis

Kidney injury molecule-1 (KIM-1) is a receptor for the "eat me" signal, phosphatidylserine, on apoptotic cells. The specific upregulation of KIM-1 by injured tubular epithelial cells (TECs) enables them to clear apoptotic cells (also known as efferocytosis), thereby protecting from acute kidney injury. Recently, we uncovered that KIM-1 binds directly to the α-subunit of heterotrimeric G₁₂ protein (Gα₁₂) and inhibits its activation by reactive oxygen species during renal ischemia-reperfusion injury (Ismail OZ, Zhang X, Wei J, Haig A, Denker BM, Suri RS, Sener A, Gunaratnam L. Am J Pathol 185: 1207-1215, 2015). Here, we investigated the role that Gα₁₂ plays in KIM-1-mediated efferocytosis by TECs. We showed that KIM-1 remains bound to Gα₁₂ and suppresses its activity during phagocytosis. When we silenced Gα₁₂ expression using small interefering RNA, KIM-1-mediated engulfment of apoptotic cells was increased significantly; in contrast overexpression of constitutively active Gα₁₂ (QLGα₁₂) resulted in inhibition of efferocytosis. Inhibition of RhoA, a key effector of Gα₁₂, using a chemical inhibitor or expression of dominant-negative RhoA, had the same effect as inhibition of Gα₁₂ on efferocytosis. Consistent with this, silencing Gα₁₂ suppressed active RhoA in KIM-1-expressing cells. Finally, using primary TECs from Kim-1^+/+ and Kim-1^-/- mice, we confirmed that engulfment of apoptotic cells requires KIM-1 expression and that silencing Gα₁₂ enhanced efferocytosis by primary TECs. Our data reveal a previously unknown role for Gα₁₂ in regulating efferocytosis and that renal TECs require KIM-1 to mediate this process. These results may have therapeutic implications given the known harmful role of Gα₁₂ in acute kidney injury.