Microbiology Virulent Protein Kinase A

Nancy R. Gough

Science's STKE, AAAS, Washington, DC 20005, USA

The Yersinia Gram-negative bacteria are pathogenic to humans causing such diseases as bubonic plague. Through the action of a type III protein secretion system, Yersinia injects bacterial proteins necessary to subvert the host cell's normal processes and to promote bacterial survival and replication. Navarro et al. report that, in addition to the previously known inhibitory activity toward guanosine triphosphatases (GTPases) Rac1 and RhoA, Yersinia protein kinase A (YpkA) also inhibits G protein (heterotrimeric guanine nucleotide-binding protein) signaling by phosphorylation of the G subunit. The actin cytoskeleton disruption in cells expressing either the full-length YpkA or a C-terminal fragment was prevented if the cells also expressed a constitutively active form of RhoA, suggesting that YpkA acted upstream of this GTPase. G protein-coupled receptors can stimulate reorganization of the actin cytoskeleton through the activation of RhoA. Exposure of cells transfected with the muscarinic receptor to carbachol stimulated G_q signaling and promoted the formation of stress fibers. Transfection of YpkA, but not a catalytically inactive version, inhibited this G_q-induced reorganization of the cytoskeleton. YpkA coimmunoprecipitated with G_q (both GDP-bound and GTP-bound forms) and G_q was phosphorylated when immunoprecipitated from cells expressing catalytically active YpkA. The phosphorylation site was mapped to a serine within the consensus guanine nucleotide-binding loop by liquid chromatography tandem mass spectrometry. Phosphorylation of this residue appeared to disrupt activation of the G protein as substitution of the serine with aspartate (to mimic phosphorylation) prevented GTP loading. Not only was signaling from the muscarinic receptor to the cytoskeleton blocked by expression of YpkA, so was nuclear translocation of tubby, a transcriptional regulator released from the plasma membrane in response to G_q-mediated stimulation of phospholipase C. In cells infected with Yersinia, both wild-type and catalytically inactive YpkA disrupted the actin cytoskeleton, presumably because the protein contains two functional domains that can inhibit the actin cytoskeleton: the C-terminal guanine nucleotide dissociation inhibitor region that directly targets RhoA and the serine-threonine kinase activity of the N-terminal region that inhibits G protein signaling to the cytoskeleton.

L. Navarro, A. Koller, R. Nordfelth, H. Wolf-Watz, S. Taylor, J. E. Dixon, Identification of a molecular target for the Yersinia protein kinase A. Mol. Cell 26, 465-477 (2007). [PubMed]