Autoproteolysis and Intramolecular Dissociation of Yersinia YscU Precedes Secretion of Its C-Terminal Polypeptide YscUCC

Type III secretion system mediated secretion and translocation of Yop-effector proteins across the eukaryotic target cell membrane by pathogenic Yersinia is highly organized and is dependent on a switching event from secretion of early structural substrates to late effector substrates (Yops). Substrate switching can be mimicked in vitro by modulating the calcium levels in the growth medium. YscU that is essential for regulation of this switch undergoes autoproteolysis at a conserved N↑PTH motif, resulting in a 10 kDa C-terminal polypeptide fragment denoted YscU_CC. Here we show that depletion of calcium induces intramolecular dissociation of YscU_CC from YscU followed by secretion of the YscU_CC polypeptide. Thus, YscU_CC behaved in vivo as a Yop protein with respect to secretion properties. Further, destabilized yscU mutants displayed increased rates of dissociation of YscU_CC in vitro resulting in enhanced Yop secretion in vivo at 30°C relative to the wild-type strain.These findings provide strong support to the relevance of YscU_CC dissociation for Yop secretion. We propose that YscU_CC orchestrates a block in the secretion channel that is eliminated by calcium depletion. Further, the striking homology between different members of the YscU/FlhB family suggests that this protein family possess regulatory functions also in other bacteria using comparable mechanisms.