Escherichia coli YaeL (EcfE) is a homolog of human site-2 protease (S2P), a membrane-bound zinc metalloprotease involved in regulated intramembrane proteolysis. We have shown previously that YaeL, having essential metalloprotease active site motifs in the cytoplasmic domain, is indispensable for viability. Here, we obtained rpoE, encoding an extracytoplasmic stress response sigma factor (sigma(E)), as a multicopy suppressor against the yaeL disruption. Whereas sigma(E) is thought to be activated by regulated cleavage of RseA on the periplasmic side by the DegS protease, we found that a degradation intermediate of RseA consisting of the transmembrane and the cytoplasmic domains accumulated in the YaeL-depleted cells. This intermediate was degraded on expression of YaeL but not of its metalloprotease motif mutants. Cells depleted of YaeL were incapable of activating a sigma(E)-dependent promoter in response to an envelope stress. It is suggested that sigma(E) activation involves two successive proteolytic cleavages: first, at a periplasmic site by DegS; second, at a cytoplasmic or intramembrane site by YaeL. Thus, YaeL is positively required for the sigma(E) extracytoplasmic stress response.