CONSTITUTIVELY SIGNALING FRAGMENTS OF TSR, THE ESCHERICHIA-COLI SERINE CHEMORECEPTOR

Tsr, the serine chemoreceptor of Escherichia coli, has two signaling m odes. One augments clockwise (CW) flagellar rotation, and the other au gments counterclockwise (CCW) rotation. To identify the portion of the Tsr molecule responsible for these activities, we isolated soluble fr agments of the Tsr cytoplasmic domain that could alter the flagellar r otation patterns of unstimulated wild-type cells. Residues 290 to 470 from wild-type Tsr generated a CW signal, whereas the same fragment wi th a single amino acid replacement (alanine 313 to valine) produced a CCW signal. The soluble components of the chemotaxis phosphorelay syst em needed for expression of these Tsr fragment signals were identified by epistasis analysis. Like full-length receptors, the fragments appe ared to generate signals through interactions with the CheA autokinase and the CheW coupling factor. CheA was required for both signaling ac tivities, whereas CheW was needed only for CW signaling. Purified Tsr fragments were also examined for effects on CheA autophosphorylation a ctivity in vitro. Consistent with the in vivo findings, the CW fragmen t stimulated CheA, whereas the CCW fragment inhibited CheA. CheW was r equired for stimulation but not for inhibition. These findings demonst rate that a 180-residue segment of the Tsr cytoplasmic domain can prod uce two active signals. The CCW signal involves a direct contact betwe en the receptor and the CheA kinase, whereas the CW signal requires pa rticipation of CheW as well. The correlation between the in vitro effe cts of Tsr signaling fragments on CheA activity and their in vivo beha vioral effects lends convincing support to the phosphorelay model of c hemotactic signaling.