Complex Regulation of the DnaJ Homolog CbpA by the Global Regulators σ ^S and Lrp, by the Specific Inhibitor CbpM, and by the Proteolytic Degradation of CbpM

Abstract

CbpA is a DnaJ homolog that functions as a DnaK cochaperone. Several cellular processes, including growth at low and high temperatures and septum formation during cell division, require either CbpA or DnaJ. CbpA is encoded in an operon with the gene for CbpM, which is a specific in vivo and in vitro inhibitor of CbpA. Here, we have cooverexpressed CbpA with CbpM in a ⌬cbpAM ⌬dnaJ strain and examined the resulting phenotypes. Under these conditions, sufficient free CbpA activity was present to support growth at low temperatures, but not at high temperatures. Defects in cell division and in replication were also partially complemented by CbpA when cooverexpressed with CbpM. Utilizing reporter fusions, we demonstrated that the cbpAM operon was maximally transcribed at the transition from exponential growth to stationary phase. Transcription was controlled by the S and Lrp global regulators, and both leucine availability and growth temperature influenced transcription. CbpA and CbpM accumulated to similar levels in stationary phase, ϳ2,300 monomers per cell. When not bound to CbpA, CbpM was unstable and was degraded by the Lon and ClpAP proteases. These data demonstrate that CbpA activity is controlled at multiple levels.

Structural Basis of the Regulation of the CbpA Co-chaperone by its Specific Modulator CbpM

“…19 It is known that CbpA and CbpM are co-expressed and accumulate to similar levels in the stationary phase of cell growth. 22 It was shown also that CbpM regulates the DNA-binding activity of CbpA, 18 and it was suggested that the inhibitory effect of CbpM on CbpA might prevent DnaK from interacting with the specific set of substrates in the cell. 22 However, the molecular mechanism underlying this inhibition is not clear.…”

“…19 However, CbpM interaction with CbpA is known to inhibit the DNA binding of CbpA. 21,22 DNA does not bind the J-domain but the region C-terminal to the Jdomain of CbpA, 19 suggesting additional role of this region in regulation by CbpM.…”

“…18 It has been shown that CbpM binds specifically to CbpA and inhibits both the co-chaperone activity and the DNA-binding activity of CbpA. 21,22 It was suggested that during certain growth or stress conditions CbpA might be released from CbpM and recruited by DnaK to function as a co-chaperone. 18 The CbpM interacting region is within the J-domain of CbpA (Jdom CbpA ) and they bind with a 1:1 ratio, 19 although the stoichiometry of the complex was not studied in detail.…”

CbpA Acts as a Modulator of HspR Repressor DNA Binding Activity in Helicobacter pylori

“…First identified as a DNA binding protein, E. coli CbpA binds DNA efficiently, with a preference for curved DNA, and has been localized to the nucleoids of stationaryphase cells (1,16,30). More recent studies have revealed that E. coli CbpA can function as a cochaperone for the DnaK/ Hsp70 chaperone system, and its activity can be modulated by CbpM, an 11-kDa protein that has structural homology to DafA of Thermus thermophilus (2,6,7,8).…”

A Conserved Acidic Amino Acid Mediates the Interaction between Modulators and Co-Chaperones in Enterobacteria

“…[11][12][13][14] The cbpA and cbpM genes form an operon and are co-expressed, and examination of sequenced genomes reveals that this operon is found in a wide variety of bacteria. 12,13 The molecular surface of CbpA that is bound by CbpM locates to the J-domain and primarily consists of positively charged amino acids (R26, R30, and H33) 7,11 (Fig.…”