Localization of the histidine kinase PilS to the poles of Pseudomonas aeruginosa and identification of a localization domain.
Transcription of the type IV pilus subunit gene of Pseudomonas aeruginosa is controlled by a two-component signal transduction system. PilS, the histidine kinase, is membrane bound and PilR, its cognate response regulator, is cytoplasmic. The signal that activates PilS is unknown. PilS has three domains: (i) The N-terminus, predicted to form six transmembrane (TM) helices; (ii) a central linker domain; and (iii) the C-terminal transmitter domain containing all the conserved residues of sensor kinases. A translational fusion of the gfp gene (green fluorescent protein) to the 3' end of pilS was used to determine the position of PilS in the bacterial cell. Epifluorescence microscopy revealed that PilS is retained to the poles of P. aeruginosa but is distributed evenly about the membrane of Escherichia coli. Deletions of the PilS-GFP fusion revealed that the TM domain was sufficient and necessary to bring GFP to the membrane of P. aeruginosa and E. coli but was not sufficient to confine GFP to the poles. Retention to the poles of P. aeruginosa required both the TM and linker domains. Replacement of the PilS TM domain with an E. coli membrane protein, MalG, still allowed polar localization. Therefore, the PilS TM domain positions the linker domain close to the membrane allowing it to interact with the putative polar anchor which is specific to P. aeruginosa.
Boyd
J M
JM
University of Calgary, Microbiology and Infectious Diseases, 3330 Hospital Drive, N.W. Calgary, Alberta T2N 4N1, Canada.
D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification*; D002451:Cell Compartmentation; D004926:Escherichia coli / Q000201:enzymology; D049452:Green Fluorescent Proteins; D000071677:Histidine Kinase; D008164:Luminescent Proteins / Q000235:genetics / Q000302:isolation & purification; D008856:Microscopy, Fluorescence; D011494:Protein Kinases / Q000235:genetics / Q000302:isolation & purification*; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification; D014157:Transcription Factors / Q000235:genetics / Q000302:isolation & purification*
D001426:Bacterial Proteins; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C080687:pilS protein, Bacteria; D049452:Green Fluorescent Proteins; D011494:Protein Kinases; D000071677:Histidine Kinase