Open AccessA genetic screen for mutations affecting temperature sensing in Bacillus subtilis
Author(s) -
A. Díaz,
Lucía Porrini,
Diego de Mendoza,
Marı́a C. Mansilla
Publication year - 2019
Publication title -
microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.019
H-Index - 179
eISSN - 1465-2080
pISSN - 1350-0872
DOI - 10.1099/mic.0.000741
Subject(s) - bacillus subtilis , histidine kinase , response regulator , histidine , kinase , biology , two component regulatory system , mutagenesis , mutant , protein kinase domain , biochemistry , microbiology and biotechnology , bacteria , computational biology , genetics , gene , amino acid
Two component systems, composed of a receptor histidine kinase and a cytoplasmic response regulator, regulate pivotal cellular processes in microorganisms. Here we describe a new screening procedure for the identification of amino acids that are crucial for the functioning of DesK, a prototypic thermosensor histidine kinase from Bacillus subtilis. This experimental strategy involves random mutagenesis of the membrane sensor domain of the DesK coding sequence, followed by the use of a detection procedure based on changes in the colony morphogenesis that take place during the sporulation programme of B. subtilis. This method permitted us the recovery of mutants defective in DesK temperature sensing. This screening approach could be applied to all histidine kinases of B. subtilis and also to kinases of other bacteria that are functionally expressed in this organism. Moreover, this reporter assay could be expanded to develop reporter assays for a variety of transcriptionally regulated systems.