A genetic screen for mutations affecting temperature sensing in Bacillus subtilis

DÍAZ, ALEJANDRA; PORRINI, L; DE MENDOZA, D; MANSILLA, MC (CORRESPONDING)

MICROBIOLOGY-UK

SOC GENERAL MICROBIOLOGY

Lugar: London; Año: 2018

1350-0872

Two component systems, composed of a receptorhistidine kinase and a cytoplasmic response regulator, regulate pivotalcellular processes in microorganisms. Here we describe a new screeningprocedure for the identification of amino acids that are crucial for the functioningof DesK, a prototypic thermosensor histidine kinase from Bacillus subtilis. This experimental strategy involves randommutagenesis of the membrane sensor domain of the DesK coding sequence, followedby the use of a detection procedure based on changes in the colonymorphogenesis that take place during the sporulation program of B. subtilis. This method permitted usthe recovery of mutants defective in DesK temperature sensing. This screening approachcould be applied to all histidine kinases of B. subtilis and also to kinases of other bacteria that arefunctionally expressed in this organism. Moreover, this reporter assay could beexpanded to develop reporter assays for a variety of transcriptionallyregulated systems.