Faculty Collaboration Database

Reciprocal Regulation of PASTA Kinase Signaling by Differential Modification. J Bacteriol 2019 May 15;201(10)

Date

03/13/2019

Pubmed ID

30858297

Pubmed Central ID

PMC6482931

DOI

10.1128/JB.00016-19

Scopus ID

2-s2.0-85065131964 (requires institutional sign-in at Scopus site)   8 Citations

Abstract

Transmembrane Ser/Thr kinases containing extracellular PASTA (penicillin-binding protein [PBP] and Ser/Thr-associated) domains are ubiquitous among Actinobacteria and Firmicutes species. Such PASTA kinases regulate critical bacterial processes, including antibiotic resistance, cell division, cell envelope homeostasis, and virulence, and are sometimes essential for viability. Previous studies of purified PASTA kinase fragments revealed they are capable of autophosphorylation in vitro, typically at multiple sites on the kinase domain. Autophosphorylation of a specific structural element of the kinase known as the activation loop is thought to enhance kinase activity in response to stimuli. However, the role of kinase phosphorylation at other sites is largely unknown. Moreover, the mechanisms by which PASTA kinases are deactivated once their stimulus has diminished are poorly understood. Enterococcus faecalis is a Gram-positive intestinal bacterium and a major antibiotic-resistant opportunistic pathogen. In E. faecalis, the PASTA kinase IreK drives intrinsic resistance to cell wall-active antimicrobials, and such antimicrobials trigger enhanced phosphorylation of IreK in vivo Here we identify multiple sites of phosphorylation on IreK and evaluate their function in vivo and in vitro While phosphorylation of the IreK activation loop is required for kinase activity, we found that phosphorylation at a site distinct from the activation loop reciprocally modulates IreK activity in vivo, leading to diminished activity (and diminished antimicrobial resistance). Moreover, this site is important for deactivation of IreK in vivo upon removal of an activating stimulus. Our results are consistent with a model in which phosphorylation of IreK at distinct sites reciprocally regulates IreK activity in vivo to promote adaptation to cell wall stresses.IMPORTANCE Transmembrane Ser/Thr kinases containing extracellular PASTA domains are ubiquitous among Actinobacteria and Firmicutes species and regulate critical processes, including antibiotic resistance, cell division, and cell envelope homeostasis. Previous studies of PASTA kinase fragments revealed autophosphorylation at multiple sites. However, the functional role of autophosphorylation and the relative impacts of phosphorylation at distinct sites are poorly understood. The PASTA kinase of Enterococcus faecalis, IreK, regulates intrinsic resistance to antimicrobials. Here we identify multiple sites of phosphorylation on IreK and show that modification of IreK at distinct sites reciprocally regulates IreK activity and antimicrobial resistance in vivo Thus, these results provide new insights into the mechanisms by which PASTA kinases can regulate critical physiological processes in a wide variety of bacterial species.

Author List

Labbe BD, Hall CL, Kellogg SL, Chen Y, Koehn O, Pickrum AM, Mirza SP, Kristich CJ

Authors

Stephanie Kellogg Research Scientist I in the Microbiology and Immunology department at Medical College of Wisconsin
Olivia Koehn Postdoctoral Fellow in the Pharmacology and Toxicology department at Medical College of Wisconsin
Christopher J. Kristich PhD Professor in the Microbiology and Immunology department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

Adaptation, Physiological
Anti-Infective Agents
Cell Wall
Enterococcus faecalis
Gene Expression Regulation, Bacterial
Phosphorylation
Protein Processing, Post-Translational
Signal Transduction