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Elucidating the Role of an Atypical Anti‐Termination Two‐Component System in Mycobacterial Gene Regulation and Pathogenesis
Author(s) -
Davenport Haley,
Rohde Kyle
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2019.33.1_supplement.lb357
Subject(s) - biology , mycobacterium smegmatis , terminator (solar) , two component regulatory system , complementation , gene , response regulator , genetics , transcriptional regulation , mutant , mycobacterium tuberculosis , transcription factor , microbiology and biotechnology , tuberculosis , medicine , ionosphere , physics , pathology , astronomy
M. tuberculosis , the etiologic agent of tuberculosis, caused 1.3 million deaths and infected 10 million people globally in 2017. During infection, many bacteria respond to their environments using two‐component systems (TCS) consisting of a membrane‐bound histidine kinase (HK) and a response regulator (RR) of transcriptional initiation. Rv3220c (HK) and Rv1626 (RR) are an atypical TCS with a cytosolic HK and an RR acting as a transcriptional anti‐terminator via its ANTAR domain. This domain causes formation of an anti‐terminator structure to allow transcription to proceed past its normal termination point. Rv1626 was previously identified by a transposon screen as involved in cholesterol metabolism, but the role of this TCS in pathogenesis is unknown. To determine the function of this TCS, knock‐out (KO) constructs were created using PCR, TOPO cloning, and recombineering. Due to the safety, faster growth, and genetic similarity of M. smegmatis to M. tuberculosis , we targeted the orthologs of the HK and RR in this surrogate organism, MSMEG_1918 and MSMEG_3246 respectively, for inactivation. After generating KO mutants, a phenotypic study was done to determine if cholesterol metabolism was inhibited by the gene deletion. Secondly, mycobacterial protein fragment complementation was used to confirm protein‐protein interactions between the putative TCS proteins. Findings from M. smegmatis will be validated in future work by conducting similar studies in pathogenic mycobacteria. Support or Funding Information The project was supported by startup funds to Dr. Kyle Rohde from University of Central Florida College of Medicine. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .