Journal article
Functional Characterization of Non-Ankyrin Repeat Domains of Orientia tsutsugamushi Ank Effectors Reveals Their Importance for Molecular Pathogenesis
Infection and immunity, Vol.90(5), pp.e0062821-e0062821
05/19/2022
DOI: 10.1128/iai.00628-21
PMCID: PMC9119115
PMID: 35435726
Abstract
Orientia tsutsugamushi is a genetically intractable obligate intracellular bacterium, causes scrub typhus, and has one of the largest known armamentariums of ankyrin repeat-containing effectors (Anks). Most have a C-terminal F-box presumed to interact with the SCF ubiquitin ligase complex primarily based on their ability to bind overexpressed Skp1. Whether all F-box-containing Anks bind endogenous SCF components and the F-box residues essential for such interactions has gone unexplored. Many O. tsutsugamushi Ank F-boxes occur as part of a PRANC (pox protein repeats of ankyrin-C-terminal) domain. Roles of the non-F-box portion of the PRANC and intervening sequence region (ISR) that links the ankyrin repeat and F-box/PRANC domains are unknown. The functional relevance of these effectors' non-ankyrin repeat domains was investigated. The F-box was necessary for Flag-tagged versions of most F-box-containing Anks to precipitate endogenous Skp1, Cul1, and/or Rbx1, while the ISR and PRANC were dispensable. Ank toxicity in yeast was predominantly F-box dependent. Interrogations of Ank1, Ank5, and Ank6 established that L1, P2, E4, I9, and D17 of the F-box consensus are key for binding native SCF components and for Ank1 and Ank6 to inhibit NF-κB. The ISR is also essential for Ank1 and Ank6 to impair NF-κB. Ectopically expressed Ank1 and Ank6 lacking the ISR or having a mutagenized F-box incapable of binding SCF components performed as dominant-negative inhibitors to block O. tsutsugamushi NF-κB modulation. This study advances knowledge of O. tsutsugamushi Ank functional domains and offers an approach for validating their roles in infection.
Details
- Title: Subtitle
- Functional Characterization of Non-Ankyrin Repeat Domains of Orientia tsutsugamushi Ank Effectors Reveals Their Importance for Molecular Pathogenesis
- Creators
- Sarika Gupta - Virginia Commonwealth University Medical CenterJason R Hunt - Virginia Commonwealth University Medical CenterHaley E Adcox - Virginia Commonwealth University Medical CenterShelby E Andersen - Roy J. and Lucille A. Carver College of MedicineJacob J Gumpf - Virginia Commonwealth University Medical CenterRyan S Green - Virginia Commonwealth University Medical CenterAndrea R Beyer - Virginia Commonwealth University Medical CenterSean M Evans - Virginia Commonwealth University Medical CenterLauren VieBrock - Virginia Commonwealth University Medical CenterCurtis B Read - Virginia Commonwealth University Medical CenterMary M Weber - Roy J. and Lucille A. Carver College of MedicineJason A Carlyon - Virginia Commonwealth University Medical Center
- Resource Type
- Journal article
- Publication Details
- Infection and immunity, Vol.90(5), pp.e0062821-e0062821
- DOI
- 10.1128/iai.00628-21
- PMID
- 35435726
- PMCID
- PMC9119115
- ISSN
- 0019-9567
- eISSN
- 1098-5522
- Grant note
- R56 AI123346 / NIAID NIH HHS R01 AI155434 / NIAID NIH HHS 20PRE35210610 / American Heart Association (AHA) R01 AI123346 / NIAID NIH HHS R01 AI150812 / NIAID NIH HHS R01 AI167857 / NIAID NIH HHS
- Language
- English
- Date published
- 05/19/2022
- Academic Unit
- Molecular Physiology and Biophysics; Microbiology and Immunology
- Record Identifier
- 9984297330802771
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