AUTHOR=Vorkapic Dina , Mitterer Fabian , Pressler Katharina , Leitner Deborah R. , Anonsen Jan Haug , Liesinger Laura , Mauerhofer Lisa-Maria , Kuehnast Torben , Toeglhofer Manuela , Schulze Adina , Zingl Franz G. , Feldman Mario F. , Reidl Joachim , Birner-Gruenberger Ruth , Koomey Michael , Schild Stefan 

TITLE=A Broad Spectrum Protein Glycosylation System Influences Type II Protein Secretion and Associated Phenotypes in Vibrio cholerae

JOURNAL=Frontiers in Microbiology

VOLUME=10

YEAR=2019

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2019.02780

DOI=10.3389/fmicb.2019.02780

ISSN=1664-302X

ABSTRACT=<p>Protein secretion plays a crucial role for bacterial pathogens, exemplified by facultative human-pathogen <italic>Vibrio cholerae</italic>, which secretes various proteinaceous effectors at different stages of its lifecycle. Accordingly, the identification of factors impacting on protein secretion is important to understand the bacterial pathophysiology. PglL<sub>Vc</sub>, a predicted oligosaccharyltransferase of <italic>V. cholerae</italic>, has been recently shown to exhibit <italic>O</italic>-glycosylation activity with relaxed glycan specificity in an engineered <italic>Escherichia coli</italic> system. By engineering <italic>V. cholerae</italic> strains to express a defined, undecaprenyl diphosphate-linked glycoform precursor, we confirmed functional <italic>O</italic>-linked protein glycosylation activity of PglL<sub>Vc</sub> in <italic>V. cholerae</italic>. We demonstrate that PglL<sub>Vc</sub> is required for the glycosylation of multiple <italic>V. cholerae</italic> proteins, including periplasmic chaperones such as DegP, that are required for efficient type II-dependent secretion. Moreover, defined deletion mutants and complementation strains provided first insights into the physiological role of <italic>O</italic>-linked protein glycosylation in <italic>V. cholerae</italic>. RbmD, a protein with structural similarities to PglL<sub>Vc</sub> and other established oligosaccharyltransferases (OTases), was also included in this phenotypical characterization. Remarkably, presence or absence of PglL<sub>Vc</sub> and RbmD impacts the secretion of proteins via the type II secretion system (T2SS). This is highlighted by altered cholera toxin (CT) secretion, chitin utilization and biofilm formation observed in Δ<italic>pglL</italic><sub>Vc</sub> and Δ<italic>rbmD</italic> single or double mutants. This work thus establishes a unique connection between broad spectrum <italic>O</italic>-linked protein glycosylation and the efficacy of type II-dependent protein secretion critical to the pathogen’s lifecycle.</p>