Functional Characterization and Transcriptional Analysis of clpP of Xanthomonas campestris pv. campestris
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Functional Characterization and Transcriptional Analysis of clpP of Xanthomonas campestris pv. campestris Chih‑En Li1 · Chao‑Tsai Liao1 · Hsueh‑Hsia Lo1 · Yi‑Min Hsiao1 Received: 24 February 2020 / Accepted: 15 June 2020 © Springer Science+Business Media, LLC, part of Springer Nature 2020
Abstract The caseinolytic protease (Clp) system is essential for survival under stress conditions and for virulence in several pathogenic bacteria. Xanthomonas campestris pv. campestris (Xcc) is a plant pathogen which causes black rot disease in crucifers. In this study, the Xcc clpP gene which is annotated to encode the proteolytic core of Clp was characterized. Mutation of clpP resulted in susceptibility to high temperature and puromycin stresses. Site-directed mutagenesis revealed that S105, H130, and D179 are critical amino acid residues for ClpP function in puromycin tolerance. Inactivation of clpP also revealed an attenuation of virulence on the host plant and a reduction in the production of extracellular cellulase, mannanase, pectinase, and protease. The affected phenotypes of the clpP mutant could be complemented to wild-type levels by the intact clpP gene. Transcriptional analysis revealed that expression of clpP is induced under heat shock condition.
Introduction Pathogenic bacteria often encounter different stresses exerted by the external environment. These stresses include antimicrobial chemicals, changes in osmolarity, pH, and temperature [1]. To conquer harmful situations, bacterial cells are equipped with various mechanisms to enable them to adapt to heterogeneous environments. Caseinolytic protease (Clp) system plays an essential role in protein quality control and stress management [2]. In addition to its involvement in protein homeostasis and stress tolerance, the Clp protein has a wide range of functions, such as expression of pathogenicity factor and regulation of developmental process in bacteria [2, 3]. The members of the Clp family composed of two functional units: proteolytic core (ClpP and ClpQ) and ATPaseactive chaperone rings (ClpA, ClpC, ClpE, ClpX, and ClpY) [4]. The ClpP protease can interact with different chaperons, namely ClpA, ClpC, ClpE, and ClpX; while ClpQ associates with ClpY to form active proteolytic machinery [4]. The ClpP is a conserved protein that is present in nearly all sequence eubacterial genomes [2]. ClpP is a serine protease * Yi‑Min Hsiao [email protected] 1
Department of Medical Laboratory Science and Biotechnology, Central Taiwan University of Science and Technology, Taichung 406, Taiwan
characterized by a unique arrangement of the active site triad, Ser-His-Asp and is found to be involved in the proteolysis of damaged and misfolded proteins, regulatory proteins, and ribosome-stalled proteins in several species [5]. In addition, ClpP is important for the degradation of proteins involved in biofilm formation, cell motility, heat stress response, metabolism, nutrient starvation, and stationary phase adaptation [5]. In a number of pathogenic bacteria, ClpP function play
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