AUTHOR=Zhang Zhiping , Li Yingbin , Luo Laixin , Hao Jianjun , Li Jianqiang 

TITLE=Characterization of cmcp Gene as a Pathogenicity Factor of Ceratocystis manginecans

JOURNAL=Frontiers in Microbiology

VOLUME=11

YEAR=2020

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2020.01824

DOI=10.3389/fmicb.2020.01824

ISSN=1664-302X

ABSTRACT=<p><italic>Ceratocystis manginecans</italic> causes mango wilt with significant economic losses. In the infection court, cerato-platanin (CP) family proteins (CPPs) are believed to involve in pathogenesis but has not been determined in <italic>C. manginecans</italic>. To confirm this function, a CP protein (CmCP) of <italic>C. manginecans</italic> was characterized in this study. A protoplast of <italic>C. manginecans</italic> was prepared by treating its mycelia with driselase and lysing enzymes. The <italic>cmcp</italic> gene was edited using CRISPR/Cas-U6-1 expression vectors in 60% PEG and 50 μg/mL hygromycin B in the medium, resulting in mutants with <italic>cmcp</italic> deletion (Δ<italic>cmcp</italic>). A complemented mutant (Δ<italic>cmcp</italic>-C) was obtained by transforming <italic>cmcp</italic> to Δ<italic>cmcp</italic>. Both Δ<italic>cmcp</italic> and Δ<italic>cmcp</italic>-C were characterized by comparing them with a wild-type strain on morphology, mycelial growth, conidial production and pathogenicity. Additionally, <italic>cmcp</italic> was transformed and expressed in <italic>Pichia pastoris</italic>, and the derived recombinant protein CmCP caused a severe necrosis on <italic>Nicotiana tabacum</italic> leaves. CmCP-treated plant leaves showed symptoms of hypersensitive response including electrolyte leakage, reactive oxygen species generation and overexpression of defense-related genes <italic>PR-1</italic>, <italic>PAD3</italic>, <italic>ERF1</italic>, <italic>HSR203J</italic>, and <italic>HIN1</italic>. All those results suggested that <italic>cmcp</italic> gene was required for the growth development of <italic>C. manginecans</italic> and functioned as a major pathogenicity factor in mango infection.</p>