AUTHOR=Men Ping , Wang Min , Li Jinda , Geng Ce , Huang Xuenian , Lu Xuefeng 

TITLE=Establishing an Efficient Genetic Manipulation System for Sulfated Echinocandin Producing Fungus Coleophoma empetri

JOURNAL=Frontiers in Microbiology

VOLUME=12

YEAR=2021

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2021.734780

DOI=10.3389/fmicb.2021.734780

ISSN=1664-302X

ABSTRACT=<p>Micafungin is an important echinocandin antifungal agent for the treatment of invasive fungal infections. In industry, micafungin is derived from the natural product FR901379, which is a non-ribosomal cyclic hexapeptide produced by the filamentous fungus <italic>Coleophoma empetri</italic>. The difficulty of genetic manipulation in <italic>C. empetri</italic> restricts the clarification of FR901379 biosynthetic mechanism. In this work, we developed an efficient genetic manipulation system in the industrial FR901379-producing strain <italic>C. empetri</italic> MEFC009. Firstly, a convenient protoplast-mediated transformation (PMT) method was developed. Secondly, with this transformation method, the essential genetic elements were verified. Selectable markers <italic>hph</italic>, <italic>neo</italic>, and <italic>nat</italic> can be used for the transformation, and promotors <italic>Ppgk</italic>, <italic>PgpdA</italic>, and <italic>PgpdAt</italic> are functional in <italic>C. empetri</italic> MEFC009. Thirdly, the frequency of homologous recombination was improved from 4 to 100% by deleting the <italic>ku80</italic> gene, resulting in an excellent chassis cell for gene-targeting. Additionally, the advantage of this genetic manipulation system was demonstrated in the identification of the polyketide synthase (PKS) responsible for the biosynthesis of dihydroxynapthalene (DHN)-melanin. This genetic manipulation system will be a useful platform for the research of FR901379 and further genome mining of secondary metabolites in <italic>C. empetri</italic>.</p>