Construction of CRISPR-Cas9 genome editing platform for white-rot fungus Cerrena unicolor BBP6 and its effects on extracellular ligninolytic enzyme biosynthesis

White-rot fungi (WRF) produce extracellular ligninolytic enzymes such as laccase and manganese-dependent peroxidase (MnP). They have a remarkable potential in degrading a wide range of pollutants including polycyclic aromatic hydrocarbons and synthetic dyes. However, the lack of WRF genetic manipulation methods limits their further applications. Cerrena unicolor BBP6 is a wild-type WRF for laccase and MnP hyper production. This study is the first attempt to develop a genome editing platform for the genus Cerrena sp. based on CRISPR-Cas9 technology. Firstly, the pyrG gene in C. unicolor BBP6 was successfully disrupted and used as an effective screening marker. Subsequently, two genes, LacA and mnp3, which respectively encode a laccase and a MnP protein in strain BBP6, were disrupted. The laccase activity of the LacA mutant strain was reduced by 53.5% compared to the wild-type strain while the MnP activity of the mnp3 mutant strain was reduced by 64.7%. Our study demonstrated that the CRISPR-Cas9 genome-editing platform for C. unicolor BBP6 was successfully established, and it was effective on perturbation of ligninolytic enzyme biosynthesis by this fungus.

Automated summary

This study successfully developed a CRISPR-Cas9 genome editing platform for Cerrena unicolor BBP6. By disrupting specific genes, the laccase and MnP activities were reduced in the strain. This research is important for further understanding of ligninolytic enzyme biosynthesis in this fungus.