The serine/threonine protein kinase MpSTE1 directly governs hyphal branching in Monascus spp

Yali Duan; Xizhu Chen; Tingya Wang; Mu Li

doi:10.1007/s00253-024-13093-7

The serine/threonine protein kinase MpSTE1 directly governs hyphal branching in Monascus spp

Appl Microbiol Biotechnol. 2024 Mar 6;108(1):255. doi: 10.1007/s00253-024-13093-7.

Authors

Yali Duan^#^{1

2}, Xizhu Chen^#^{1

2}, Tingya Wang^{1

2}, Mu Li^{3

4}

Affiliations

¹ Key Laboratory of Environment Correlative Dietology, College of Food Science and Technology, Hubei International Scientific and Technological Cooperation Base of Traditional Fermented FoodsHuazhong Agricultural UniversityHubei Province, Wuhan, 430070, China.
² College of Food Science and Technology, Huazhong Agricultural University, Hubei Province, Wuhan, 430070, China.
³ Key Laboratory of Environment Correlative Dietology, College of Food Science and Technology, Hubei International Scientific and Technological Cooperation Base of Traditional Fermented FoodsHuazhong Agricultural UniversityHubei Province, Wuhan, 430070, China. limu@mail.hzau.edu.cn.
⁴ College of Food Science and Technology, Huazhong Agricultural University, Hubei Province, Wuhan, 430070, China. limu@mail.hzau.edu.cn.

^# Contributed equally.

Abstract

Monascus spp. are commercially important fungi due to their ability to produce beneficial secondary metabolites such as the cholesterol-lowering agent lovastatin and natural food colorants azaphilone pigments. Although hyphal branching intensively influenced the production of these secondary metabolites, the pivotal regulators of hyphal development in Monascus spp. remain unclear. To identify these important regulators, we developed an artificial intelligence (AI)-assisted image analysis tool for quantification of hyphae-branching and constructed a random T-DNA insertion library. High-throughput screening revealed that a STE kinase, MpSTE1, was considered as a key regulator of hyphal branching based on the hyphal phenotype. To further validate the role of MpSTE1, we generated an mpSTE1 gene knockout mutant, a complemented mutant, and an overexpression mutant (OE::mpSTE1). Microscopic observations revealed that overexpression of mpSTE1 led to a 63% increase in branch number while deletion of mpSTE1 reduced the hyphal branching by 68% compared to the wild-type strain. In flask cultures, the strain OE::mpSTE1 showed accelerated growth and glucose consumption. More importantly, the strain OE::mpSTE1 produced 9.2 mg/L lovastatin and 17.0 mg/L azaphilone pigments, respectively, 47.0% and 30.1% higher than those of the wild-type strain. Phosphoproteomic analysis revealed that MpSTE1 directly phosphorylated 7 downstream signal proteins involved in cell division, cytoskeletal organization, and signal transduction. To our best knowledge, MpSTE1 is reported as the first characterized regulator for tightly regulating the hyphal branching in Monascus spp. These findings significantly expanded current understanding of the signaling pathway governing the hyphal branching and development in Monascus spp. Furthermore, MpSTE1 and its analogs were demonstrated as promising targets for improving production of valuable secondary metabolites. KEY POINTS: • MpSTE1 is the first characterized regulator for tightly regulating hyphal branching • Overexpression of mpSTE1 significantly improves secondary metabolite production • A high-throughput image analysis tool was developed for counting hyphal branching.

Keywords: Filamentous fungi; Hyphal branching; Protein kinase; Secondary metabolites; Signaling pathway.

MeSH terms

Artificial Intelligence
Hyphae*
Lovastatin
Monascus* / genetics
Protein Serine-Threonine Kinases
Serine
Threonine

Substances

Protein Serine-Threonine Kinases
Lovastatin
Threonine
Serine