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LsSpt23p is a regulator of triacylglycerol synthesis in the oleaginous yeast Lipomyces starkeyi

  • Applied Genetics and Molecular Biotechnology
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Abstract

The oleaginous yeast Lipomyces starkeyi has considerable potential in industrial application, since it can accumulate a large amount of triacylglycerol (TAG), which is produced from sugars under nitrogen limitation condition. However, the regulation of lipogenesis in L. starkeyi has not been investigated in depth. In this study, we compared the genome sequences of wild-type and mutants with increased TAG productivity, and identified a regulatory protein, LsSpt23p, which contributes to the regulation of TAG synthesis in L. starkeyi. L. starkeyi mutants overexpressing LsSPT23 had increased TAG productivity compared with the wild-type strain. Quantitative real-time PCR analysis showed that LsSpt23p upregulated the expression of GPD1, which encodes glycerol 3-phosphate dehydrogenase; the Kennedy pathway genes SCT1, SLC1, PAH1, DGA1, and DGA2; the citrate-mediated acyl-CoA synthesis pathway-related genes ACL1, ACL2, ACC1, FAS1, and FAS2; and OLE1, which encodes ∆9 fatty acid desaturase. Chromatin immunoprecipitation-quantitative PCR assays indicated that LsSpt23p acts as a direct regulator of SLC1 and PAH1, all the citrate-mediated acyl-CoA synthesis pathway–related genes, and OLE1. These results indicate that LsSpt23p regulates TAG synthesis. Phosphatidic acid is a common substrate of phosphatidic acid phosphohydrolase, which is used for TAG synthesis, and phosphatidate cytidylyltransferase 1 for phospholipid synthesis in the Kennedy pathway. LsSpt23p directly regulated PAH1 but did not affect the expression of CDS1, suggesting that the preferred route of carbon is the Pah1p-mediated TAG synthesis pathway under nitrogen limitation condition. The present study contributes to understanding the regulation of TAG synthesis, and will be valuable in future improvement of TAG productivity in oleaginous yeasts.

Key points

LsSpt23p was identified as a positive regulator of TAG biosynthesis

LsSPT23 overexpression enhanced TAG biosynthesis gene expression and TAG production

LsSPT23 M1108T overexpression mutant showed fivefold higher TAG production than control

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Data availability

The raw data supporting the conclusion of this article will be made available by the authors, without undue reservation.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

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Funding

This research was partially funded by the New Energy and Industrial Technology Development Organization (NEDO), “Development of Production Techniques for Highly Functional Biomaterials Using Smart Cells of Plants and Other Organisms (Smart Cell Project)”, “Development of bio-based production technology to accelerate Carbon Recycling”, and JSPS KAKENHI (Grant Number 22H02250).

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Authors and Affiliations

  1. Department of Applied Life Sciences, Niigata University of Pharmacy and Applied Life Sciences, 265-1 Higashijima, Akiha-Ku, Niigata, 956-8603, Japan

    Hiroaki Takaku, Haruka Kazama, Rikako Sato, Satoshi Ara & Harutake Yamazaki

  2. Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, 744 Motooka, Nishi-Ku, Fukuoka, 819-0395, Japan

    Kazuki Mori, Kosuke Tashiro & Satoru Kuhara

  3. Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Toyohira-Ku, Sapporo, 062-8517, Japan

    Koji Ishiya

  4. Bioproduction Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba, Ibaraki, 305-8566, Japan

    Tomohiko Matsuzawa, Katsuro Yaoi & Sachiyo Aburatani

  5. Research Institute for Creating the Future, FUJI OIL HOLDINGS INC, 4-3 Kinunodai, Tsukubamirai-Shi, Ibaraki, 300-2497, Japan

    Hideo Araki

  6. Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan

    Yosuke Shida & Wataru Ogasawara

  7. Department of Applied Biological Science, Faculty of Agriculture, Kagawa University, 2393 Ikenobe, Miki-Cho, Kagawa, Kita-Gun, 761-0795, Japan

    Tomohiko Matsuzawa

  8. Department of Science of Technology Innovation, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan

    Wataru Ogasawara

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  1. Hiroaki Takaku
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  2. Haruka Kazama
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Contributions

Conceptualization, H.T., S.K., H.Y., and S. Abratani; formal analysis, H.T., H.K., R.S., K.M., S. Ara, and K.I.; investigation, H.T., H.K., R.S., K.M., S. Ara, K.I., T.M., K.Y., H.A., Y.S., W.O., K.T., S.K., and H.Y.; writing—original draft preparation, H.T., K.I. and S. Abratani; writing—review and editing, H.T., T.M., K.Y., H.A., Y.S., W.O., K.T., H.Y., and S. Abratani; supervision, H.T. and S. Abratani; project administration, H.T. and S. Abratani; funding acquisition, H.T., K.Y., S.K., and S.Abratani. All authors have read and agreed to the published version of the manuscript.

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Correspondence to Hiroaki Takaku.

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Takaku, H., Kazama, H., Sato, R. et al. LsSpt23p is a regulator of triacylglycerol synthesis in the oleaginous yeast Lipomyces starkeyi. Appl Microbiol Biotechnol 107, 1269–1284 (2023). https://doi.org/10.1007/s00253-023-12361-2

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