Validated Growth Rate-Dependent Regulation of Lipid Metabolism in Yarrowia lipolytica

Abstract

Given the strong potential of Yarrowia lipolytica to produce lipids for use as renewable fuels and oleochemicals, it is important to gain in-depth understanding of the molecular mechanism underlying its lipid accumulation. As cellular growth rate affects biomass lipid content, we performed a comparative proteomic analysis of Y. lipolytica grown in nitrogen-limited chemostat cultures at different dilution rates. After confirming the correlation between growth rate and lipid accumulation, we were able to identify various cellular functions and biological mechanisms involved in oleaginousness. Inspection of significantly up- and downregulated proteins revealed nonintuitive processes associated with lipid accumulation in this yeast. This included proteins related to endoplasmic reticulum (ER) stress, ER–plasma membrane tether proteins, and arginase. Genetic engineering of selected targets validated that some genes indeed affected lipid accumulation. They were able to increase lipid content and were complementary to other genetic engineering strategies to optimize lipid yield.

Urea is a drop-in nitrogen source alternative to ammonium sulphate in Yarrowia lipolytica

“…The strain OKYL049 is a genetically engineered obese strain carrying DGA1 overexpression (YALI1_E38810g) and are1 deletion (YALI1_F09747g). 23 The strain JFYL007, 24 also referred to as Q4 strain, is incapable of synthesizing the storage lipid triacylglycerol or sterol esters due to the deletion of four genes, Δare1 (YALI1_F09747g), Δlro1 (YALI1_E20049g), Δdga1 (YALI1_E38810g), and Δdga2 (YALI1_D10264g). 5 OKYL029 25 has no modifications of the lipid production and displays a wild-type lipid phenotype.…”

“… 25 N/A OKYL049 Konzock et al. 24 N/A JFYL007 Poorinmohammad et al. 23 N/A Software and algorithms DASware® control 5 version 5.30 Eppendorf https://www.eppendorf.com/gb-en/eShop-Products/Bioprocess/Bioprocess-Software/DASware-control-p-PF-133797 bcl2fastq_v2.20.0.422 Illumina https://support.illumina.com/downloads/bcl2fastq-conversion-software-v2-20.html RNAseq pipeline Nf-core https://github.com/nf-core/rnaseq RStudio version 4.2.1 RStudio https://www.rstudio.com OmicsBox version 2.0.24 BioBam https://www.biobam.com/omicsbox/ Other DASGIP® Process Computer Eppendorf 76DGPCR DASGIP® Bioblock Stirrer Vessels: 1 L Vessel Eppendorf 76SR0700ODLS DASGIP® Bioblock Eppendorf 76DGTBLOCK DASGIP® MX Modules for TMFC Gas Mixing Eppendorf 76DGMX44H DASGIP® MP8 Multi Pump Modules Eppendorf 76DGMP8X DASGIP® TC4SC4 for Temperature and Agitation Control Eppendorf 76DGTC4SC4B DASGIP® PH4PO4 for Monitoring of pH, DO, Redox and/or Level Eppendorf 76DGPH4PO4 DASGIP® GA for Exhaust Analysis Eppendorf 76DGGA1EX DO sensors Eppendorf 78108026 pH sensors Eppendorf …”

“…JFYL007 (ST6512 Δmhy1 Δare1 Δlro1 Δdga1 Δdga2) is a low lipid accumulating strain carrying a deletion of ARE1 to abolish sterol ester formation, a deletion of LRO1 (triacylglycerol formation with phospholipids as acyl donors), a deletion of DGA1 (acyl-CoA dependent triacylglycerol synthase), and a deletion of DGA2 (member of the type 1 acyl-CoA:diacylglycerol acyltransferase family) to decrease TAG accumulation; to prevent hyphae formation, MHY1 is also deleted. 24 Since our strains are derived from the W29/CLIB89 background, our gene annotation follows the YALI1 system, but translation into YALI0/CLIB122) can be done with the S2 table of. 39 …”

Abolishing storage lipids induces protein misfolding and stress responses inYarrowia lipolytica

“…To prevent hyphae formation, the MHY1 gene is also deleted in this strain. The Q4 strain JFYL007 (ST6512 ∆mhy1 ∆are1 ∆lro1 ∆dga1 ∆dga2) is a low-lipid accumulating strain that carries deletions of the ARE1, LRO1, DGA1, and DGA2 genes to decrease TAG accumulation, and a deletion of the MHY1 gene to prevent hyphae formation 18 . The gene annotation for these strains follows the YALI1 system, but translation into YALI0/CLIB122 can be done with the S2 table of 48 .…”

“…A Y. lipolytica strain carrying DGA1, DGA2, LRO1, and ARE1 deletions (referred to as Q4 strain) lacks storage lipid accumulation 13,18 . While abolishing storage lipid accumulation is a justifiable strategy to increase the production of desired chemicals in Y. lipolytica, the consequences on cell physiology and regulation have not been investigated yet.…”