Deletion of MHY1 abolishes hyphae formation in Yarrowia lipolytica without negative effects on stress tolerance

Abstract

There is a need for development of sustainable production processes for production of fats/ oils and lipid derived chemicals. The dimorphic oleaginous yeast Yarrowia lipolytica is a promising organism for conversion of biomass hydrolysate to lipids, but in many such processes hyphae formation will be problematic. We have therefore constructed and compared the performance of strains carrying deletions in several published gene targets suggested to abolish hyphae formation (MHY1, HOY1 and CLA4). The MHY1-deletion was the only of the tested strains which did not exhibit hyphae formation under any of the conditions tested. The MHY1-deletion also had a weak positive effect on lipid accumulation without affecting the total fatty acid composition, irrespective of the nitrogen source used. MHY1 has been suggested to constitute a functional homolog of the stress responsive transcription factors MSN2/4 in Saccharomyces cerevisiae, the deletion of which are highly stress sensitive. However, the deletion of MHY1 displayed only minor difference on survival of a range of acute or long term stress and starvation conditions. We conclude that the deletion of MHY1 in Y.lipolytica is a reliable way of abolishing hyphae formation with few detectable negative side effects regarding growth, stress tolerance and lipid accumulation and composition.

Altering the fatty acid profile of Yarrowia lipolytica to mimic cocoa butter by genetic engineering of desaturases

“…To establish a food oil production strain, it is crucial to alter its fatty acid profile while maintaining high lipid titers. We first compared the fatty acid composition of a non-engineered strain (OKYL029, mhy1Δ to avoid undesired pseudohyphal growth) [ 20 ] with a derived strain engineered for lipid production (OKYL049—deletion of sterol ester production (are1Δ) and increased TAG production (DGA1 overexpression)) [ 21 ], leading to a 2.5-fold increase in lipid content. We noted that OKYL049 displayed a decrease in linoleic acid (C18:2) and an increase in stearic acid (C18:0), both of which properties are desirable for CBE-production (Fig.…”

“…Displayed is the mean and standard error of n ≥ 4 replicates. Data for OKYL029 from [ 20 ]. Yl Y. lipolytica , Gt G. trabeum , Rt R. toruloides , Aa A. adeninivorans , WT wild type, Δ deletion …”

“…ATCC20460/CBS7504). The initial starting strain of this study was OKYL029 which carries the MHY1 deletion to prevent filamentous growth [ 20 ].…”

Tolerance of Yarrowia lipolytica to inhibitors commonly found in lignocellulosic hydrolysates

“…OKYL049 is an obese strain in which overexpression of DGA1 was first introduced, followed by deletion of ARE1 to increase TAG accumulation and abolish sterol ester formation. MHY1 was deleted to prevent hyphae formation [ 40 ]. The strains SZYL002 (overexpressing XK and XDH) and SZYL004 (overexpressing XK, XDH, and XR) are derived from OKYL049 (Table 1 ).…”

“…Y. lipolytica strains were cultivated in LPU media and samples were taken after 24, 48, 72 and 96 h for fatty acid methyl ester (FAME) extraction to measure cellular lipid content. The protocol used was previously described [ 40 , 43 ]. In short, 100 μL of cell culture was spun down, the supernatant was discarded, and the cells were washed twice with 1 mL water.…”