Fig. 4: Five strategies used to validate TAG cycling in Y. lipolytica.

a Description of two strategies used to inhibit TAG cycling. Knocking out the TGL4 gene (strategy 1, red line) and overexpressing the PEX10 gene (strategy 2, blue line) decreased the free FA content (dotted line) and increased the acetyl-CoA content (thick line). b C16:0 and C20:5 contents of the engineered strains YIP56, YIP56-ΔTGL4, and YlP56-PEX10. c Description of two strategies to accelerate TAG cycling. Overexpression of the TGL4 gene (strategy 3, green line) and knockout of the PEX10 gene (strategy 4, purple line) increased the free FA content (thick line) and decreased the acetyl-CoA content (dotted line). d C16:0 and C20:5 contents of the engineered strains YIP56, YIP56-TGL4, and YlP56-ΔPEX10. e Description of the inhibition of the expression of the Des12 gene (strategy 5) and the corresponding changes in the C18:1, C18:2, and C20:5 contents. The inducer was added at 168 h, the red line represents the fatty acid content in the presence of the inducer, and the blue line represents the fatty acid content in the absence of the inducer. The red arrow represents the time point of inducer addition. f The abundance of ¹³C-labeled fatty acids. In (b, d–f), the data are the means ± s.d. from three (n = 3) biologically independent replicates.