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Background Docosahexaenoic acid (DHA, C22:6) and odd-chain fatty acids (OCFAs, C15:0 and C17:0) have attracted great interest, since they have been widely used in food and therapeutic industries, as well as chemical industry, such as biodiesel production and improvement. The oil-producing heterotrophic microalgae Schizochytrium sp. 31 is one of main DHA-producing strains. Recently, it was found that Schizochytrium can also synthesize OCFAs; however, contents and titers of DHA and OCFAs in Schizochytrium are still low, which limit its practical application. Results In this study, we found that acetyl-CoA carboxylase suffered from a feedback inhibition by C16-CoA in Schizochytrium, and relief of the inhibition resulted in improved both lipid content and the ratio of OCFAs in total fatty acids. Based on this finding, a novel strategy for elevating both DHA and OCFAs contents was established. First, the total lipid accumulation was increased by overexpressing a malic enzyme from Crypthecodinium cohnii to elevate NADPH supply. Second, the inhibition effect on acetyl-CoA carboxylase was relieved by overexpressing a codon-optimized ELO3 gene from Mortierella alpina, which encodes an elongase enzyme responsible for converting C16 into C18 fatty acids. After the above two-step engineering, contents of DHA and OCFAs were increased by 1.39- and 3.30-fold, reaching a level of 26.70 and 25.08% of dry cell weight, respectively, which are the highest contents reported so far for Schizochytrium. The titers of DHA and OCFAs were elevated by 1.08- and 2.57-fold, reaching a level of 3.54 and 3.32 g/L, respectively. Notably, the OCFAs titer achieved was 2.66-fold higher than the highest reported in Escherichia coli (1.25 g/L), implying potential value for industry application. To reveal the potential metabolic mechanism for the enhanced biosynthesis of both DHA and OCFAs, LC–MS metabolomic analysis was employed and the results showed that the pentose phosphate pathway and the glycolysis pathway were strengthened and intracellular propionyl-CoA concentration were also significantly increased in the engineered Schizochytrium, suggesting an increased supply of NADPH, acetyl-CoA, and propionyl-CoA for DHA and OCFAs accumulation. Conclusions The discovery provides a new source of OCFAs production, and proposes a new strategy to improve contents and titers of both DHA and OCFAs in Schizochytrium. These will be valuable for improving commercial potential of Schizochytrium and guiding the engineering strategy in other fatty acids producing heterotrophic microalga. Electronic supplementary material The online version of this article (10.1186/s13068-019-1484-x) contains supplementary material, which is available to authorized users.

Homologs of the velvet protein family are encoded by the ve1, vel2, and vel3 genes in Trichoderma reesei. To test their regulatory functions, the velvet protein-coding genes were disrupted, generating Δve1, Δvel2, and Δvel3 strains. The phenotypic features of these strains were examined to identify their functions in morphogenesis, sporulation, and cellulase expression. The three velvet-deficient strains produced more hyphal branches, indicating that velvet family proteins participate in the morphogenesis in T. reesei. Deletion of ve1 and vel3 did not affect biomass accumulation, while deletion of vel2 led to a significantly hampered growth when cellulose was used as the sole carbon source in the medium. The deletion of either ve1 or vel2 led to the sharp decrease of sporulation as well as a global downregulation of cellulase-coding genes. In contrast, although the expression of cellulase-coding genes of the ∆vel3 strain was downregulated in the dark, their expression in light condition was unaffected. Sporulation was hampered in the ∆vel3 strain. These results suggest that Ve1 and Vel2 play major roles, whereas Vel3 plays a minor role in sporulation, morphogenesis, and cellulase expression.

The lignocellulose degradation capacity of filamentous fungi has been widely studied because of their cellulase hypersecretion. The p24 proteins in eukaryotes serve important functions in this secretory pathway. However, little is known about the functions of the p24 proteins in filamentous fungi. In this study, four p24 proteins were identified in Penicillium oxalicum. Six p24 double-deletion strains were constructed and further studies were carried out with the ΔerpΔpδ strain. The experimental results suggested that Erp and Pδ form a p24 heterodimer in vivo. This p24 heterodimer participates in important morphogenetic events, including sporulation, hyphal growth and lateral branching. The results suggested that the p24 heterodimer mediates protein transport, particularly that of cellobiohydrolase. Analysis of the intracellular proteome revealed that the ΔerpΔpδ double mutant is under secretion stress due to attempts to remove proteins that are jammed in the endomembrane system. These results suggest that the p24 heterodimer participates in morphogenesis and protein transport. Compared with P. oxalicum Δerp, a greater number of cellular physiological pathways were impaired in ΔerpΔpδ. This finding may provide new insights into the secretory pathways of filamentous fungi.

Despite the important role of MAPKs in signal transduction, their functions in the cellulase hyper-producing filamentous fungus Hypocrea jecorina haven't been studied except for the Hog1-like Tmk3. In this work, we constructed and explored the features of H. jecorina Δtmk2 to identify the role of this Slt2-homologous Tmk2. It is suggested from the results that Tmk2 is involved in cell wall integrity, sporulation and cellulase production. Although bearing similar roles in cell wall integrity maintenance, Tmk2 and Tmk3 appear to also have distinct functions: Tmk3 participates in high osmolarity resistance while Tmk2 does not; Tmk2 participates in sporulation but not Tmk3; Tmk3 is involved in promoting cellulase production while Tmk2 is involved in repressing cellulase formation. These studies provide the first insight into the function of Tmk2 in H. jecorina and contribute to understanding the signal transduction processes leading to the regulation of cellulase production in this important cellulase hyper-producer.