High-dose rapamycin exerts a temporary impact on T. reesei RUT-C30 through gene trFKBP12

Abstract

Background Knowledge with respect to regulatory systems for cellulase production is prerequisite for exploitation of such regulatory networks to increase cellulase production, improve fermentation efficiency and reduce the relevant production cost. The target of rapamycin (TOR) signaling pathway is considered as a central signaling hub coordinating eukaryotic cell growth and metabolism with environmental inputs. However, how and to what extent the TOR signaling pathway and rapamycin are involved in cellulase production remain elusive. Result At the early fermentation stage, high-dose rapamycin (100 μM) caused a temporary inhibition effect on cellulase production, cell growth and sporulation of Trichoderma reesei RUT-C30 independently of the carbon sources, and specifically caused a tentative morphology defect in RUT-C30 grown on cellulose. On the contrary, the lipid content of T. reesei RUT-C30 was not affected by rapamycin. Accordingly, the transcriptional levels of genes involved in the cellulase production were downregulated notably with the addition of rapamycin. Although the mRNA levels of the putative rapamycin receptor trFKBP12 was upregulated significantly by rapamycin, gene trTOR (the downstream effector of the rapamycin–FKBP12 complex) and genes associated with the TOR signaling pathways were not changed markedly. With the deletion of gene trFKBP12, there is no impact of rapamycin on cellulase production, indicating that trFKBP12 mediates the observed temporary inhibition effect of rapamycin. Conclusion Our study shows for the first time that only high-concentration rapamycin induced a transient impact on T. reesei RUT-C30 at its early cultivation stage, demonstrating T. reesei RUT-C30 is highly resistant to rapamycin, probably due to that trTOR and its related signaling pathways were not that sensitive to rapamycin. This temporary influence of rapamycin was facilitated by gene trFKBP12. These findings add to our knowledge on the roles of rapamycin and the TOR signaling pathways play in T. reesei.

Glutamine involvement in nitrogen regulation of cellulase production in fungi

“…By contrast, the mRNA level of trFKBP12 was increased remarkably. This was similar to what we found previously that genes related to TOR signaling pathways were not changed significantly but trFKBP12 was significantly upregulated by rapamycin in T. reesei grown on cellulose condition [ 41 ]. Nevertheless, there were still several genes in the TOR signaling pathways affected by glutamine, including the upregulated expression of the cellular receptor of rapamycin trFKBP12, and the downregulated expression of transcription initiation factor Rrn3, stress-responding transcription factors Msn2 and Msn4, as well as the primary amino acid receptor Ssy1.…”

“…8 B). For genes associated with TOR signal pathways including 15 genes for ribosome biogenesis, 8 genes for cell cycle/growth, 25 genes for nutrient uptake, 1 gene for stress, 5 genes for lipid metabolism, 6 genes for cell wall integrity, and 5 genes for autophagy [ 41 ] (Additional file 6 : Table S6), only four genes (Rrn3, M419DRAFT_136493, M419DRAFT_91864 and M419DRAFT_97298) were differentially expressed (Fig. 8 C).…”

“…By contrast, the cellular receptor of rapamycin trFKBP12 was noticeably upregulated by 2 4.10 (17.14)-fold with 2.92 g/L glutamine. To see whether trFKBP12 was involved in the negative impact of glutamine on cellulase production, the trFKBP12 deletion strain ΔtrFKBP12 engineered in our previous study [ 41 ] was treated with glutamine. Similar to the deletion of genes ooc1 and ooc2 , the knockout of trFKBP12 exacerbated the cellulase depression influence of glutamine (Fig.…”

“…(Hemi)cellulase activity assay, confocal imaging, spore counting, RNA-seq analysis, and RT-PCR were performed as described in our previous research [ 41 , 65 ]. T. reesei RUT-C30 cultured for 24 h with or without 2.92 g/L glutamine were utilized for RNA-seq.…”

Cellulase production and efficient saccharification of biomass by a new mutant Trichoderma afroharzianum MEA-12

“…This matched the superior power of the cellulase production model strain T. reesei Rut-C30. T. reesei Rut-C30 has been developed for decades and is currently an industrial strain with a relatively complete cellulase system [ 28 ]. The comparable protein secretion level between MEA-12 and T. reesei Rut-C30 and the higher production of β-glucosidase by MEA-12 (Table 2 ) suggest its great potential for biotechnological application, especially for lignocellulose biomass hydrolysis.…”