Constitutive Overexpression\nof CRZ1 in Trichoderma reesei Reveals\nIts Ability to Enhance\nRecombinant Lipase Production

The filamentous fungus Trichoderma reesei is extensively utilized in the realm of recombinant protein expression\nowing to its well-established protein production systems. However,\nthe potential for efficient and convenient protein production in T. reesei has not been fully harnessed. To further\nincrease the production of recombinant lipase Candida\nantarctica lipase B (CalB), we overexpressed seven\ntranscription activators and found that overexpression of the calcineurin\nCRZ1 could significantly enhance CalB production by 2.08-fold, and\nthen through employing a synthetic strong promoter Pcbh7IR and knocking\nout <i>ace1</i>, a competitor of <i>crz1</i>,\nCalB production was further increased 4.2-fold. Furthermore, we found\nthat increasing the Ca<sup>2+</sup> concentration enabled the enhancement\nof CalB production in the CRZ1-overexpressing strains. Additionally,\ntranscription analysis revealed a significant upregulation of calcium-related\npathway genes and ER chaperone genes in T. reesei, revealing that overexpression of CRZ1 can increase the production\nof the recombinant protein by activation of the calcium signaling\npathway. In this study, by systematically manipulating the recombinant\nprotein expression module and its regulatory transcription activators,\nwe found a positive effect of <i>crz1</i>-mediated calcium\nchannels in boosting the expression of the recombinant protein.