Trichoderma reesei encodes 11 putative β-glucosidases in genome, which play an important role in cellulase induction and production. However, current understanding does not provide a global perspective on the role and function of these β-glucosidases. This study comprehensively explored the cellular distribution and function of 10 known β-glucosidases (CEL3B, CEL3E, CEL3F, CEL3H, CEL3J, CEL1A, CEL3C, CEL1B, CEL3G and CEL3D) in T. reesei by fusing red fluorescent protein (DsRed) to the C terminus of β-glucosidases. Overexpression of cel3b or cel3g promoted β-glucosidase production, while cel1b overexpression severely inhibited cellulase production on cellulose. Overexpression of cel3b, cel3e, cel3f, cel3h, cel3j, cel3c or cel3g delayed endoglucanase (CMC) production. Four types of cellular distribution patterns were first identified: 1) CEL3B, CEL3E, CEL3F, and CEL3G forming clearly separated protein secretion vesicles in the cytoplasm; 2) CEL3H and CEL3J diffusing the whole endomembrane as well as cell membrane with protein aggregation, like a reticular network; 3) CEL1A and CEL3D in vacuoles; 4) CEL3C in the nuclear. Seven β-glucosidases CEL1A, CEL3B, CEL3E, CEL3F, CEL3G, CEL3H, and CEL3J were proved to be extracellular. Amont them, CEL3B, CEL3E, CEL3F, CEL3H and CEL3G were secreted through tip-directed conventional secretion pathway, and CEL1A via vacuole-mediated one, while CEL3J via an unconventional protein pathway bypassing ER and Golgi. These findings advance our understanding of β-glucosidase properties and secretory pathways in filamentous fungi.

Dissecting cellular function and distribution of β-glucosidases in Trichoderma reesei

ACS Fall 2021

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