Abstract
Objective
The regulator in glycerol repression of Pichia pastoris AOX1 promoter (P AOX1 ) is still unclear.
Results
A Cys2His2 zinc finger transcriptional repressor PpNrg1 localized to nucleus and participated in the repression of P AOX1 in P. pastoris in glucose and glycerol. Quantitative real-time PCR revealed that PpNrg1 repressed expression of numerous genes involved in methanol utilization and peroxisome biogenesis in 0.02 % glucose and 1 % (v/v) glycerol. Electrophoretic mobility shift assay and DNase I footprinting assay revealed that PpNrg1 bound to five sites of P AOX1 , including two binding sites of PpMxr1, which is an indispensable activator of P AOX1 in P. pastoris.
Conclusion
Transcriptional repressor PpNrg1 suppresses P AOX1 in glucose and glycerol by directly binding to five sites of P AOX1 , including two binding sites of transcriptional activator PpMxr1.
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Acknowledgments
This work was supported by Chinese National High Technology Research and Development Program (No. 2014AA093501), National Special Fund for State Key Laboratory of Bioreactor Engineering (No. 2060204), and Grants of Young and Middle-aged Leading Science and Technology Innovation Talents from Ministry of Science and Technology of China.
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Supplementary Table 1—Oligonucleotide primers used in this study.
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Wang, X., Cai, M., Shi, L. et al. PpNrg1 is a transcriptional repressor for glucose and glycerol repression of AOX1 promoter in methylotrophic yeast Pichia pastoris . Biotechnol Lett 38, 291–298 (2016). https://doi.org/10.1007/s10529-015-1972-4
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DOI: https://doi.org/10.1007/s10529-015-1972-4