Hansenula polymorpha Hac1p Is Critical to Protein N-Glycosylation Activity Modulation, as Revealed by Functional and Transcriptomic Analyses

ABSTRACT Aggregation of misfolded protein in the endoplasmic reticulum (ER) induces a cellular protective response to ER stress, the unfolded protein response (UPR), which is mediated by a basic leucine zipper (bZIP) transcription factor, Hac1p/Xbp1. In this study, we identified and studied the molecular functions of a HAC1 homolog from the thermotolerant yeast Hansenula polymorpha (Hp HAC1 ). We found that the Hp HAC1 mRNA contains a nonconventional intron of 177 bp whose interaction with the 5′ untranslated region is responsible for the translational inhibition of the Hp HAC1 mRNA. The H. polymorpha hac1 -null (Hp hac1 Δ) mutant strain grew slowly, even under normal growth conditions, and was less thermotolerant than the wild-type (WT) strain. The mutant strain was also more sensitive to cell wall-perturbing agents and to the UPR-inducing agents dithiothreitol (DTT) and tunicamycin (TM). Using comparative transcriptome analysis of the WT and Hp hac1 Δ strains treated with DTT and TM, we identified Hp HAC1 -dependent core UPR targets, which included genes involved in protein secretion and processing, particularly those required for N -linked protein glycosylation. Notably, different glycosylation and processing patterns of the vacuolar glycoprotein carboxypeptidase Y were observed in the WT and Hp hac1 Δ strains. Moreover, overexpression of active HpHac1p significantly increased the N -linked glycosylation efficiency and TM resistance. Collectively, our results suggest that the function of HpHac1p is important not only for UPR induction but also for efficient glycosylation in H. polymorpha.