Dataset Information

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 (HpHAC1). We found that the HpHAC1 mRNA contains a nonconventional intron of 177 bp whose interaction with the 5' untranslated region is responsible for the translational inhibition of the HpHAC1 mRNA. The H. polymorpha hac1-null (Hphac1?) 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 Hphac1? strains treated with DTT and TM, we identified HpHAC1-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 Hphac1? 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.

SUBMITTER: Moon HY

PROVIDER: S-EPMC4579441 | biostudies-literature | 2015 Oct

REPOSITORIES: biostudies-literature

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

Moon Hye-Yun HY Cheon Seon Ah SA Kim Hyunah H Agaphonov M O MO Kwon Ohsuk O Oh Doo-Byoung DB Kim Jeong-Yoon JY Kang Hyun Ah HA

Applied and environmental microbiology 20150731 20

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 (HpHAC1). We found that the HpHAC1 mRNA contains a nonconventional intron of 177 bp whose interaction with the 5' untranslat ...[more]

PMID: 26231645

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Project description:BACKGROUND: Methylotrophic yeast species (e.g. Hansenula polymorpha, Pichia pastoris) can grow on methanol as sole source of carbon and energy. These organisms are important cell factories for the production of recombinant proteins, but are also used in fundamental research as model organisms to study peroxisome biology. During exponential growth on glucose, cells of H. polymorpha typically contain a single, small peroxisome that is redundant for growth while on methanol multiple, enlarged peroxisomes are present. These organelles are crucial to support growth on methanol, as they contain key enzymes of methanol metabolism.In this study, changes in the transcriptional profiles during adaptation of H. polymorpha cells from glucose- to methanol-containing media were investigated using DNA-microarray analyses. RESULTS: Two hours after the shift of cells from glucose to methanol nearly 20% (1184 genes) of the approximately 6000 annotated H. polymorpha genes were significantly upregulated with at least a two-fold differential expression. Highest upregulation (> 300-fold) was observed for the genes encoding the transcription factor Mpp1 and formate dehydrogenase, an enzyme of the methanol dissimilation pathway. Upregulated genes also included genes encoding other enzymes of methanol metabolism as well as of peroxisomal beta-oxidation.A moderate increase in transcriptional levels (up to 4-fold) was observed for several PEX genes, which are involved in peroxisome biogenesis. Only PEX11 and PEX32 were higher upregulated. In addition, an increase was observed in expression of the several ATG genes, which encode proteins involved in autophagy and autophagy processes. The strongest upregulation was observed for ATG8 and ATG11.Approximately 20% (1246 genes) of the genes were downregulated. These included glycolytic genes as well as genes involved in transcription and translation. CONCLUSION: Transcriptional profiling of H. polymorpha cells shifted from glucose to methanol showed the expected downregulation of glycolytic genes together with upregulation of the methanol utilisation pathway. This serves as a confirmation and validation of the array data obtained. Consistent with this, also various PEX genes were upregulated. The strong upregulation of ATG genes is possibly due to induction of autophagy processes related to remodeling of the cell architecture required to support growth on methanol. These processes may also be responsible for the enhanced peroxisomal beta-oxidation, as autophagy leads to recycling of membrane lipids. The prominent downregulation of transcription and translation may be explained by the reduced growth rate on methanol (td glucose 1 h vs td methanol 4.5 h).

Dataset Information

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

Publications

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

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