We identified the AGS1 and AGS3 genes by their ability to partially complement an ags mutant (RC1707) which is supersensitive to various aminoglycoside antibiotics (J. F. Ernst and R. K. Chan, J. Bacteriol. 163:8–14, 1985).AGS1 is located in proximity to the centromere of chromosome III and encodes a small protein of 88 amino acids. The size of the AGS1 transcript, which in wild-type cells is 1 kb, is reduced to 0.75 kb in mutant RC1707. Disruption of AGS1rendered strains supersensitive to hygromycin B and increased their resistance to vanadate. In addition, ags1Δ strains underglycosylated invertase but had normal carboxypeptidase Y glycosylation, suggesting that Ags1p is required for the elaboration of outer N-glycosyl chains. AGS3 was found to be identical toPHO80 (TUP7), which encodes a cyclin activating the Pho85p protein kinase. Deletion of either PHO80 orPHO85 led to aminoglycoside supersensitivity;pho80Δ ags1Δ strains showed an enhanced-sensitivity phenotype compared to single mutants. pho80 andpho85 mutants were rendered resistant by deletion ofPHO4, indicating that activation of the Pho4p transcription factor is required for increased aminoglycoside sensitivity. Thus, both the Pho80p-Pho85p kinase complex (by Pho4p phosphorylation) and a novel component of the N glycosylation pathway contribute to basal levels of aminoglycoside resistance in Saccharomyces cerevisiae.
In eukaryotes, Dom34 upregulates translation by securing levels of activatable ribosomal subunits. We found that in the yeast Saccharomyces cerevisiae and the human fungal pathogen Candida albicans, Dom34 interacts genetically with Pmt1, a major isoform of protein O-mannosyltransferase. In C. albicans, lack of Dom34 exacerbated defective phenotypes of pmt1 mutants, while they were ameliorated by Dom34 overproduction that enhanced Pmt1 protein but not PMT1 transcript levels. Translational effects of Dom34 required the 5′-UTR of the PMT1 transcript, which bound recombinant Dom34 directly at a CA/AC-rich sequence and regulated in vitro translation. Polysomal profiling revealed that Dom34 stimulates general translation moderately, but that it is especially required for translation of transcripts encoding Pmt isoforms 1, 4 and 6. Because defective protein N- or O-glycosylation upregulates transcription of PMT genes, it appears that Dom34-mediated specific translational upregulation of the PMT transcripts optimizes cellular responses to glycostress. Its translational function as an RNA binding protein acting at the 5′-UTR of specific transcripts adds another facet to the known ribosome-releasing functions of Dom34 at the 3′-UTR of transcripts.
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