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1.  The Saccharomyces cerevisiae Hot1p regulated gene YHR087W (HGI1) has a role in translation upon high glucose concentration stress 
BMC Molecular Biology  2012;13:19.
While growing in natural environments yeasts can be affected by osmotic stress provoked by high glucose concentrations. The response to this adverse condition requires the HOG pathway and involves transcriptional and posttranscriptional mechanisms initiated by the phosphorylation of this protein, its translocation to the nucleus and activation of transcription factors. One of the genes induced to respond to this injury is YHR087W. It encodes for a protein structurally similar to the N-terminal region of human SBDS whose expression is also induced under other forms of stress and whose deletion determines growth defects at high glucose concentrations.
In this work we show that YHR087W expression is regulated by several transcription factors depending on the particular stress condition, and Hot1p is particularly relevant for the induction at high glucose concentrations. In this situation, Hot1p, together to Sko1p, binds to YHR087W promoter in a Hog1p-dependent manner. Several evidences obtained indicate Yhr087wp’s role in translation. Firstly, and according to TAP purification experiments, it interacts with proteins involved in translation initiation. Besides, its deletion mutant shows growth defects in the presence of translation inhibitors and displays a slightly slower translation recovery after applying high glucose stress than the wild type strain. Analyses of the association of mRNAs to polysome fractions reveals a lower translation in the mutant strain of the mRNAs corresponding to genes GPD1, HSP78 and HSP104.
The data demonstrates that expression of Yhr087wp under high glucose concentration is controlled by Hot1p and Sko1p transcription factors, which bind to its promoter. Yhr087wp has a role in translation, maybe in the control of the synthesis of several stress response proteins, which could explain the lower levels of some of these proteins found in previous proteomic analyses and the growth defects of the deletion strain.
PMCID: PMC3441895  PMID: 22720784
Saccharomyces cerevisiae; High glucose osmotic stress; Gene YHR087W; Gene expression; Translation; Hot1p; Hog1p; Polysomes
2.  Saccharomyces cerevisiae Signature Genes for Predicting Nitrogen Deficiency during Alcoholic Fermentation▿ †  
Applied and Environmental Microbiology  2007;73(16):5363-5369.
Genome-wide analysis of the wine yeast strain Saccharomyces cerevisiae PYCC4072 identified 36 genes highly expressed under conditions of low or absent nitrogen in comparison with a nitrogen-replete condition. Reverse transcription-PCR analysis for four of these transcripts with this strain and its validation with another wine yeast strain underlines the usefulness of these signature genes for predicting nitrogen deficiency and therefore the diagnosis of wine stuck/sluggish fermentations.
PMCID: PMC1950961  PMID: 17601813
3.  Transcriptional Response of Saccharomyces cerevisiae to Different Nitrogen Concentrations during Alcoholic Fermentation▿ †  
Gene expression profiles of a wine strain of Saccharomyces cerevisiae PYCC4072 were monitored during alcoholic fermentations with three different nitrogen supplies: (i) control fermentation (with enough nitrogen to complete sugar fermentation), (ii) nitrogen-limiting fermentation, and (iii) the addition of nitrogen to the nitrogen-limiting fermentation (refed fermentation). Approximately 70% of the yeast transcriptome was altered in at least one of the fermentation stages studied, revealing the continuous adjustment of yeast cells to stressful conditions. Nitrogen concentration had a decisive effect on gene expression during fermentation. The largest changes in transcription profiles were observed when the early time points of the N-limiting and control fermentations were compared. Despite the high levels of glucose present in the media, the early responses of yeast cells to low nitrogen were characterized by the induction of genes involved in oxidative glucose metabolism, including a significant number of mitochondrial associated genes resembling the yeast cell response to glucose starvation. As the N-limiting fermentation progressed, a general downregulation of genes associated with catabolism was observed. Surprisingly, genes encoding ribosomal proteins and involved in ribosome biogenesis showed a slight increase during N starvation; besides, genes that comprise the RiBi regulon behaved distinctively under the different experimental conditions. Here, for the first time, the global response of nitrogen-depleted cells to nitrogen addition under enological conditions is described. An important gene expression reprogramming occurred after nitrogen addition; this reprogramming affected genes involved in glycolysis, thiamine metabolism, and energy pathways, which enabled the yeast strain to overcome the previous nitrogen starvation stress and restart alcoholic fermentation.
PMCID: PMC1892875  PMID: 17337556
4.  The yeast FBP1 poly(A) signal functions in both orientations and overlaps with a gene promoter. 
Nucleic Acids Research  1998;26(20):4588-4596.
This report provides an analysis of a region of chromosome XII in which the FBP1 and YLR376c genes transcribe in the same direction. Our investigation indicates that the Saccharomyces cerevisiae FBP1 gene contains strong signals for polyadenylation and transcription termination in both orientations in vivo . A (TA)14 element plays a major role in directing polyadenylation in both orientations. While this region has four nonoverlapping copies of a TATATA hexanucleotide, which is a very potent polyadenylation efficiency element in yeast, it alone is not sufficient for full activation in the reverse orientation of a cluster of downstream poly(A) sites, and an additional upstream sequence is required. The putative RNA hairpin formed from the (TA)14 element is not involved in 3'-end formation. Surprisingly, deletion of the entire (TA)14 stretch affects transcription termination in the reverse orientation, in contrast to our previous results with the forward orientation, indicating that the transcription termination element operating in the reverse orientation has very different sequence requirements. Promoter elements for the YLR376c gene overlap with the signal for FBP1 3'-end formation. To our knowledge, this is the first time that overlapping of both types of regulatory signals has been found in two adjacent yeast genes.
PMCID: PMC147910  PMID: 9753725
5.  AZT treatment induces molecular and ultrastructural oxidative damage to muscle mitochondria. Prevention by antioxidant vitamins. 
AIDS patients who receive zidovudine (AZT) frequently suffer from myopathy. This has been attributed to mitochondrial (mt) damage, and specifically to the loss of mtDNA. This study examines whether AZT causes oxidative damage to DNA in patients and to skeletal muscle mitochondria in mice, and whether this damage may be prevented by supranutritional doses of antioxidant vitamins. Asymptomatic HIV-infected patients treated with AZT have a higher urinary excretion (355+/-100 pmol/kg/d) of 8-oxo-7, 8-dihydro-2'-deoxyguanosine (8-oxo-dG) (a marker of oxidative damage to DNA) than untreated controls (asymptomatic HIV-infected patients) (182+/-29 pmol/kg/d). This was prevented (110+/-79 pmol/kg/d) by simultaneous oral treatment with AZT plus antioxidant vitamins (C and E). Mice treated with AZT also had a significantly higher urinary excretion of 8-oxo-dG than controls. Skeletal muscle mtDNA of mice treated with AZT had more 8-oxo-dG than controls. mt lipoperoxidation was also increased and skeletal muscle glutathione was oxidized. These effects may be due to an increased peroxide production by muscle mitochondria of AZT-treated animals. Dietary supplements with vitamins C and E at supranutritional doses protect against oxidative damage to skeletal muscle mitochondria caused by AZT.
PMCID: PMC509058  PMID: 9649550
6.  Utilization of hemin and hemoglobin by Vibrio vulnificus biotype 2. 
The eel pathogen Vibrio vulnificus biotype 2 is able to use hemoglobin (Hb) and hemin (Hm) to reverse iron limitation. In this stud, the adjuvant effect of both compounds on eel pathogenicity has been evaluated and confirmed. Further, we have studied the heme-iron acquisition mechanism displayed by this bacterium. Whole cells were capable of binding Hb and Hm, independently of (i) iron levels in growth medium and (ii) the presence of polysaccharide capsules on bacterial surface. The Hb- and Hm-binding capacity was retained by the outer membrane protein (OMP) fraction and was abolished after proteolytic digestion of OMP samples. Western blotting (immunoblotting) of denatured OMPs revealed that two major protein bands of 36 and 32 kDa were involved in both Hm and Hb binding. The expression of these proteins was not affected by iron levels. In addition, V. vulnificus biotype 2 produced extracellular proteases, not regulated by iron, that were active against native Hb. In conclusion, the overall data suggest that the eel pathogen V. vulnificus biotype 2 can obtain iron by means of a mechanism which involves a direct interaction between the heme moiety and constitutive OMPs.
PMCID: PMC168066  PMID: 8702273

Results 1-6 (6)