Screening for Theileria parva secretory gene products by functional analysis in Saccharomyces cerevisiae

SR 31747 is a novel immunosuppressant agent that arrests cell proliferation in the yeast Saccharomyces cerevisiae, SR 31747-treated cells accumulate the same aberrant sterols as those found in a mutant impaired in delta 8- delta 7-sterol isomerase. Sterol isomerase activity is also inhibited by SR 31747 in in vitro assays. Overexpression of the sterol isomerase-encoding gene, ERG2, confers enhanced SR resistance. Cells growing anaerobically on ergosterol-containing medium are not sensitive to SR. Disruption of the sterol isomerase-encoding gene is lethal in cells growing in the absence of exogenous ergosterol, except in SR-resistant mutants lacking either the SUR4 or the FEN1 gene product. The results suggest that sterol isomerase is the target of SR 31747 and that both the SUR4 and FEN1 gene products are required to mediate the proliferation arrest induced by ergosterol depletion.

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Two systems of glucose repression of the GAL1 promoter in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.10.9.4757-4769.1990 ◽

1990 ◽

Vol 10 (9) ◽

pp. 4757-4769

Author(s):

J S Flick ◽

M Johnston

Keyword(s):

Saccharomyces Cerevisiae ◽

Base Pair ◽

Glucose Repression ◽

Gene Products ◽

Gal1 Promoter ◽

Base Pair Fragment ◽

Common Signal ◽

Multiple Sequences ◽

Pair Fragment ◽

Gal1 Gene

Expression of the GAL1 gene in Saccharomyces cerevisiae is strongly repressed by growth on glucose. We show that two sites within the GAL1 promoter mediate glucose repression. First, glucose inhibits transcription activation by GAL4 protein through UASG. Second, a promoter element, termed URSG, confers glucose repression independently of GAL4. We have localized the URSG sequences responsible for glucose repression to an 87-base-pair fragment located between UASG and the TATA box. Promoters deleted for small (20-base-pair) segments that span this sequence are still subject to glucose repression, suggesting that there are multiple sequences within this region that confer repression. Extended deletions across this region confirm that it contains at least two and possibly three URSG elements. To identify the gene products that confer repression upon UASG and URSG, we have analyzed glucose repression mutants and found that the GAL83, REG1, GRR1, and SSN6 genes are required for repression mediated by both UASG and URSG. In contrast, GAL82 and HXK2 are required only for UASG repression. A mutation designated urr1-1 (URSG repression resistant) was identified that specifically relieves URSG repression without affecting UASG repression. In addition, we observed that the SNF1-encoded protein kinase is essential for derepression of both UASG and URSG. We propose that repression of UASG and URSG is mediated by two independent pathways that respond to a common signal generated by growth on glucose.

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The HML mating-type cassette of Saccharomyces cerevisiae is regulated by two separate but functionally equivalent silencers

Molecular and Cellular Biology ◽

10.1128/mcb.9.11.4621-4630.1989 ◽

1989 ◽

Vol 9 (11) ◽

pp. 4621-4630

Author(s):

D J Mahoney ◽

J R Broach

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Binding Sites ◽

The Other ◽

Functional Domains ◽

Gene Products ◽

Cis Acting ◽

Full Expression ◽

Mating Type Genes ◽

Chromosome Iii

Mating-type genes resident in the silent cassette HML at the left arm of chromosome III are repressed by the action of four SIR gene products, most likely mediated through two cis-acting sites located on opposite sides of the locus. We showed that deletion of either of these two cis-acting sites from the chromosome did not yield any detectable derepression of HML, while deletion of both sites yielded full expression of the locus. In addition, each of these sites was capable of exerting repression of heterologous genes inserted in their vicinity. Thus, HML expression is regulated by two independent silencers, each fully competent for maintaining repression. This situation was distinct from the organization of the other silent locus, HMR, at which a single silencer served as the predominant repressor of expression. Examination of identifiable domains and binding sites within the HML silencers suggested that silencing activity can be achieved by a variety of combinations of various functional domains.

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The RAD7 and RAD16 genes, which are essential for pyrimidine dimer removal from the silent mating type loci, are also required for repair of the nontranscribed strand of an active gene in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.14.9.6135-6142.1994 ◽

1994 ◽

Vol 14 (9) ◽

pp. 6135-6142

Author(s):

R Verhage ◽

A M Zeeman ◽

N de Groot ◽

F Gleig ◽

D D Bang ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Mating Type ◽

Excision Repair ◽

Complementation Group ◽

Pyrimidine Dimer ◽

Gene Products ◽

Pyrimidine Dimers ◽

Nucleotide Excision ◽

Active Genes ◽

Silent Mating Type Loci

The rad16 mutant of Saccharomyces cerevisiae was previously shown to be impaired in removal of UV-induced pyrimidine dimers from the silent mating-type loci (D. D. Bang, R. A. Verhage, N. Goosen, J. Brouwer, and P. van de Putte, Nucleic Acids Res. 20:3925-3931, 1992). Here we show that rad7 as well as rad7 rad16 double mutants have the same repair phenotype, indicating that the RAD7 and RAD16 gene products might operate in the same nucleotide excision repair subpathway. Dimer removal from the genome overall is essentially incomplete in these mutants, leaving about 20 to 30% of the DNA unrepaired. Repair analysis of the transcribed RPB2 gene shows that the nontranscribed strand is not repaired at all in rad7 and rad16 mutants, whereas the transcribed strand is repaired in these mutants at a fast rate similar to that in RAD+ cells. When the results obtained with the RPB2 gene can be generalized, the RAD7 and RAD16 proteins not only are essential for repair of silenced regions but also function in repair of nontranscribed strands of active genes in S. cerevisiae. The phenotype of rad7 and rad16 mutants closely resembles that of human xeroderma pigmentosum complementation group C (XP-C) cells, suggesting that RAD7 and RAD16 in S. cerevisiae function in the same pathway as the XPC gene in human cells. RAD4, which on the basis of sequence homology has been proposed to be the yeast XPC counterpart, seems to be involved in repair of both inactive and active yeast DNA, challenging the hypothesis that RAD4 and XPC are functional homologs.

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Functional analysis of Rpn4-like proteins from Komagataella (Pichia) pastoris and Yarrowia lipolytica on a genetic background of Saccharomyces cerevisiae

Applied Biochemistry and Microbiology ◽

10.1134/s0003683815070029 ◽

2015 ◽

Vol 51 (7) ◽

pp. 757-765

Author(s):

E. N. Grineva ◽

A. T. Leinsoo ◽

D. S. Spasskaya ◽

D. S. Karpov ◽

V. L. Karpov

Keyword(s):

Saccharomyces Cerevisiae ◽

Functional Analysis ◽

Pichia Pastoris ◽

Yarrowia Lipolytica ◽

Genetic Background

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TWO RECESSIVE SUPPRESSORS OF SACCHAROMYCES CEREVISIAE CHO1 THAT ARE UNLINKED BUT FALL IN THE SAME COMPLEMENTATION GROUP

Genetics ◽

10.1093/genetics/111.1.1 ◽

1985 ◽

Vol 111 (1) ◽

pp. 1-6

Author(s):

Katharine D Atkinson

Keyword(s):

Saccharomyces Cerevisiae ◽

Meiotic Recombination ◽

Genetic Locus ◽

Complementation Group ◽

Complementation Analysis ◽

Gene Products ◽

Chromosome X ◽

Recessive Mutations ◽

Recessive Mutant ◽

Phenotypic Reversion

ABSTRACT Phenotypic reversion of ethanolamine-requiring Saccharomyces cerevisiae cho 1 mutants is predominantly due to recessive mutations at genes unlinked to the chromosome V cho 1 locus. The recessive suppressors do not correct the primary cho 1 defect in phosphatidylserine synthesis but circumvent it with a novel endogenous supply of ethanolamine. One suppressor (eam1) was previously mapped to chromosome X, and 135 suppressor isolates were identified as eam1 alleles by complementation analysis. Additional meiotic recombination studies have identified a second genetic locus, eam2, that falls in the eam1 complementation group but maps close to the centromere of chromosome IV. Although the normal EAM1 and EAM2 alleles are fully dominant over recessive mutant alleles, their dominance fails in diploids heterozygous for defects in both genes simultaneously. The unusual complementation pattern could be explained by interaction of the gene products in formation of the same enzyme.

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Lysis Genes of the Bacillus subtilisDefective Prophage PBSX

Journal of Bacteriology ◽

10.1128/jb.180.8.2110-2117.1998 ◽

1998 ◽

Vol 180 (8) ◽

pp. 2110-2117 ◽

Cited By ~ 37

Author(s):

Susanne Krogh ◽

Steen T. Jørgensen ◽

Kevin M. Devine

Keyword(s):

Functional Analysis ◽

Host Cell ◽

Expression System ◽

Cell Lysis ◽

Lethal Gene ◽

Gram Negative Bacteria ◽

Gene Products ◽

Gram Negative ◽

Gram Positive Bacteria ◽

Exported Protein

ABSTRACT Four genes identified within the late operon of PBSX show characteristics expected of a host cell lysis system; they arexepA, encoding an exported protein; xhlA, encoding a putative membrane-associated protein; xhlB, encoding a putative holin; and xlyA, encoding a putative endolysin. In this work, we have assessed the contribution of each gene to host cell lysis by expressing the four genes in different combinations under the control of their natural promoter located on the chromosome of Bacillus subtilis 168. The results show thatxepA is unlikely to be involved in host cell lysis. Expression of both xhlA and xhlB is necessary to effect host cell lysis of B. subtilis. Expression ofxhlB (encoding the putative holin) together withxlyA (encoding the endolysin) cannot effect cell lysis, indicating that the PBSX lysis system differs from those identified in the phages of gram-negative bacteria. Since host cell lysis can be achieved when xlyA is inactivated, it is probable that PBSX encodes a second endolysin activity which also uses XhlA and XhlB for export from the cell. The chromosome-based expression system developed in this study to investigate the functions of the PBSX lysis genes should be a valuable tool for the analysis of other host cell lysis systems and for expression and functional analysis of other lethal gene products in gram-positive bacteria.

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