Control of gag-pol gene expression in the Candida albicans retrotransposon Tca2

Persistent Link:
http://hdl.handle.net/10150/610060
Title:
Control of gag-pol gene expression in the Candida albicans retrotransposon Tca2
Author:
Forbes, Elaine; Nieduszynska, Sian; Brunton, Fiona; Gibson, Joanne; Glover, L. A.; Stansfield, Ian
Affiliation:
University of Aberdeen, School of Medical Sciences, Institute of Medical Sciences, Foresterhill, Aberdeen AB25 2ZD, UK; Current address : Dept. of Pediatrics, University of Arizona, College of Medicine, Tucson, AZ 85724, USA; Current address : School of Medicine, King's College London, Strand, London, WC2R 2LS, UK
Issue Date:
2007
Publisher:
BioMed Central
Citation:
BMC Molecular Biology 2007, 8:94 doi:10.1186/1471-2199-8-94
Journal:
BMC Molecular Biology
Rights:
© 2007 Forbes et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0)
Collection Information:
This item is part of the UA Faculty Publications collection. For more information this item or other items in the UA Campus Repository, contact the University of Arizona Libraries at repository@u.library.arizona.edu.
Abstract:
BACKGROUND:In the C. albicans retrotransposon Tca2, the gag and pol ORFs are separated by a UGA stop codon, 3' of which is a potential RNA pseudoknot. It is unclear how the Tca2 gag UGA codon is bypassed to allow pol expression. However, in other retroelements, translational readthrough of the gag stop codon can be directed by its flanking sequence, including a 3' pseudoknot.RESULTS:The hypothesis was tested that in Tca2, gag stop codon flanking sequences direct translational readthrough and synthesis of a gag-pol fusion protein. Sequence from the Tca2 gag-UGA-pol junction (300 nt) was inserted between fused lacZ and luciferase (luc) genes in a Saccharomyces cerevisiae dual reporter construct. Although downstream of UGA, luc was expressed, but its expression was unaffected by inserting additional stop codons at the 3' end of lacZ. Luc expression was instead being driven by a previously unknown minor promoter activity within the gag-pol junction region. Evidence together indicated that junction sequence alone cannot direct UGA readthrough. Using reporter genes in C. albicans, the activities of this gag-pol junction promoter and the Tca2 long terminal repeat (LTR) promoter were compared. Of the two promoters, only the LTR promoter was induced by heat-shock, which also triggers retrotransposition. Tca2 pol protein, epitope-tagged in C. albicans to allow detection, was also heat-shock induced, indicating that pol proteins were expressed from a gag-UGA-pol RNA.CONCLUSION:This is the first demonstration that the LTR promoter directs Tca2 pol protein expression, and that pol proteins are translated from a gag-pol RNA, which thus requires a mechanism for stop codon bypass. However, in contrast to most other retroelement and viral readthrough signals, immediate gag UGA-flanking sequences were insufficient to direct stop readthrough in S. cerevisiae, indicating non-canonical mechanisms direct gag UGA bypass in Tca2.
EISSN:
1471-2199
DOI:
10.1186/1471-2199-8-94
Version:
Final published version
Additional Links:
http://www.biomedcentral.com/1471-2199/8/94

Full metadata record

DC FieldValue Language
dc.contributor.authorForbes, Elaineen
dc.contributor.authorNieduszynska, Sianen
dc.contributor.authorBrunton, Fionaen
dc.contributor.authorGibson, Joanneen
dc.contributor.authorGlover, L. A.en
dc.contributor.authorStansfield, Ianen
dc.date.accessioned2016-05-20T08:57:40Z-
dc.date.available2016-05-20T08:57:40Z-
dc.date.issued2007en
dc.identifier.citationBMC Molecular Biology 2007, 8:94 doi:10.1186/1471-2199-8-94en
dc.identifier.doi10.1186/1471-2199-8-94en
dc.identifier.urihttp://hdl.handle.net/10150/610060-
dc.description.abstractBACKGROUND:In the C. albicans retrotransposon Tca2, the gag and pol ORFs are separated by a UGA stop codon, 3' of which is a potential RNA pseudoknot. It is unclear how the Tca2 gag UGA codon is bypassed to allow pol expression. However, in other retroelements, translational readthrough of the gag stop codon can be directed by its flanking sequence, including a 3' pseudoknot.RESULTS:The hypothesis was tested that in Tca2, gag stop codon flanking sequences direct translational readthrough and synthesis of a gag-pol fusion protein. Sequence from the Tca2 gag-UGA-pol junction (300 nt) was inserted between fused lacZ and luciferase (luc) genes in a Saccharomyces cerevisiae dual reporter construct. Although downstream of UGA, luc was expressed, but its expression was unaffected by inserting additional stop codons at the 3' end of lacZ. Luc expression was instead being driven by a previously unknown minor promoter activity within the gag-pol junction region. Evidence together indicated that junction sequence alone cannot direct UGA readthrough. Using reporter genes in C. albicans, the activities of this gag-pol junction promoter and the Tca2 long terminal repeat (LTR) promoter were compared. Of the two promoters, only the LTR promoter was induced by heat-shock, which also triggers retrotransposition. Tca2 pol protein, epitope-tagged in C. albicans to allow detection, was also heat-shock induced, indicating that pol proteins were expressed from a gag-UGA-pol RNA.CONCLUSION:This is the first demonstration that the LTR promoter directs Tca2 pol protein expression, and that pol proteins are translated from a gag-pol RNA, which thus requires a mechanism for stop codon bypass. However, in contrast to most other retroelement and viral readthrough signals, immediate gag UGA-flanking sequences were insufficient to direct stop readthrough in S. cerevisiae, indicating non-canonical mechanisms direct gag UGA bypass in Tca2.en
dc.language.isoenen
dc.publisherBioMed Centralen
dc.relation.urlhttp://www.biomedcentral.com/1471-2199/8/94en
dc.rights© 2007 Forbes et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0)en
dc.titleControl of gag-pol gene expression in the Candida albicans retrotransposon Tca2en
dc.typeArticleen
dc.identifier.eissn1471-2199en
dc.contributor.departmentUniversity of Aberdeen, School of Medical Sciences, Institute of Medical Sciences, Foresterhill, Aberdeen AB25 2ZD, UKen
dc.contributor.departmentCurrent address : Dept. of Pediatrics, University of Arizona, College of Medicine, Tucson, AZ 85724, USAen
dc.contributor.departmentCurrent address : School of Medicine, King's College London, Strand, London, WC2R 2LS, UKen
dc.identifier.journalBMC Molecular Biologyen
dc.description.collectioninformationThis item is part of the UA Faculty Publications collection. For more information this item or other items in the UA Campus Repository, contact the University of Arizona Libraries at repository@u.library.arizona.edu.en
dc.eprint.versionFinal published versionen
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