Azf1p is a nuclear-localized zinc-finger protein that is preferentially expressed under non-fermentative growth conditions in Saccharomyces cerevisiae.
In previous studies the AZF1 gene has been identified as a second high-copy number suppressor for a special mutant of the gene for the mitochondrial core enzyme of RNA polymerase. The first high-copy number suppressor of this mutant turned out to be the specificity factor MTF1 for mitochondrial transcription. Up to now, the influence of AZF1 on mitochondrial transcription, its precise localization in the cell and the regulation of its expression has not been determined. The putative protein contains a long stretch of poly-asparagine amino acids and a typical zinc-finger domain for DNA binding. These characteristic structural features were used to create the abbreviation AZF1 (Asparagine-rich Zinc Finger protein). An initial computer analysis of the sequence gave no conclusive results for the presence of a mitochondrial import sequence or a typical nuclear-targeting sequence. A recent more-detailed analysis identified a possible nuclear localization signal in the middle of the protein. Disruption of the gene shows no effect on plates with glucose-rich medium or glycerol. In this report a specific polyclonal antibody against Azf1p was prepared and used in cell-fractionation experiments and in electron-microscopic studies. Both of these clearly demonstrate that the AZF1 protein is localized exclusively in the nucleus of the yeast cell. Northern analysis for the expression of the AZF1 messenger RNA under different growth conditions was therefore performed to obtain new insights into the regulation of this gene. Together with the respective protein-expression analysis these data demonstrate that Azf1p is preferentially synthezised in higher amounts under non-fermentable growth conditions. Over-expression of Azf1p in the yeast cell does not influence the expression level of the mitochondrial transcription factor Mtf1p, indicating that the influence of Azf1p on the suppression of the special mitochondrial RNA polymerase mutant is an indirect one. Subcellular investigation of the deletion mutant by electron microscopy identifies specific ultrastructural cell-division defects in comparison to the wild-type.
Stein
T
T
Botanisches Institut I, Heinrich-Heine-Universität Düsseldorf, Universitätsstrasse 1, D-40225 Düsseldorf, Germany.
Kricke
J
J
Becher
D
D
Lisowsky
T
T
D002352:Carrier Proteins / Q000235:genetics*; D002455:Cell Division / Q000235:genetics*; D005285:Fermentation / Q000502:physiology; D005656:Fungal Proteins / Q000235:genetics; D018628:Gene Dosage; D015966:Gene Expression Regulation, Fungal / Q000235:genetics*; D049452:Green Fluorescent Proteins; D007150:Immunohistochemistry; D008164:Luminescent Proteins / Q000235:genetics; D016253:Microscopy, Immunoelectron; D008928:Mitochondria / Q000201:enzymology / Q000235:genetics; D024101:Mitochondrial Proteins; D019913:Nuclear Localization Signals / Q000235:genetics; D009687:Nuclear Proteins / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion / Q000235:genetics; D013489:Suppression, Genetic / Q000235:genetics; D014157:Transcription Factors; D014158:Transcription, Genetic / Q000235:genetics; D016335:Zinc Fingers / Q000235:genetics*
C116478:AZF1 protein, S cerevisiae; D002352:Carrier Proteins; D005656:Fungal Proteins; D008164:Luminescent Proteins; C071720:MTF1 protein, S cerevisiae; D024101:Mitochondrial Proteins; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins