ABSTRACT
BACKGROUND INFORMATION: The CBK1 gene of Saccharomyces cerevisiae encodes a protein kinase that is a member of the NDR (nuclear Dbf2-related) family of protein kinases, which are involved in morphogenesis and cell proliferation. Previous studies have shown that deletion of CBK1 leads to a loss of polarity and the formation of large aggregates of cells. This aggregation phenotype is due to the loss of the daughter cell-specific accumulation of the transcription factor Ace2p, which is responsible for the transcription of genes whose products are necessary for the final separation of the mother and the daughter at the end of cell division. RESULTS: We show that the daughter cell-specific localization of Ace2p does not occur via a specific localization of the ACE2 mRNA and that, in vivo, the transcription of CTS1, one of the principal targets of Ace2p, is daughter cell-specific. We have shown that extragenic suppressors of the Deltacbk1 aggregation phenotype are located in the nuclear exportin CRM1 and ACE2. These mutations disrupt the interaction of Ace2p and Crm1p, thus impairing Ace2p export and resulting in the accumulation of the protein in both mother and daughter cell nuclei. CONCLUSIONS: We propose that in the daughter cell nucleus Cbk1p phosphorylates the Ace2p nuclear export signal, and that this phosphorylation blocks the export of Ace2p via Crm1p, thus promoting the daughter cell-specific nuclear accumulation of Ace2p.
Subject(s)
Cell Nucleus/metabolism , DNA-Binding Proteins/metabolism , Karyopherins/genetics , Mutation , Receptors, Cytoplasmic and Nuclear/genetics , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae/metabolism , Transcription Factors/metabolism , Active Transport, Cell Nucleus , Cell Division , Cell Nucleus/genetics , Chitinases/genetics , DNA-Binding Proteins/analysis , DNA-Binding Proteins/genetics , Fungal Proteins/analysis , Fungal Proteins/genetics , Fungal Proteins/metabolism , Intracellular Signaling Peptides and Proteins , Karyopherins/metabolism , Protein Serine-Threonine Kinases , Protein Transport , Receptors, Cytoplasmic and Nuclear/metabolism , Repressor Proteins/analysis , Repressor Proteins/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae Proteins/analysis , Saccharomyces cerevisiae Proteins/genetics , Species Specificity , Transcription Factors/analysis , Transcription Factors/genetics , Exportin 1 ProteinABSTRACT
In Saccharomyces cerevisiae the nicotinic acid moiety of NAD+ can be synthesized from tryptophan using the kynurenine pathway or incorporated directly using nicotinate phosphoribosyl transferase (NPT1). We have identified the genes that encode the enzymes of the kynurenine pathway and for BNA5 (YLR231c) and BNA6 (YFR047c) confirmed that they encode kynureninase and quinolinate phosphoribosyl transferase respectively. We show that deletion of genes encoding kynurenine pathway enzymes are co-lethal with the Deltanpt1, demonstrating that no other pathway for the synthesis of nicotinic acid exists in S. cerevisiae. Also, we show that under anaerobic conditions S. cerevisiae is a nicotinic acid auxotroph.