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K F Cooper

Publications and source records attributed to K F Cooper.

8 recordsLinked to original sources
PubMed2000

Ama1p is a meiosis-specific regulator of the anaphase promoting complex/cyclosome in yeast.

Meiosis is the developmental program by which diploid organisms produce haploid gametes capable of sexual reproduction. Here we describe the yeast gene AMA1, a new member of the Cdc20 protein family that regulates the multisubunit ubiquitin ligase termed the anaphase promoting complex/cyclosome (APC/C). AMA1 is developmentally regulated in that its transcription and splicing occur only in meiotic cells. The meiosis-specific processing of AMA1 mRNA depends on the previously described MER1 splicing factor. Several results indicate that Ama1p is required for APC/C function during meiosis. First, coimmunoprecipitation assays indicate that Ama1p associates with the APC/C in vivo. Second, Ama1p is required for the degradation of the B-type cyclin Clb1p, an APC/C substrate in both meiotic and mitotic cells. Third, ectopic overexpression of AMA1 is able to stimulate ubiquitination of Clb1p in vitro and degradation of Clb1p in vivo. Mutants lacking AMA1 revealed that it is required for the first meiotic division but not the mitotic-like meiosis II. In addition, ama1 mutants are defective for both spore wall assembly and the expression of late meiotic genes. In conclusion, this study indicates that Ama1p directs a meiotic APC/C that functions solely outside mitotic cell division. The requirement of Ama1p only for meiosis I and spore morphogenesis suggests a function for APC/C(Ama1) specifically adapted to germ cell development.

Amino Acid Sequence↗
PubMed1999

Oxidative stress-induced destruction of the yeast C-type cyclin Ume3p requires phosphatidylinositol-specific phospholipase C and the 26S proteasome.

The yeast UME3 (SRB11/SSN3) gene encodes a C-type cyclin that represses the transcription of the HSP70 family member SSA1. To relieve this repression, Ume3p is rapidly destroyed in cells exposed to elevated temperatures. This report demonstrates that Ume3p levels are also reduced in cultures subjected to ethanol shock, oxidative stress, or carbon starvation or during growth on nonfermentable carbons. Of the three elements (RXXL, PEST, and cyclin box) previously shown to be required for heat-induced Ume3p destruction, only the cyclin box regulates Ume3p degradation in response to these stressors. The one exception observed was growth on nonfermentable carbons, which requires the PEST region. These findings indicate that yeast cells contain multiple, independent pathways that mediate stress-induced Ume3p degradation. Ume3p destruction in response to oxidative stress, but not to ethanol treatment, requires DOA4 and UMP1, two factors required for 26S proteasome activity. This result for the first time implicates ubiquitin-mediated proteolysis in C-type cyclin regulation. Similarly, the presence of a membrane stabilizer (sorbitol) or the loss of phosphatidylinositol-specific phospholipase C (PLC1) protects Ume3p from oxidative-stress-induced degradation. Finally, a ume3 null allele suppresses the growth defect of plc1 mutants in response to either elevated temperature or the presence of hydrogen peroxide. These results indicate that the growth defects observed in plc1 mutants are due to the failure to downregulate Ume3p. Taken together, these findings support a model in which Plc1p mediates an oxidative-stress signal from the plasma membrane that triggers Ume3p destruction through a Doa4p-dependent mechanism.

Cyclin-Dependent Kinases↗
PubMed1999

Functional analysis of the Ume3p/ Srb11p-RNA polymerase II holoenzyme interaction.

The yeast C-type cyclin Ume3p/Srb11p and its cyclin-dependent kinase (Cdk) Ume5p are required for the full repression of genes involved in the stress response or meiosis. This cyclin-Cdk kinase copurifies with the RNA polymerase II holoenzyme complex, suggesting it functions through modification of the transcriptional machinery. This report describes two domains required for Ume3p-RNA Pol II holoenzyme association. One domain contains the highly conserved cyclin box that directs cyclin-Cdk interaction and requires Ume5p for holoenzyme binding. The second domain, termed HAD for holoenzyme associating domain, is located within the amino-terminal region of the cyclin and is sufficient for holoenzyme binding independent of Ume5p or the cyclin box. In addition to its role in RNA Pol II holoenzyme association, the HAD is also required for Ume3p-dependent repression in vivo. Finally, HAD mutations do not affect the ability of the Ume3p-Ume5p kinase to phosphorylate in vitro the carboxy-terminal domain (CTD) of RNA polymerase II, a reported target of cyclin C-Cdk activity. In conclusion, this study demonstrates that the association of the Ume3p to the holoenzyme is complex, involving two independent domains, both of which are required for full Ume3p-dependent repression in vivo. Furthermore, HAD-dependent repression does not appear to involve CTD phosphorylation, suggesting a different role for this domain in directing Ume3p-Ume5p activity.

Adenosine Triphosphatases↗
PubMed1997

Stress and developmental regulation of the yeast C-type cyclin Ume3p (Srb11p/Ssn8p).

The ume3-1 allele was identified as a mutation that allowed the aberrant expression of several meiotic genes (e.g. SPO11, SPO13) during mitotic cell division in Saccharomyces cerevisiae. Here we report that UME3 is also required for the full repression of the HSP70 family member SSA1. UME3 encodes a non-essential C-type cyclin (Ume3p) whose levels do not vary through the mitotic cell cycle. However, Ume3p is destroyed during meiosis or when cultures are subjected to heat shock. Ume3p mutants resistant to degradation resulted in a 2-fold reduction in SPO13 mRNA levels during meiosis, indicating that the down-regulation of this cyclin is important for normal meiotic gene expression. Mutational analysis identified two regions (PEST-rich and RXXL) that mediate Ume3p degradation. A third destruction signal lies within the highly conserved cyclin box, a region that mediates cyclin-cyclin-dependent kinase (Cdk) interactions. However, the Cdk activated by Ume3p (Ume5p) is not required for the rapid destruction of this cyclin. Finally, Ume3p destruction was not affected in mutants defective for ubiquitin-dependent proteolysis. These results support a model in which Ume3p, when exposed to heat shock or sporulation conditions, is targeted for destruction to allow the expression of genes necessary for the cell to respond correctly to these environmental cues.

Adenosine Triphosphatases↗
PubMed1993

Structure of the sequences adjacent to the centromeric alphoid satellite DNA array on the human Y chromosome.

Eighteen yeast artificial chromosome (YAC) clones containing alphoid satellite DNA and adjacent sequences from the human Y chromosome have been identified from three different YAC libraries. Restriction site mapping of the genomic alphoid arrays and the YACs has allowed seven of the alphoid clones to be positioned on the arrays. Three clones extend into flanking sequences. At one edge the alphoid DNA is highly diverged and is flanked by a small block of the 48 base-pair satellite, dispersed moderately repeated sequences and a separate short alphoid array. More distal sequences are Y-specific. At the other edge there is much less divergence and the alphoid DNA is flanked by an Alu sequence and the five base-pair satellite.

Centromere↗
PubMed1993

The major centromeric array of alphoid satellite DNA on the human Y chromosome is non-palindromic.

We have determined the orientation of the major centromeric alphoid array on the human Y chromosome. A PCR assay was used to analyse the vector-insert junctions of seven YAC clones previously positioned on two independent Y chromosomes. The orientation is the same at all 10 positions measured. This suggests that the alphoid array is a simple unidirectional repeat throughout and that human centromere structure thus differs from the palindromic organisation found in Schizosaccharomyces pombe.

Base Sequence↗
PubMed1992

Structure of the pericentric long arm region of the human Y chromosome.

We have analysed the sequence organization of the DNA in the pericentric region of the long arm of the human Y chromosome. The structures of one cosmid and three yeast artificial chromosome clones were determined. The region consists of a mosaic of the known 5, 48 and 68 base-pair tandemly repeated sequences and at least five novel repeated sequence families. A long range-map of approximately 3.5 x 10(6) base-pairs of genomic DNA was constructed that placed the clones between about 500 x 10(3) and 850 x 10(3) base-pairs from the long arm edge of the centromeric alphoid DNA array.

Animals↗