Protein phosphorylation is now recognized to be one of the most important means of regulating protein activity. An approach was taken that was aimed at identifying new protein Serine/Threonine kinase genes in the fruit fly Drosophila melanogaster. Three of the kinases identified were chosen for molecular characterization: a Map kinase-activated protein kinase-2 homolog (DmMAPKAPK-2); a novel female germline specific kinase ( loki); and the homolog of the vertebrate cdk7 genes (Dmcdk7). Among those, Dmcdk7 was chosen for in depth molecular and genetic characterization. Cdk7 has previously been shown in vertebrate systems to phosphorylate and activate many different Cyclin-dependent kinases (Cdks) in vitro. However, conclusive evidence that Cdk7 could act as a Cdk-activating kinase (CAK) in vivo had remained elusive, and became even controversial. Adding to the controversy was the fact that in the budding yeast S. cerevisiae, CAK activity is provided by the CAK1/Civ1 protein which is unrelated to Cdk7. It was therefore proposed that the CAK activity of Cdk7 may be an in vitro artefact. In an attempt to resolve this issue null and temperature sensitive mutations of the Dmcdk7 gene have been created. The results obtained using these mutant alleles of Dmcdk7 demonstrate that cdk7 is necessary for CAK activity in vivo in a multicellular organism. It is shown that cdk7 activity is required for the activation of both Cdc2/Cyclin A and Cdc2/Cyclin B complexes, and for cell division. In addition to validate the function of Cdk7 as a bona fide regulator of the cell cycle, these results suggest that there may be a fundamental difference in the way metazoans and budding yeast effect a key modification of Cdks. Phosphorylation events at different sites (including the T-loop) are also known to be involved in stabilizing the Cdk7/Cyclin H dimer in vitro, and have been shown to occur in vivo. Surprisingly, the in vivo analysis of different phosphorylation mutant forms of DmCdk7 f
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.37548 |
| Date | January 1998 |
| Creators | Larochelle, Stéphane. |
| Contributors | Suter, Beat (advisor) |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Biology.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001610659, proquestno: NQ44487, Theses scanned by UMI/ProQuest. |
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