The CDP/Cux transcription factor contains four DNA binding domains: three Cut repeats (CR1, CR2 and CR3) and the Cut homeodomain (HD). The CCAAT-displacement protein (CDP) was first identified as it represses transcription of certain promoters by competing for the occupancy of the CCAAT sequence. CDP was then found to be the ortholog of the Drosophila Cut protein (Cux: Cu t homeobox&barbelow;). The goal of my project was to define the modes of interactions of CDP/Cux with DNA. I demonstrated that high affinity DNA binding requires the cooperation between at least two of the CDP/Cux DNA binding domains. Among all combinations of domains, Cut repeats 1 and 2 (CR1CR2) and Cut repeat 3 and the Cut homeodomain (CR3HD) exhibited the highest DNA binding affinities, but with different kinetics and specificities. Whereas CR1CR2 bound with fast kinetics to dimers of CART or CGAT, CR3HD bound to ATCGAT with slow kinetics. CR1CR2 was shown to be responsible for the CCAAT displacement activity of CDP/Cux. Surprisingly, the full-length CDP/Cux protein, p200, exhibited DNA binding properties similar to that of CR1CR2, indicating that CR3HD is inactive in this context. However, an amino terminally truncated 110 KDa CDP/Cux isoform capable of stable interaction with DNA was identified in cellular extracts. The p110 isoform was shown to be generated by proteolytic cleavage of p200 at the G1/S transition. The p200 and p110 isoforms displayed different transcriptional activities in reporter assays, as only p110 was able to activate transcription from the DNA polymerase a gene promoter. Interestingly, expression of p110 was found to be increased in human uterine leiomyomas as compared to the adjacent normal myometrium, raising the possibility that proteolytic processing of CDP/Cux is activated in cancer cells. Finally, cyclin D/CDK4 was shown to interact with CDP/Cux, phosphorylate it on several serine residues and inhibit proteolytic processing as well as DNA binding by CR3HD. Repla
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.38244 |
| Date | January 2002 |
| Creators | Moon, Nam Sung, 1972- |
| Contributors | Nepveu, Alain (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 Biochemistry.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001864556, proquestno: NQ78738, Theses scanned by UMI/ProQuest. |
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