Fbw7/hCdc4/Ago and Endometrial Cancer Ashmita Chatterjee Biology 169 April 4, 2006

Fbw7/hCdc4/Ago Is the key protein that targets cyclin E for destruction Is the key protein that targets cyclin E for destruction Belongs to a large family of F-box proteins Belongs to a large family of F-box proteins F-box proteins are a subunit of E3, the ubiquitin ligase F-box proteins are a subunit of E3, the ubiquitin ligase

Cell Proliferation Control Done by modulating the abundance of cyclins Done by modulating the abundance of cyclins Achieved by alternating transcription with their degradation Achieved by alternating transcription with their degradation Ensures that each cyclin reaches the necessary threshold to activate its CDK Ensures that each cyclin reaches the necessary threshold to activate its CDK

A common example: Cyclin E Typically low in Early G1 Typically low in Early G1 Rises to a peak in late G1 Rises to a peak in late G1 Activates CDK2 around the G1-S transition Activates CDK2 around the G1-S transition Falls again during S, G2, and M phases Falls again during S, G2, and M phasesTherefore….. Cyclin E-CDK2 functions in promoting the initiation of genome replication during S-phase

And as we studied in class… Insufficient cyclin E results in cell arrest in G1 Insufficient cyclin E results in cell arrest in G1 Too much cyclin E leads to premature entry into S phase, genomic instability, and the formation of tumors Too much cyclin E leads to premature entry into S phase, genomic instability, and the formation of tumors

Ubiquitin Mediated Proteolysis Courtesy Dr. Duronio 2005

How hCdc4 Functions Schwab and Tyers 2001

hCdc4 binds to Phosphorylated Cyclin E Rajagopalan et al. 2004

Evidence of Important Biological Role Koepp et al. 2001

Drosophila Mutants Dramatic eye and wing phenotype Dramatic eye and wing phenotype Have persistently elevated levels of cyclin E protein without increased levels of cyclin E RNA Have persistently elevated levels of cyclin E protein without increased levels of cyclin E RNA

Mouse Knockouts Heterozygous: Heterozygous: Normal, no spontaneous tumor formation Homozygous Mutant: Homozygous Mutant: Died 10.5 days postcoitus Tetzlaff et al. 2004

Heart Formation in KO Mice A-D: WT Mice A-D: WT Mice E-H: KO Mice E-H: KO Mice K- WT Heart- well differentiated K- WT Heart- well differentiated L-KO Heart- linear heart tube L-KO Heart- linear heart tube Tetzlaff et al. 2004

WT and KO Cell Phenotypes Mutant cells contain more cyclin E and increased DNA synthesis Mutant cells contain more cyclin E and increased DNA synthesis Tetzlaff et al. 2004

Linking hCdc4 to Cancer Hubalek et al Bartek and Lukas 2001

Linking hCdc4 to Cancer hCdc4 is therefore a tumor-suppressor hCdc4 is therefore a tumor-suppressor Have not yet identified mutations in the Fbw7 gene in tumors Have not yet identified mutations in the Fbw7 gene in tumors However, Fbw7 maps to 4q32, a site of loss of heterozygosity in 30% of all cancers However, Fbw7 maps to 4q32, a site of loss of heterozygosity in 30% of all cancers

Endometrial Cancer originates in the inner lining of the uterus (which is known as the endometrium) originates in the inner lining of the uterus (which is known as the endometrium) accounts for about 90% of all uterine cancers accounts for about 90% of all uterine cancers Uterine cancer is the most common type of cancer that originates in the female reproductive system Uterine cancer is the most common type of cancer that originates in the female reproductive system

Types of Endometrial Cancer Adenocarcinoma Adenocarcinoma Papillary Serous Carcinoma Papillary Serous Carcinoma Clear Cell Carcinoma Clear Cell Carcinoma

Incidence and Prevalence In the US each year: In the US each year: - 37,000 diagnoses - 6,000 deaths - 75% of cases in postmenopausal women - average age at diagnosis: 60 years More common in Caucasian women than African American More common in Caucasian women than African American

Current Treatments Surgery Surgery Radiation Therapy Radiation Therapy Hormone Therapy Hormone Therapy Chemotherapy Chemotherapy

Future Treatments? Inhibitors of cyclin E-CDK2 Inhibitors of cyclin E-CDK2 Modulators of ubiquitin-dependent proteolysis Modulators of ubiquitin-dependent proteolysis

References Bartek, Jiri, and Jiri Lukas. “Order From Destruction.” Science 294 (2001): Bartek, Jiri, and Jiri Lukas. “Order From Destruction.” Science 294 (2001): Hubalek, Michael, A. Widschwendter, M. Erdel, A. Gschwendtner, H. Fiegl, H. Müller, G. Goebel, E. Mueller-Holzner, C. Marth, C. Spruck, S.Reed, and M. Widschwendter. “Cyclin E dysregulation and chromosomal instability in endometrial cancer.” Oncogene 23 (2004): Hubalek, Michael, A. Widschwendter, M. Erdel, A. Gschwendtner, H. Fiegl, H. Müller, G. Goebel, E. Mueller-Holzner, C. Marth, C. Spruck, S.Reed, and M. Widschwendter. “Cyclin E dysregulation and chromosomal instability in endometrial cancer.” Oncogene 23 (2004): Koepp, Deanna, L. Schaefer, X. Ye, K. Keyomarsi, C. Chu, J. Harper, and S. Elledge. “Phosphorylation-Dependent Ubiquitination of Cyclin E by the SCF Fbw7 Ubiquitin Ligase.” Science 294 (2001): Koepp, Deanna, L. Schaefer, X. Ye, K. Keyomarsi, C. Chu, J. Harper, and S. Elledge. “Phosphorylation-Dependent Ubiquitination of Cyclin E by the SCF Fbw7 Ubiquitin Ligase.” Science 294 (2001): Moberg, Kenneth, D. Bell, D. Wahrer, D. Haber, and I. Hariharan. “Archipelago regulates Cyclin E levels in Drosophila and is mutated in human cancer cell lines.” Nature 413 (2001): Moberg, Kenneth, D. Bell, D. Wahrer, D. Haber, and I. Hariharan. “Archipelago regulates Cyclin E levels in Drosophila and is mutated in human cancer cell lines.” Nature 413 (2001): Rajagopalan, Harith, P. Jallepalli, C. Rago, V. Velculescu, K. Kinzler, B. Vogelstein, and C. Lengauer. “Inactivation of hCDC4 can cause chromosomal instability.” Nature 428 (2004): Rajagopalan, Harith, P. Jallepalli, C. Rago, V. Velculescu, K. Kinzler, B. Vogelstein, and C. Lengauer. “Inactivation of hCDC4 can cause chromosomal instability.” Nature 428 (2004): Reed, Steven. “Ratchets and Clocks: The Cell Cycle, Ubiquitylation, and Protein Turnover.” Molecular Cell Biology 4 (2003): Reed, Steven. “Ratchets and Clocks: The Cell Cycle, Ubiquitylation, and Protein Turnover.” Molecular Cell Biology 4 (2003): Schwab, Michael, and Mike Tyers. “Cell cycle: Archipelago of destruction.” Nature 413 (2001): Schwab, Michael, and Mike Tyers. “Cell cycle: Archipelago of destruction.” Nature 413 (2001): Tetzlaff, Michael, W. Yu, M. Li, P. Zhang, M. Finegold, K. Mahon, J. Harper, R. Schwartz, and S. Elledge. “Defective cardiovascular development and elevated cyclin E and Notch proteins in mice lacking the Fbw7 F-box protein.” PNAS 101 (2004): Tetzlaff, Michael, W. Yu, M. Li, P. Zhang, M. Finegold, K. Mahon, J. Harper, R. Schwartz, and S. Elledge. “Defective cardiovascular development and elevated cyclin E and Notch proteins in mice lacking the Fbw7 F-box protein.” PNAS 101 (2004):