Regulated cyclin-dependent kinase (CDK) levels and activities are critical for the proper progression of the cell division cycle. p12DOC-1 is a growth suppressor isolated from normal keratinocytes. We report that p12DOC-1 associates with CDK2. More specifically, p12DOC-1 associates with the monomeric nonphosphorylated form of CDK2 (p33CDK2). Ectopic expression of p12DOC-1 resulted in decreased cellular CDK2 and reduced CDK2-associated kinase activities and was accompanied by a shift in the cell cycle positions of p12DOC-1 transfectants (1G1 and2S). The p12DOC-1-mediated decrease of CDK2 was prevented if the p12DOC-1 transfectants were grown in the presence of the proteosome inhibitor clasto-lactacystin b-lactone, suggesting that p12DOC-1 may target CDK2 for proteolysis. A CDK2 binding mutant was created and was found to revert p12DOC-1-mediated, CDK2-associated cell cycle phenotypes. These data support p12DOC-1 as a specific CDK2-associated protein that negatively regulates CDK2 activities by sequestering the monomeric pool of CDK2 and/or targets CDK2 for proteolysis, reducing the active pool of CDK2.

Large scale gene expression pro

A novel p53-related gene, p73, was recently isolated and cytogenetically mapped to chromosome region 1p36. Functionally, p73 expression induces p21waf and suppresses tumor cell growth. We mapped p73 using radiation hybrids and localized the gene to an interval that putatively harbors a melanoma tumor suppressor locus. We then analyzed p73 transcripts from 24 melanoma cell lines using reverse transcription-PCR/single strand conformation polymorphism and identified nine polymorphic sequence changes (three novel and six previously published polymorphisms); furthermore, we found evidence of biallelic transcription in our cell lines. However, we did not detect any deleterious mutations. These data suggest that the p73 gene is unlikely to be essential in melanoma tumorigenesis.

We prospectively screened (through chart review) 913 consecutive patients who were seen at the Massachusetts General Hospital (MGH) Melanoma Clinic over a 6-month period (December 1997 to August 1998) and identified 519 patients with invasive melanomas. TheMGHMelanoma Clinic is a referral-based clinic for patients with a history of melanomaand high-risk individuals with a history of atypical moles or a family history of melanoma. We identified 172 patients whowere or had been diagnosed as having an invasive or metastatic melanoma before they were 40 years old. The diagnoses did not have to have been rendered during the ascertainment period; the average interval between melanoma diagnosis and the study visit was 5.9 years (range, 0 [time at diagnosis] to 30 years). Only diagnoses that were histologically confirmed by a member of the MGH Dermatopathology Unit were included. Patients with a diagnosis of melanoma in situ were excluded, given the occasional ambiguity between melanoma in situ and severely dysplastic nevus. At the time of the clinic visit, each of the 172 patients was provided with a detailed letter describing the study. Those patients expressing interest then underwent an extended discussion regarding the study content and risks. A total of 49 patients (28%) consented to the study and, in accordance with a protocol approved by the Dana Farber Cancer Institute and theMGHInstitutional Review Board, donated 20mLof blood for genetic analysis. The blood samples were coded, and the confidentiality of the patients was maintained. The results were not available to the patients. Medical and family histories and demographic details of participants were recorded through chart review and direct interview. All 49 patients were white and were unrelated to one another. Forty-three blood specimens from normal, healthy blood donors at heMGHblood bank were used for analysis of the polymorphism.