Young Tae Kim and Min Zhao phorylate their substrates. The cyclin D-dependent kinases (CDK4 and CDK6) can phosphorylate Rb family members (Rb, p107, and p130), thus helping to inactivate their transcriptional corepressor activities. Aberrantly expressed CDK4 could play an im- portant role in cervical tumorigenesis. It is pos- tulated that CDK4 oscillates between the INK4 and KIP inhibitors, blocking their suppressor acti- vity. In cervical cancer, the demonstrated lower levels of INK4 molecules and the high levels of CDK4 would favor binding of the more abundant KIP inhibitors to these kinases, undermining their inhibition of cyclin E. Thus, in this situation Cyclin D is expendable. The E7 would deregulate pRb initially, unleashing E2F-induced cyclin E expression; the overexpressed CDK4 would tether the KIP molecules, allowing cyclin E to become sufficiently active to phosphorylate and inactivate pRb and p27, perpetuating its own activity and that of E7.4,36,37 Yoshinouchi et al. found overexpression of CDK4 in 72.6% of cervical cancer specimens;38 this value was consistent with previous studies of cervical carcinoma.33,34 In another study, CDK4 gene amplification was described in 25% of cer- vical cancers, whereas no mutations in exon 2 of the CDK4 gene were found.34 To determine whether alterations of p16 might be involved in HPV-positive cervical cancers, Yoshinouchi looked for gene alterations and changes in the ability of the p16 protein to interact with CDK4 in 5 cervical cancer cell lines. No alteration of this gene was detected, and the p16 and CDK4 pro- teins were normally expressed. Additionally, the ability of p16 to interact with CDK4 was not abrogated in these cell lines. These cell lines were HPV-positive and carried wild-type p53 genes. These findings suggest that phosphorylation of pRb by CDK4 is not critical in the carcinogenesis or in the establishment of HPV-positive cervical cancer cell lines, since the HPV viral-transforming proteins E6 or E7 inactivate p53 and pRb tumor suppressor protein function, resulting in deregu- lated progression of the cell cycle.38 So far, re- search studies have not found a correlation between amplification and overexpression of CDK4 and patient age, histological tumor type, or tumor grade or stage.33,34 Yonsei Med J Vol. 46, No. 5, 2005 To release cells from G1 arrest and to promote entry into S-phase, pRb is phosphorylated, and thereby inactivated, by the cyclin D1/CDK4 com- plex. This inactivation may also be achieved by the interaction of pRb with the viral oncoprotein E7, leading to the hypothesis that up-regulation of positive regulators upstream of pRb might be dispensable in high-risk HPV-infected cervical carcinomas. Interestingly, very recent work has indicated that cyclin D/CDK4 complexes also phosphorylate Smad3, negatively regulating the functions of transcriptional complexes that medi- ate cell growth inhibition by proteins of the TGF family.39 Importantly, several lines of evidence indicate that cyclin D/CDK complexes play a second noncatalytic role in G1 progression by sequestering proteins of the Cip/Kip family, in- cluding p27KIP1 and p21CIP1, two potent inhibitors of CDK2.40,41 pRb/E2F The retinoblastoma tumor suppressor gene (Rb) encodes the nuclear phosphoprotein pRb (p105), which has been found mutated or deleted in several types of human cancer. p105Rb and other Rb family members p107 and p130 regulate the activity of E2F transcription factors.42 Complexes consisting of E2F and hypophosphorylated p105Rb repress the transcription of genes that are required for cell cycle progression, and repression is re- lieved by CDK-mediated phosphorylation of p105Rb.43 pRb is subject to regulation by many factors, including E2F and cyclin D1. The hypo- phosphorylated pRb, complexed with a transcrip- tion factor, serves as a transcriptional activator of cyclin D1 by binding to its promoter. On the other hand, inactivation of pRb by phosphorylation via the cyclin D/CDK complex in late G1 would not only unleash E2F transcription factors, but would also decrease cyclin D1 expression.3,44 D-Type cyclins interact with pRb pockets through their NH2-terminal L-X-C-X-E motifs. These are similar to DNA virus oncoprotein region 2, such as region 2 from HPV E7.45 Thus, the oncoprotein may occupy the pRb pocket and displace E2F factors, preventing pRb/E2F from inducing cyclin D1 transcription and undermining its normal growth- suppressive function. Some immunohistochemical studies of pRb in