Functional Consequences of Alterations in Cell Cycle Components

29 May

In general, as shown from studies in yeast (Hartwell and Kastan, 1994), significant alterations in the expression of any one of the components of the cell cycle may, over time, lead to derepres- sion of the cycle, aneuploidy, and chromosomal abnormalities. However, in most instances one sees abnormalities in more than one component of the cell cycle within any neoplastic cell. Sim- ple overexpression of cyclins D1 or E leads to accelerated transition through G1, reduced growth factor dependency.  However, in the case of these two cyclins, simple overexpression  does not appear to result in cell transformation in most instances (cf. Michalides, 1999), although associ- ation with alterations in p53 may do so. Amplification of cyclin D1 is a very common event in many human neoplasms (Hall and Peters, 1996).

Alterations in cyclin kinase, especially Cdk4, as well as cyclin-dependent  kinase inhibi- tors, especially p16, have also been characterized  in a number of human neoplasms (Hall and Peters, 1996). However, several of the cyclin kinase inhibitors such as p21 and p27 have rarely if at all been found in a mutation form in neoplasms. In human ovarian carcinoma, a lack of muta- tion of the p16 gene was found, although there was expression of aberrant p16 RNA transcripts (Suh et al., 2000). On the other hand, since p21 expression is regulated by p53 and mutations in p53 are found in a number of malignancies, loss of p53 function may result in loss of the normal functioning of p21 and other checkpoint control components (Orlowski and Furlanetto, 1996). Despite the lack of mutations in p27 found in human cancer, deletions of this gene have been seen in a number of aggressive carcinomas in the human (cf. Millard and Koff, 1998). Like p16, p18 is commonly mutated in a number of human neoplasms (cf. Millard and Koff, 1998).

As noted earlier (Chapter 6), a variety of “stress signals”— including DNA damage, hy- poxia, etc.—lead to activation of the p53 tumor suppressor gene. This, in turn, can potentially lead to a variety of different functions, as noted in Figure 15.5. E2F-1 induces p53 expression which, like DNA damage via the DNA kinase, may cause arrest of the cell cycle in normal cells.

The p19ARF  protein interacts with E2F-1 and is important  in the transcriptional  activation  by E2F-1. Furthermore, p53 induces mdm-2, which subsequently binds and destabilizes p53 in the feedback loop noted in Figure 15.5. Although mdm-2 has been termed a proto-oncogene, it was originally discovered as one of several genes in a transformed cell line resulting from amplifica- tion of double minute particles, from which the terminology mouse double minute (mdm) was derived. The mdm-2 protein physically associates with the p53 tumor suppressor protein, block- ing its transactivational  effects. In addition,  it seems to stimulate  degradation  of the protein (Haines, 1997). mdm gene amplification  is quite common in a number of neoplasms (Haines,

1997; Flørenes et al., 1994). E2F-1 enhances the expression of p53 and also neutralizes mdm-2 through its stimulation of the expression of p19/arf (cf. Michalides, 1999). The ATM gene codes for a multifunctional  enzyme having various kinase activities and involved in the response to DNA damage (Lavin and Shiloh, 1997). p53 also transactivates both the p21 cyclin-dependent inhibitor  as well as BAX, which enhances  apoptosis  (cf. Brady and Gil-Gómez,  1998). En- hanced expression of BAX has been noted in prostatic neoplasia (Johnson et al., 1998), but low levels of BAX expression have been associated with a poor clinical outcome in breast cancer (Krajewski et al., 1995). Alternatively, p53 may induce apoptosis in cells. Mutations in this tu- mor suppressor  gene thus lead to a lack of apoptotic response to DNA damage in neoplastic cells, which is quite commonly seen in many neoplasms.

Cell Cycle Checkpoints in Neoplasia

As noted in Figures 15.4 and 9.5, the checkpoints of the cycle occur at the G1/S interface, at the end of G2, and in M. Defects in each of these checkpoints have been noted in neoplasia. In addi- tion, the restriction (R) point at which the decision is made whether to enter the mitotic cycle is another component that may be defective in neoplastic cells. The (R) and G1/S checkpoint de- fects may be in part due to the overexpression of D and E-type cyclins seen in so many neoplasms (Hunter and Pines, 1994). Defects in p53 also result in defective G1/S checkpoint  function in neoplastic cells (cf. Sherr, 1996) as well as genetic instability (cf. Morgan and Kastan, 1997). Defects in the cyclin B-Cdk1 (Cdc2) phosphorylation,  resulting in activation of the complex in neoplastic cells, allow the neoplastic cell to enter mitosis with damaged DNA (cf. Hunter and Pines, 1994). Mutations in the Bub1 gene (Figure 15.7), which controls the mitotic checkpoint and chromosome segregation, are seen in a variety of human neoplasms (Cahill et al., 1998). Al- terations in this gene and related genes involved in the completion of normal mitosis can lead to aneuploidy, a hallmark characteristic of cells in the stage of progression (cf. Molinari, 2000).

Figure 15.6 Mechanisms  for potential  roles of DNA methylation  in neoplastic  development.  Unfilled circles denote unmethylated  CpGs; filled circles indicate methylated CpGs. a. Aberrant hypermethylation of CpG islands could result in their transcriptional  silencing  or repression.  In all examples,  the gene is indicated by the filled rectangle. b. Hypomethylation  of CpG dinucleotides in protooncogene  promoter re- gions may facilitate an increased expression of the proto-oncogene.  (Adapted from Laird, 1997, with per- mission of author and publisher.)

Thus, it may be noted that numerous defects occur in the cell cycle of neoplastic cells and contribute ultimately to the evolving karyotypic instability characteristic of the stage of progres- sion (Chapter 9). Since, as noted earlier, enhanced mitotic activity may lay the foundation for enhanced aberrations in the cell cycle on the basis of mutation and/or selection, other mecha- nisms of regulating DNA expression may be found to be abnormal. Knowledge and understand- ing of such mechanisms  may allow the elucidation  of the transition  of preneoplastic  cells exhibiting few if any mutational alterations into neoplastic cells in the stage of progression that exhibit the profound mutational alterations discussed above. One such mechanism may be found in the alterations of DNA methylation, discussed below.

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