Genes VII

The common theme in the role of oncogenes in tumorigenesis is that increased or altered activity of the gene product is oncogenic. Whether the oncogene is introduced by a virus or results from a mutation in the genome, it is dominant over its allelic proto-oncogene(s). A mutation that activates a single allele is tumorigenic. Tumorigenesis then results from gain of a function.

Certain tumors are caused by a different mechanism: loss of both alleles at a locus is tumorigenic. Propensity to form such tumors may be inherited through the germline; it also occurs as the result of somatic change in the individual. Such cases identify tumor suppressors: genes whose products are needed for normal cell function, and whose loss of function causes tumors. The two best characterized genes of this class code for the proteins RB and p53.

Retinoblastoma is a human childhood disease, involving a tumor of the retina. It occurs both as a heritable trait and sporadically (by somatic mutation). It is often associated with deletions of band q14 of human chromosome 13. The RB gene has been localized to this region by molecular cloning.

Figure 28.21 summarizes the situation. Retinoblastoma arises when both copies of the RB gene are inactivated. In the inherited form of the disease, one parental chromosome carries an alteration in this region. A somatic event in retinal cells that causes loss of the other copy of the RB gene causes a tumor. In the sporadic form of the disease, the parental chromosomes are normal, and both RB alleles are lost by (individual) somatic events (Cavanee et al., 1983).

The cause of retinoblastoma is therefore loss of protein function, usually resulting from mutations that prevent gene expression (as opposed to point mutations that affect function of the protein product). Loss of RB is involved also in other forms of cancer, including osteosarcomas and small cell lung cancers.

RB is a nuclear phosphoprotein that influences the cell cycle, as discussed previously in 27 Cell cycle and growth regulation. In resting (G0/G1) cells, RB is not phosphorylated. RB is phosphorylated during the cell cycle by cyclin/cdk complexes, most particularly at the end of G1; it is dephosphorylated during mitosis. The nonphosphorylated form of RB specifically binds several proteins, and these interactions therefore occur only during part of the cell cycle (prior to S phase). Phosphorylation releases these proteins.

The target proteins include the E2F group of transcription factors, which activate target genes whose products are essential for S phase. Binding to RB inhibits the ability of E2F to activate transcription, which suggests that RB may repress the expression of genes dependent on E2F. In this way, RB indirectly prevents cells from entering S phase. Also, the RB-E2F complex directly represses some target genes, so its dissociation allows them to be expressed.

Certain viral tumor antigens bind specifically to the nonphosphorylated form of RB. The best characterized are SV40 T antigen and adenovirus E1A. This suggests the model shown in Figure 28.22. Nonphosphorylated RB prevents cell proliferation; this activity must be suppressed in order to pass through the cell cycle, which is accomplished by the cyclic phosphorylation. And it may also be suppressed when a tumor antigen sequesters the nonphosphorylated RB. Because the RB-tumor antigen complex does not bind E2F, the E2F is permanently free to allow entry into S phase (and the RB-E2F complex is not available to repress its target genes) (Sarnow et al., 1982).

Over-expression of RB impedes cell growth. An indication of the importance of RB for cell proliferation is given by the properties of an osteosarcoma cell line that lacks RB; when RB is introduced into this cell line, its growth is impeded. However, the inhibition can be overcome by expression of D cyclins, which form cdk-cyclin combinations that phosphorylate RB. RB is not the only protein of its type: proteins with related sequences, called p107 and p130, have similar properties.

The connection between the cell cycle and tumorigenesis is illustrated in Figure 28.23. Several regulators are identified as tumor suppressors by the occurrence of inactivating mutations in tumors. In addition to RB itself, there are the small inhibitory proteins (most notably p16 and possibly p21), and D cyclin(s). Although these proteins (most notably RB) play a role in the cycle of a proliferating cell, the role that is relevant for tumorigenesis is more probably their function in the quiescent (G0) state. In quiescent cells, RB is not phosphorylated, D cyclin levels are low or absent, and p16, p21, and p27 ensure inactivity of cdk-cyclin complexes. The loss of this circuit is necessary for unrestrained growth.