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Lodish H, Berk A, Zipursky SL, et al. Molecular Cell Biology. 4th edition. New York: W. H. Freeman; 2000.

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Molecular Cell Biology. 4th edition.

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Section 24.2Proto-Oncogenes and Tumor-Suppressor Genes

As noted in the previous section, tumor cells differ from their normal counterparts in many respects: growth control, morphology, cell-to-cell interactions, membrane properties, cytoskeletal structure, protein secretion, and gene expression. We also saw that two broad classes of genes — proto-oncogenes (e.g., ras) and tumor-suppressor genes (e.g., APC) — play a key role in cancer induction. These genes encode many kinds of proteins that help control cell growth and proliferation; mutations in these genes can contribute to the development of cancer (Figure 24-9). Most cancers have inactivating mutations in one or more proteins that normally function to restrict progression through the G1 stage of the cell cycle (e.g., Rb and p16), although colon carcinomas usually do not. Virtually all human tumors have inactivating mutations in proteins such as p53 that normally function at crucial cell-cycle checkpoints, stopping the cycle if a previous step has occurred incorrectly or if DNA has been damaged. Likewise, a constitutively active Ras is found in several human tumors of different origin. Thus normal growth control and malignancy are two faces of the same coin.

Figure 24-9. The seven types of proteins that participate in controlling cell growth.

Figure 24-9

The seven types of proteins that participate in controlling cell growth. Cancer can result from expression of mutant forms of these proteins: growth factors (I), growthfactor receptors (II), signal-transduction (more...)

In this section, we describe the types of mutations that are oncogenic and how oncogenes and tumor-suppressor genes were first discovered. In later sections, we discuss specific examples of precisely how oncogenic mutations cause the abnormalities characteristic of tumor cells. Certain viruses also cause cancer, and we indicate how certain virus-encoded proteins also can subvert normal cell control mechanisms.

Gain-of-Function Mutations Convert Proto-Oncogenes into Oncogenes

Recall that an oncogene is any gene that encodes a protein able to transform cells in culture or to induce cancer in animals. Of the many known oncogenes, all but a few are derived from normal cellular genes (i.e., proto-oncogenes) whose products participate in cellular growth-controlling pathways. For example, the ras gene discussed previously is a proto-oncogene that encodes an intracellular signal-transduction protein; the mutant rasD gene derived from ras is an oncogene, whose encoded oncoprotein provides an excessive or uncontrolled growth-promoting signal. Because most proto-oncogenes are basic to animal life, they have been highly conserved over eons of evolutionary time.

Conversion, or activation, of a proto-oncogene into an oncogene generally involves a gain-of-function mutation. At least three mechanisms can produce oncogenes from the corresponding proto-oncogenes.

  • Point mutations in a proto-oncogene that result in a constitutively acting protein product
  • Localized reduplication (gene amplification) of a DNA segment that includes a proto-oncogene, leading to overexpression of the encoded protein
  • Chromosomal translocation that brings a growth-regulatory gene under the control of a different promoter and that causes inappropriate expression of the gene

An oncogene formed by the first mechanism encodes an oncoprotein that differs slightly from the normal protein encoded by the corresponding proto-oncogene. In contrast, the latter two mechanisms generate oncogenes whose protein products are identical with the normal proteins; their oncogenic effect is due to their being expressed at higher-than-normal levels or in cells where they normally are not expressed. However they arise, the gain-of-function mutations that convert proto-oncogenes to oncogenes act dominantly; that is, mutation in only one of the two alleles is sufficient for induction of cancer.

Oncogenes Were First Identified in Cancer-Causing Retroviruses

Evidence that viruses could cause cancer first came from a series of studies by Peyton Rous beginning in 1911. He excised fibrosarcomas (connective tissue tumors) from chickens, ground them up, and removed cells and debris by centrifugation. After passing the supernatant through filters with very small pores, which retained even the smallest bacteria, Rous injected the filtrate into chicks. Most of the injected chicks developed sarcomas. The transforming agent in the filtrate eventually was shown to be a virus, called Rous sarcoma virus (RSV). Some 50 years later, in 1966, Rous was awarded the Nobel prize for his pioneering work. The long delay in recognizing the importance of his discovery was due to the absence of any obvious molecular mechanism by which a virus could cause cancer, either in birds or in humans.

Later generations of molecular biologists showed that RSV is a retrovirus whose RNA genome is reversetranscribed into DNA, which is incorporated into the host-cell genome (see Figure 6-22). Nontransforming retroviruses contain the genes gag, pol, and env, which encode the virus structural proteins and the reverse transcriptase. In addition to these “normal” retroviral genes, oncogenic transforming viruses like RSV contain the v-src gene. Subsequent studies with mutant forms of RSV demonstrated that only the v-src gene, not the gag, pol, or env genes, was required for cancer induction. One revealing mutation in the v-src gene was temperature-sensitive; transformed cells were generated at 30 °C, but these cells reverted to normal morphology at 39 °C. The v-src gene thus was identified as an oncogene.

The next breakthrough came in 1977 when Michael Bishop and Harold Varmus showed that normal cells from chickens and other species contain a gene that is closely related to the RSV v-src gene. This normal cellular gene, a proto-oncogene, commonly is distinguished from the viral gene by the prefix “c” (c-src). The landmark discovery of the close relationship between a viral oncogene and cellular proto-oncogene fundamentally reoriented thinking in cancer research because it showed that cancer may be induced by the action of normal, or nearly normal, genes. RSV and other oncogenic viruses are thought to have arisen by incorporating, or transducing, a normal cellular protooncogene into their genome. Subsequent mutation in the transduced gene then converted it into an oncogene.

As discussed below, v-Src protein is a constitutively active mutant form of c-Src protein, a protein-tyrosine kinase. In cells containing an integrated RSV genome, not only is v-src transcribed at inappropriately high rate levels, but the unregulated activity of v-Src protein causes continuous and inappropriate phosphorylation of target proteins. Because v-src can induce cell transformation in the presence of the normal c-src proto-oncogene, v-src is said to be a dominant gain-of-function mutant of c-src, analogous to the rasD form of the ras proto-oncogene discussed previously. Many other oncogenes derived from cellular proto-oncogenes have been found in different retroviruses, implying that the normal vertebrate genome contains many potential cancer-causing genes.

Earlier we described the critical DNA transfection experiment establishing the existence of dominant gain-of-function oncogenes in human bladder tumors (see Figure 24-4), which led to the molecular cloning of a ras gene with a single point mutation. This oncogene, designated Ha-ras, also is present in Harvey sarcoma virus, a retrovirus. Similar experiments with DNA from many other tumors, both human and experimental, have led to the cloning of numerous oncogenes from tumor-cell DNA. Many of these cancer-causing genes are also found in various animal retroviruses.

Slow-Acting Carcinogenic Retroviruses Can Activate Cellular Proto-Oncogenes

Because its genome carries the v-src oncogene, Rous sarcoma virus induces tumors within days. Most oncogenic retroviruses, however, induce cancer only after a period of months or years. The genomes of the slow-acting retroviruses differ from those of transducing viruses such as RSV in one crucial respect: they lack an oncogene. Thus, slow-acting, or “long latency,” retroviruses have no direct affect on growth of cells in culture.

The mechanism by which avian leukosis viruses cause cancer appears to operate in all slow-acting retroviruses. Like other retroviruses, avian leukosis virus DNA generally integrates into cellular chromosomes more or less at random. However, the finding that the site of integration in the cells from tumors caused by these viruses is near the c-myc gene suggested that these slow-acting viruses cause disease by activating expression of c-Myc. As noted earlier, c-Myc is required for transcription of many genes that encode cellcycle proteins. These viruses act slowly both because integration near c-myc is a random, rare event and because additional mutations have to occur before a full-fledged tumor becomes evident.

In some tumors, the avian leukosis proviral DNA is found at the 5′ end of the myc gene in the same transcriptional orientation. In such cases, the right-hand LTR of the integrated retrovirus — which usually serves as a terminator — is believed to act as a promoter, initiating synthesis of RNA transcripts from the c-myc gene (Figure 24-10a). In other tumors, the proviral DNA is found in the opposite transcriptional orientation; in this case, it is thought to exert an indirect enhancer activity (Figure 24-10b). Whether the inserted proviral DNA acts as a promoter or enhancer of c-myc transcription, the expressed c-Myc protein apparently is perfectly normal. The enhanced level of c-Myc resulting from the strong promoting or enhancing activity of the retroviral LTR partly explains the oncogenic effect of avian leukosis viruses. A second aspect is that c-myc expression is usually down-regulated when cells are induced to differentiate, but the LTR-driven expression of c-myc does not respond to such signals, and thus cells that normally would differentiate instead undergo DNA replication and cell division. These mechanisms of oncogene activation — called promoter insertion and enhancer insertion — operate in a variety of oncogenes and have been implicated in many animal tumors induced by slow-acting retroviruses.

Figure 24-10. Activation of the c-myc proto-oncogene by retroviral promoter and enhancer insertions.

Figure 24-10

Activation of the c-myc proto-oncogene by retroviral promoter and enhancer insertions. (a) The promoter can be activated when the retrovirus inserts upstream (5′) of the c-myc exons. (more...)

In natural bird and mouse populations, slow-acting retroviruses are much more common than oncogenecontaining retroviruses such as Rous sarcoma virus. Thus, insertional oncogene activation is probably the major mechanism whereby retroviruses cause cancer.

Many DNA Viruses Also Contain Oncogenes

Most animal cells infected by small DNA viruses such as SV40 are killed, but a very small proportion integrate the viral DNA into the host-cell genome. Although these cells survive infection, they become permanently transformed because the viral DNA contains one or more oncogenes. For example, many warts and other benign tumors of epithelial cells are caused by the DNA-containing papillomaviruses. Human genital warts are caused by one such virus that can induce stable transformation and transient mitogenic stimulation of a variety of cultured cells.

Unlike retroviral oncogenes, which are derived from normal cellular genes and have no function for the virus, the known oncogenes of DNA viruses are integral parts of the viral genome required for viral replication. As discussed later, the oncoproteins expressed from integrated viral DNA in infected cells act in various ways to stimulate cell growth and proliferation.

Loss-of-Function Mutations in Tumor-Suppressor Genes Are Oncogenic

Tumor-suppressor genes generally encode proteins that in one way or another inhibit cell proliferation. Loss of one or more of these “brakes” contributes to the development of many cancers. Five broad classes of proteins are generally recognized as being encoded by tumor-suppressor genes:

  • Intracellular proteins, such as the p16 cyclin-kinase inhibitor, that regulate or inhibit progression through a specific stage of the cell cycle
  • Receptors for secreted hormones (e.g., tumorderived growth factor β) that function to inhibit cell proliferation
  • Checkpoint-control proteins that arrest the cell cycle if DNA is damaged or chromosomes are abnormal
  • Proteins that promote apoptosis
  • Enzymes that participate in DNA repair

Although DNA-repair enzymes do not directly function to inhibit cell proliferation, cells that have lost the ability to repair errors, gaps, or broken ends in DNA accumulate mutations in many genes, including those that are critical in controlling cell growth and proliferation. Thus loss-of-function mutations in the genes encoding DNA-repair enzymes promote inactivation of other tumor-suppressor genes as well as activation of oncogenes.

Since generally one copy of a tumor-suppressor gene suffices to control cell proliferation, both alleles of a tumor-suppressor gene must be lost or inactivated in order to promote tumor development. Thus oncogenic loss-of-function mutations in tumor-suppressor genes act recessively. Tumor-suppressor genes in many cancers have deletions or point mutations that prevent production of any protein or lead to production of a nonfunctional protein.

The First Tumor-Suppressor Gene Was Identified in Patients with Inherited Retinoblastoma

We saw earlier that individuals who inherit a mutant allele of APC, a tumor-suppressor gene, have a high risk of developing colon cancer. Inheriting one mutant allele of another tumor-suppressor gene increases to almost 100 percent the probability that a person will develop a specific tumor. Indeed, genetic studies on cancer-prone families led to the initial identification of many tumor-suppressor genes. A classic case is retinoblastoma, which is caused by loss of function of RB, the first tumor-suppressor gene to be identified. As discussed later, the protein encoded by RB helps regulate progress through the cell cycle.

Children with hereditary retinoblastoma inherit a single defective copy of the RB gene, sometimes seen as a small deletion on chromosome 13. They develop retinal tumors early in life and generally in both eyes (Figure 24-11). Each tumor that develops is derived from a single transformed cell. The developing retina contains about 4×106 cells, but only about 1 in 106 cells actually gives rise to a tumor cell. This finding shows that the defective RB allele is acting recessively at the cellular level, and that other genetic events are needed to bring on the transformed state. One essential event is the deletion or mutation of the normal RB gene on the other chromosome, giving rise to a cell that produces no functional Rb protein (see Figure 8-7). Individuals with sporadic retinoblastoma, in contrast, inherit two normal RB alleles each of which has undergone a somatic loss-offunction mutation in a single retinal cell. Because this is an unlikely occurrence, sporadic retinoblastoma is rare, develops late in life, and usually affects only one eye.

Figure 24-11. Children with hereditary retinoblastoma develop retinal tumors early in life and generally in both eyes.

Figure 24-11

Children with hereditary retinoblastoma develop retinal tumors early in life and generally in both eyes. They inherit one mutant allele of the RB gene. Somatic mutation of the other allele (more...)

If retinal tumors are removed before they become malignant, children with hereditary retinoblastoma often survive until adulthood and produce children. Molecular cloning of the RB gene established that these individuals inherited one normal and one mutant RB allele. On average, they will pass on the mutant allele to half their children and the normal allele to the other half. Children who inherit the normal allele are normal if their other parent has two normal RB alleles. However, those who inherit the mutant allele have the same enhanced predisposition to develop retinal tumors as their affected parent, even though they inherit a normal RB allele from their other, normal parent. Thus hereditary retinoblastoma is inherited as an autosomal dominant trait (Figure 24-12). As discussed below, many human tumors (not just retinal tumors) contain mutant RB alleles; most of these arise as the result of somatic mutations.

Figure 24-12. Hereditary retinoblastoma is inherited as an autosomal dominant trait, as illustrated in this pedigree.

Figure 24-12

Hereditary retinoblastoma is inherited as an autosomal dominant trait, as illustrated in this pedigree. Affected individuals (red), who inherit one mutant allele of RB, a tumor-suppressor (more...)

A similar hereditary predisposition for breast cancer has been linked to BRCA1, another tumor-suppressor gene. Women who inherit one mutant BRCA1 allele have a 60 percent probability of developing breast cancer by age 50, whereas those who inherit two normal BRCA1 alleles have only a 2 percent probability of doing so. In women with hereditary breast cancer, loss of the second BRCA1 allele, together with other mutations, is required for a normal breast duct cell to become malignant. However, BRCA1 generally is not mutated in sporadic, noninherited breast cancer.

Loss of Heterozygosity of Tumor-Suppressor Genes Occurs by Mitotic Recombination or Chromosome Mis-segregation

In a somatic cell that contains one mutant and one normal allele of a tumor-suppressor gene, how is the normal allele lost or inactivated? That is, what mechanisms can result in loss of heterozygosity (LOH) of the normal allele? Point mutations are an unlikely cause because any such mutations that occur in the normal allele usually are repaired except in cells that are defective in certain DNA-repair systems (Chapter 12).

One common mechanism for LOH involves missegregation of the chromosomes bearing the heterozygous tumor-suppressor gene during mitosis. In this process, also referred to as nondisjunction, one daughter cell inherits only one normal chromosome (and probably dies), while the other inherits three, the other normal chromosome as well as two bearing the mutant allele. Such mis-segregation is caused by failure of a mitotic checkpoint, which would normally prevent a metaphase cell with an abnormal mitotic spindle from completing mitosis (see Figure 13-34). Subsequent loss of one chromosome often occurs, restoring the 2n complement; if the normal chromosome is lost, the resultant cell will contain two copies of the “mutant” chromosome (Figure 24-13a).

Figure 24-13. Loss of heterozygosity (LOH) of tumorsuppressor genes.

Figure 24-13

Loss of heterozygosity (LOH) of tumorsuppressor genes. A cell containing one normal and one mutant allele of a tumor-suppressor gene is generally phenotypically normal. (a) If formation of (more...)

Another likely mechanism for LOH is mitotic recombination between a chromatid bearing the wild-type allele and a homologous chromatid bearing a mutant allele. As illustrated in Figure 24-13b, the products of such a recombination are two normal chromatids and two mutant chromatids. Subsequent chromosome segregation can generate three types of daughter cells: one homozygous for the mutant tumor-suppressor allele; one homozygous for the normal allele; and one like the parental cell, heterozygous for the mutant allele.

SUMMARY

  •  Dominant gain-of-function mutations in protooncogenes and recessive loss-of-function mutations in tumor-suppressor genes are oncogenic.
  •  Among the proteins encoded by proto-oncogenes are positive-acting growth factors and their receptors, signal-transduction proteins, transcription factors, and cell-cycle control proteins (see Figure 24-9).
  •  An activating mutation of one of the two alleles of a proto-oncogene converts it to an oncogene, which can induce transformation in cultured cells or cancer in animals.
  •  Activation of a proto-oncogene into an oncogene can occur by point mutation, gene amplification, and gene translocation.
  •  The first recognized oncogene, v-src, was identified in Rous sarcoma virus, a cancer-causing retrovirus. Retroviral oncogenes arose by transduction of cellular proto-oncogenes into the viral genome and subsequent mutation.
  •  The first human oncogene to be identified encodes a constitutively active form of Ras, a signal-transduction protein. This oncogene was isolated from a human bladder carcinoma (see Figure 24-4).
  •  Slow-acting retroviruses can cause cancer by integrating near a proto-oncogene in such a way that gene transcription is activated continuously and inappropriately.
  •  Tumor-suppressor genes encode proteins that slow or inhibit progression through a specific stage of the cell cycle, checkpoint-control proteins that arrest the cell cycle if DNA is damaged or chromosomes are abnormal, receptors for secreted hormones that function to inhibit cell proliferation, proteins that promote apoptosis, and DNA repair enzymes.
  •  Inherited mutations causing retinoblastoma led to the identification of RB, the first tumor-suppressor gene to be recognized.
  •  Inheritance of a single mutant allele of many tumor-suppressor genes (e.g., RB, APC, and BRCA1) increases to almost 100 percent the probability that a specific kind of tumor will develop.
  •  Loss of heterozygosity of tumor-suppressor genes occurs by mitotic recombination or chromosome missegregation (see Figure 24-13).

By agreement with the publisher, this book is accessible by the search feature, but cannot be browsed.

Copyright © 2000, W. H. Freeman and Company.
Bookshelf ID: NBK21662