6. Oncogenes code for components of signal transduction cascades

28.5 Insertion, translocation, or amplification may activate proto-oncogenes

Key terms defined in this section
Reciprocal translocation exchanges part of one chromosome with part of another chromosome.

A variety of genomic changes can activate proto-oncogenes, sometimes involving a change in the target gene itself, sometimes activating it without changing the protein product. Insertion, translocation, and amplification can be causative events in tumorigenesis.




Figure 17.29 Amplified copies of the dhfr gene produce a homogeneously staining region (HSR) in the chromosome. Photograph kindly provided by Robert Schimke.


Figure 17.30 Amplified extrachromosomal dhfr genes take the form of double-minute chromosomes, as seen in the form of the small white dots. Photograph kindly provided by Robert Schimke.

Many tumor cell lines have visible regions of chromosomal amplification, as shown by homogeneously staining regions (see Figure 17.29) or double minute chromosomes (see Figure 17.30). The amplified region may include an oncogene. Examples of oncogenes that are amplified in various tumors include c-myc, c-abl, c-myb, c-erbB, c-K-ras, and Mdm2.


Established cell lines are prone to amplify genes (along with other karyotypic changes to which they are susceptible). The presence of known oncogenes in the amplified regions, and the consistent amplification of particular oncogenes in many independent tumors of the same type, strengthens the correlation between increased expression and tumor growth.


Some proto-oncogenes are activated by events that change their expression, but which leave their coding sequence unaltered. The best characterized is c-myc, whose expression is elevated by several mechanisms. One common mechanism is the insertion of a nondefective retrovirus in the vicinity of the gene.


The ability of a retrovirus to transform without expressing a v-onc sequence was first noted during analysis of the bursal lymphomas caused by the transformation of B lymphocytes with avian leukemia virus. Similar events occur in the induction of T-cell lymphomas by murine leukemia virus. In each case, the transforming potential of the retrovirus is due to the ability of its LTR (the long terminal repeat of the integrated form) to cause expression of cellular gene(s).




Figure 28.11 Insertions of ALV at the c-myc locus occur at various positions, and activate the gene in different ways.

In many independent tumors, the virus has integrated into the cellular genome within or close to the c-myc gene. Figure 28.11 summarizes the types of insertions. The retrovirus may be inserted at a variety of locations relative to the c-myc gene.


The gene consists of three exons; the first represents a long nontranslated leader, and the second two code for the c-Myc protein. The simplest insertions to explain are those that occur within the first intron. The LTR provides a promoter, and transcription reads through the two coding exons. Transcription of c-myc under viral control differs from its usual control: the level of expression is increased (because the LTR provides an efficient promoter); expression cannot be switched off in B or T cells in response to the usual differentiation signals; and the transcript lacks its usual nontranslated leader (which may usually limit expression). All of these changes add up to increased constitutive expression.


Activation of c-myc in the other two classes of insertions reflects different mechanisms. The retroviral genome may be inserted within or upstream of the first intron, but in reverse orientation, so that its promoter points in the wrong direction. The retroviral genome also may be inserted downstream of the c-myc gene. In these cases, the enhancer in the viral LTR may be responsible for activating transcription of c-Myc, either from its normal promoter or from a fortuitous promoter.


In all of these cases, the coding sequence of c-myc is unchanged, so oncogenicity is attributed to the loss of normal control and increased expression of the gene.


Other oncogenes that are activated in tumors by the insertion of a retroviral genome include c-erbB, c-myb, c-mos, c-H-ras, and c-raf. Up to 10 other cellular genes (not previously identified as oncogenes by their presence in transforming viruses) are implicated as potential oncogenes by this criterion. The best characterized among this latter class are wnt1 and int2. The wnt1 gene codes for a protein involved in early embryogenesis that is related to the wingless gene of Drosophila; int2 codes for an FGF (fibroblast growth factor).




Figure 28.12 A chromosomal translocation is a reciprocal event that exchanges parts of two chromosomes. Translocations that activate the human c-myc proto-oncogene involve Ig loci in B cells and TcR loci in T cells.

Translocation to a new chromosomal location is another of the mechanisms by which oncogenes are activated. A reciprocal translocation occurs when an illegitimate recombination occurs between two chromosomes as illustrated in Figure 28.12. The involvement of such events in tumorigenesis was discovered via a connection between the loci coding immunoglobulins and the occurrence of certain tumors. Specific chromosomal translocations are often associated with tumors that arise from undifferentiated B lymphocytes. The common feature is that an oncogene on one chromosome is brought by translocation into the proximity of an Ig locus on another chromosome. Similar events occur in T lymphocytes to bring oncogenes into the proximity of a TcR locus (for review see Showe and Croce, 1987).


In both man and mouse, the nonimmune partner is often the c-myc locus. In man, the translocations in B-cell tumors usually involve chromosome 8, which carries c-myc, and chromosome 14, which carries the IgH locus; ~10% involve chromosome 8 and either chromosome 2 (kappa locus) or chromosome 22 (lambda locus). The translocations in T-cell tumors often involve chromosome 8, and either chromosome 14 (which has the TcR α locus at the other end from the Ig locus) or chromosome 7 (which carries TcR β locus). Analogous translocations occur in the mouse.


Translocations in B cells fall into two classes, reflecting the two types of recombination that occur in immunoglobulin genes. One type is similar to those involved in somatic recombination to generate the active genes, involving the consensus sequences used for V-D-J recombination. These can occur at all the Ig loci. In the other type, the translocation occurs at a switching site at the IgH locus, presumably reflecting the operation of the system for class switching.


When c-myc is translocated to the Ig locus, its level of expression is usually increased. The increase varies considerably among individual tumors, generally being in the range from 2 V10 . Why does translocation activate the c-myc gene? The event has two consequences: c-myc is brought into a new region, one in which an Ig or TcR gene was actively expressed; and the structure of the c-myc gene may itself be changed (but usually not involving the coding regions). It seems likely that several different mechanisms can activate the c-myc gene in its new location (just as retroviral insertions activate c-myc in a variety of ways).


The correlation between the tumorigenic phenotype and the activation of c-myc by either insertion or translocation suggests that continued high expression of c-Myc protein is oncogenic. Expression of c-myc must be switched off to enable immature lymphocytes to differentiate into mature B and T cells; failure to turn off c-myc maintains the cells in the undifferentiated (dividing) state.


The oncogenic potential of c-myc has been demonstrated directly by the creation of transgenic mice. Mice carrying a c-myc gene linked to a B lymphocyte-specific enhancer (the IgH enhancer) develop lymphomas. The tumors represent both immature and mature B lymphocytes, suggesting that over-expression of c-myc is tumorigenic throughout the B cell lineage. Transgenic mice carrying a c-myc gene under the control of the LTR from a mouse mammary tumor virus, however, develop a variety of cancers, including mammary carcinomas. This suggests that increased or continued expression of c-myc transforms the type of cell in which it occurs into a corresponding tumor (for review see Cory and Adams, 1988; Adams and Cory, 1991).


c-myc exhibits three means of oncogene activation: retroviral insertion, chromosomal translocation, and gene amplification. The common thread among them is deregulated expression of the oncogene rather than a qualitative change in its coding function, although in at least some cases the transcript has lost the usual (and possibly regulatory) nontranslated leader. c-myc provides the paradigm for oncogenes that may be effectively activated by increased (or possibly altered) expression.


Translocations are now known in many types of tumors. Often a specific chromosomal site is commonly involved, creating the supposition that a locus at that site is involved in tumorigenesis. However, every translocation generates reciprocal products; sometimes a known oncogene is activated in one of the products, but in other cases it is not evident which of the reciprocal products has responsibility for oncogenicity. Also, it is not axiomatic that the gene(s) at the breakpoint have responsibility; for example, the translocation could provide an enhancer that activates another gene nearby.


A variety of translocations found in B and T cells have identified new oncogenes. In some cases, the translocation generates a hybrid gene, in which an active transcription unit is broken by the translocation. This has the result that the exons of one gene may be connected to another. In such cases, there are two potential causes of oncogenicity The proto-oncogene part of the protein may be activated in some way that is independent of the other part, for example, because it is over-expressed under its new management (a situation directly comparable to the example of c-myc). Or the other partner in the hybrid gene may have some positive effect that generates a gain-of-function in the part of the protein coded by the proto-oncogene.


One of the best characterized cases in which a translocation creates a hybrid oncogene is provided by the Philadelphia (PH1) chromosome present in patients with chronic myelogenous leukemia (CML). This reciprocal translocation is too small to be visible in the karyotype, but links a 5000 kb region from the end of chromosome 9 carrying c-abl to the bcr gene of chromosome 22. The bcr (breakpoint cluster region) was originally named to describe a region of ~5.8 kb within which breakpoints occur on chromosome 22.




Figure 28.13 Translocations between chromosome 22 and chromosome 9 generate Philadelphia chromosomes that synthesize bcr-abl fusion transcripts that are responsible for two types of leukemia.

The consequences of this translocation are summarized in Figure 28.13. The bcr region lies within a large (>90 kb) gene, which is now known as the bcr gene. The breakpoints in CML usually occur within one of two introns in the middle of the gene. The same gene is also involved in translocations that generate another disease, ALL (acute lymphoblastic leukemia); in this case, the breakpoint in the bcr gene occurs in the first intron.


The c-abl gene is expressed by alternative splicing that uses either of the first two exons. The breakpoints in both CML and ALL occur in the intron that precedes the first common exon. Although the exact breakpoints on both chromosomes 9 and 22 vary in individual cases, the common outcome is the production of a transcript coding for a Bcr-Abl fusion protein, in which N-terminal sequences derived from bcr are linked to c-abl sequences. In ALL, the fusion protein has ~45 kD of the Bcr protein; in CML the fusion protein has ~70 kD of the Bcr protein. In each case, the fusion protein contains ~140 kD of the usual ~145 kD c-Abl protein, that is, it has lost just a few N-terminal amino acids of the c-Abl sequence.


Why is the fusion protein oncogenic? The Bcr-Abl protein appears to activate the Ras pathway for transformation. It may have multiple ways of doing so, including activation of the adaptors Grb2 and Shc (see 26 Signal transduction). Both the Bcr and Abl regions of the joint protein may be important in transforming activity.


Changes at the N-terminus are involved in activating the oncogenic activity of v-abl, a transforming version of the gene carried in a retrovirus. The c-abl gene codes for a tyrosine kinase activity; this activity is essential for transforming potential in oncogenic variants. Deletion (or replacement) of the N-terminal region activates the kinase activity and transforming capacity. So the N-terminus provides a domain that usually regulates kinase activity; its loss may cause inappropriate activation.



Reviews
Adams, J. M. and Cory, S. (1991). Transgenic models of tumor development. Science 254, 1161-1167.
Cory, S. and Adams, J. M. (1988). Transgenic mice and oncogenesis. Ann. Rev. Immunol. 6, 25-48.
Showe, L. C. and Croce, C. M. (1987). The role of chromosomal translocations in B- and T-cell neoplasia. Ann. Rev. Immunol. 5, 253-277.



Genes VII
Genes VII
ISBN: B000R0CSVM
EAN: N/A
Year: 2005
Pages: 382

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