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Oncogenes and cell proliferation
Oncogenes
and cell proliferation
Editorial overview Chris Marshall* and Andrew
Wyllief
Addmsses * institute of Cancer research, Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, UK t Department of Pathology, University Medical School, Teviot Place, Edinburgh EHE QAG, UK Cunent
Opinion in Genetics and Development
1 QQ6,6, l-3
Q Current Biology Ltd ISSN 0959-437X Abbreviations interleukin-1 B-converting enzyme ICE HNPCC hereditary non-polyposis colon cancer
A programme for cell death The past few years have included remarkable advances in the field of cell death. Possibly the most significant single development has been the clarification of a basic biochemical pathway that appears to be common to death in a wide variety of circumstances. The existence of such a pathway was originally predicted (in 1972) leading to the introduction of the term apoptosis - on the basis of the distinctive structural changes that are common to mammalian and many invertebrate cells dying in the course of normal development, tissue homeostasis, function-dependent cellular selection (as in the negative selection of lymphocytes in the immune system), the response to certain cytotoxic agents, and in the targets of cytotoxic T-cell attack. It is now clear that such structural changes reflect the activation of a common effector pathway, driven by members of an unusual family of cysteine proteases. These are referred to collectively as ICE-family proteases, after the first identified family member, interleukin-lb converting enzyme, and are the subject of the article by Takahashi and Earnshaw (pp 50-55). It is now a matter of debate whether ICE itself is involved as an endogenous mediator of death, but the case is strong that this role is played by other family members, notably the protease designated CPP32, yama or apopain. Different ICE-family proteases mediate apoptosis in different cell lineages, or many may be involved together, in parallel or in tandem. The physiological activators and substrates of these proteases have not yet been satisfactorily defined, but these are autocatalytic enzymes, competent to participate in a self-amplifying cascade. They also cleave proteins with well-recognised roles in the cytoskeleton (e.g. fodrin), the nuclear envelope (lamin) and the response to DNA strand breaks (poly (ADP-ribose) polymerase), proteolysis of each of which might contribute to the structural changes characteristic of apoptosis. Significantly, at least one of the serine proteases
released from cytotoxic T-cell granules (granzyme B) is an activator of ICE-family proteases. Not surprisingly, activation of this effector pathway for suicide is under tight, complex control. One element in this is provided by a family of proteins of which the first discovered member was B&Z, an oncogenic protein whose dysregulation leads to prolonged survival of B-cells in certain human lymphomas. The review here (in the article by Farrow and Brown [pp 45-491) explores several new members of this family which appear to act in opposition to each other, competing in the formation of homodimers and heterodimers that either protect from, or potentiate, death depending on their composition. Differential production or stabilisation of these B&2 family proteins thus has the capacity to modify the sensitivity of the cell to lethal stimuli. There also appear to be proteins with similar roles outwith the B&2 family. Discovery of all these essential elements in the mammalian death pathway was foreshadowed by results of genetic analysis in invertebrates, notably Cacno&r~&r Agans, but more recently Dmsophila. These studies, which were commenced in the 197Os, did much to establish beyond doubt that specific genes are required to mediate and protect cells from developmentally programmed death. In the past five years, however, they have also revealed that many of the death-related genes in mammals are direct homologues of those found in invertebrates. Interesting conceptual problems arise from these striking homologies. Developmental cell death in C. e&am does not involve the selection of a small population of survivors from a much larger set doomed to die, as in the development of the mammalian nervous system or immune system. In all, there are only 131 cell death events in the lifetime of the organism, which possesses just over 1000 cells. Although these deaths are very precisely defined in space and time in nematode development, they are not used for extensive sculpting and moulding processes, and mutants that are constitutionally unable to activate death are not remarkably abnormal. Yet the death pathway in this simple invertebrate includes an ICE-family protease (ted 3) and a member of the B&2 family (ted 9). Invertebrate systems continue to provide new insights into the genetics of programmed death, and are discussed in the article by Hengartner (pp 34-38). , What then are the stimuli for death and how are they transduced? The first identified surface receptor whose activation induces apoptosis was a transmembrane receptor protein that appears on activated lymphocytes and several other cell types, called APO-l, fas or CD95. It is a member
2
Oncomma and cell pmlifwaUon
of the TNF receptor family, and its physiologicalligand (in the TNF family) has also been cloned. Advancescontinue to be made in the biology and downstream signalling mechanisms of both these receptors, and in this issue Tewari and Diit (pp 3e44) describe the TNF signalling pathway. Specific receptor-driven mechanisms such as CD95 and its ligand are not required for all apoptotic events, however, as mutant mouse strains deficient in each exist and are developmentally normal. There are also pathways that couple cell injury to the activation of apoptosis. The best known of these are sensitive to DNA breaks and activate apoptosis in a p53dependent manner, but there are certainly others. DNA-PK, a protein kinase complex activated through binding to DNA in the vicinity of a double strand break, may also contribute to signalling DNA damage. There is, at present, great uncertainty over why some cells proceed to apoptosis after DNA damage, whilst others successfully complete repair and survive. The nature of the cilluiar decisions to arrest proliferation, epter apoptosis or undertake repair is discussed in the reviews by Bates and Vousden(pp 12-18) and Jackson(pp 19-25). Further articles describe the use of powerful yeast systems to analyst this decision-making process (Lydall and Weinert [pp 4-101) and studies of the polyfunctional transcription complex TFIIH, which may lie at the heart of the decision-making process (Hoeijmakers, Egly and Vermeulen [pp 26-331).
Most studies to identify oncosuppressorgenes in human cancer, however, depend upon analyses of linkage with turnout susceptibility in cancer families, and positional cloning of common regions of allelic deletion on sporadic turnours. Both A!F,? (the gene responsible for the rarer type of familial neurofibromatosisreviewed by Gusella, Ramesh, MacCollin and Jacoby [pp 87-921) and the two breast cancer-susceptibilitygenesBRCAI and BRCAZ (reviewed by Stratton and Wooster [pp 93-971) were identified by these routes. Becauseno presuppositionsare made about the function of the oncosuppressorgene in this scenario, cloning of the gene represents only a first step towards the identification of the protein product and its biological roles, first in normal then in neoplastic cells. Whereas much of this still has to be explored in BRCAl and BRCA2, the NF2 protein is now known to be a member of the protein 4.1 family located at, or near, the cell membraneand is probably concernedwith connecting intra-membranous proteins to the cytoskeleton. Other proteins with these general characteristics have been shown to be significant in the organisationof cell shape, movement, attachment or communication.
A further approach, frequently adopted in conjunction with positional cloning based on loss of heterozygosity,is the testing of candidates that map to the deleted region. In view of the very widespread association of loss of p53 with tumour growth, transcriptional targets of p53 would appearto be good candidatesfor testing as potential oncosuppressorgenes.The review by Harper and Elledge (5664) summarisesrecent work on the family of cyclin New human kcee genes kinase inhibitors, of which one (pZZm/-‘) is known to be The articles in this section describerecent advancesin the transactivatedby wild-type ~53. The data show, however, detection of human cancer genes.The earliest method for that - of the cyclin kinase inhibitors tested - only detecting cancer genes depended upon transfer of tumour DNA into non-transformed cells in oitro and subsequent pl6 exerts a critical oncosuppressoractivity. Loss of function of the pl6 cyclin kinase inhibitor is evident in selection on the basis of dominant transforming activity. sporadic schwannomas,meningiomasand some malignant Most familial tumours showing a dominant pattern of mesotheliomas;as predicted for an oncosuppressorgene, inheritance of tumour susceptibility, however, do not arise through activation of such cytologically-dominant all the mutations result in truncated proteins which are oncogenes, but rather through loss of oncosuppressor presumably functionally inactive. genes, a phenomenon which cannot be detected by any simple DNA transfer method. It therefore came as some- A completely different mode of carcinogenesisis discussed thing of a surprise when the RET gene, responsible for in the article by Dunlop (pp 76-81). Defects in the transmission of the rare tumour-susceptibility syndrome genes responsible for DNA nucleotide m ismatch repair multiple endocrine neoplasia type 2, and some instances lead to a characteristic phenotype in which the incidence of medullary carcinoma of thyroid, turned out to be a of point mutations (usually by small frame shifts in transforming oncogene in the tyrosine kinase family, as m icrosatellite regions) are common. Patients inheriting reviewed here by Mak and Ponder (pp 82-86). such defective genes may be found as members of the familial syndromeof hereditary non-polyposiscolon cancer Another means of identifying dominant oncogenesis by (HNPCC). The full impact in human carcinogenesis searchingfor consistent translocationsin neoplasms.The of defects in m ismatch repair ,genes has not yet been translocation creates a dysregulated fusion gene which evaluated. Around l-5% of all colorectal cancers arise in can be identified by cloning the breakpoint region. The members of classic HNPCC kindreds, but between 10 review by Cooper (pp 71-75) summarisessome of the and 20% of all (apparently sporadic) colorectal cancers genetic loci at which such translocations occur in soft show the clonal appearanceof non-germline m icrosatellite tissue turnours. Surprisingly specific relationships emerge alleles, characteristicof m ismatchrepair deficiency. It is far between different translocationsand histologically-defined from clear what drives these cancers;this article explores turnout types. the interesting hypothesis that some of the patients may
EdItorId
be constitutionally mosaic for mutations in other colorectal cancer genes, as a result of failure of mismatch repair early in development. Finally, this section includes a critique (Williams and Jacks [pp 65-701) of the use of mouse strains bearing specific gene modifications for the analysisof mechanisms
ovotvlew
Marshall and Wyllii
3
of carcinogenicity.This has sometimes proved a definitive step in the dismissal or acceptance of a candidate cancer gene isolated initially on the basis of positional information. However, the technology now availableallows investigation of subtle interactions between different genes, and (in chimaeric animals) between cells of different genetic constitution.
and cell proliferation
Editorial overview Chris Marshall* and Andrew
Wyllief
Addmsses * institute of Cancer research, Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, UK t Department of Pathology, University Medical School, Teviot Place, Edinburgh EHE QAG, UK Cunent
Opinion in Genetics and Development
1 QQ6,6, l-3
Q Current Biology Ltd ISSN 0959-437X Abbreviations interleukin-1 B-converting enzyme ICE HNPCC hereditary non-polyposis colon cancer
A programme for cell death The past few years have included remarkable advances in the field of cell death. Possibly the most significant single development has been the clarification of a basic biochemical pathway that appears to be common to death in a wide variety of circumstances. The existence of such a pathway was originally predicted (in 1972) leading to the introduction of the term apoptosis - on the basis of the distinctive structural changes that are common to mammalian and many invertebrate cells dying in the course of normal development, tissue homeostasis, function-dependent cellular selection (as in the negative selection of lymphocytes in the immune system), the response to certain cytotoxic agents, and in the targets of cytotoxic T-cell attack. It is now clear that such structural changes reflect the activation of a common effector pathway, driven by members of an unusual family of cysteine proteases. These are referred to collectively as ICE-family proteases, after the first identified family member, interleukin-lb converting enzyme, and are the subject of the article by Takahashi and Earnshaw (pp 50-55). It is now a matter of debate whether ICE itself is involved as an endogenous mediator of death, but the case is strong that this role is played by other family members, notably the protease designated CPP32, yama or apopain. Different ICE-family proteases mediate apoptosis in different cell lineages, or many may be involved together, in parallel or in tandem. The physiological activators and substrates of these proteases have not yet been satisfactorily defined, but these are autocatalytic enzymes, competent to participate in a self-amplifying cascade. They also cleave proteins with well-recognised roles in the cytoskeleton (e.g. fodrin), the nuclear envelope (lamin) and the response to DNA strand breaks (poly (ADP-ribose) polymerase), proteolysis of each of which might contribute to the structural changes characteristic of apoptosis. Significantly, at least one of the serine proteases
released from cytotoxic T-cell granules (granzyme B) is an activator of ICE-family proteases. Not surprisingly, activation of this effector pathway for suicide is under tight, complex control. One element in this is provided by a family of proteins of which the first discovered member was B&Z, an oncogenic protein whose dysregulation leads to prolonged survival of B-cells in certain human lymphomas. The review here (in the article by Farrow and Brown [pp 45-491) explores several new members of this family which appear to act in opposition to each other, competing in the formation of homodimers and heterodimers that either protect from, or potentiate, death depending on their composition. Differential production or stabilisation of these B&2 family proteins thus has the capacity to modify the sensitivity of the cell to lethal stimuli. There also appear to be proteins with similar roles outwith the B&2 family. Discovery of all these essential elements in the mammalian death pathway was foreshadowed by results of genetic analysis in invertebrates, notably Cacno&r~&r Agans, but more recently Dmsophila. These studies, which were commenced in the 197Os, did much to establish beyond doubt that specific genes are required to mediate and protect cells from developmentally programmed death. In the past five years, however, they have also revealed that many of the death-related genes in mammals are direct homologues of those found in invertebrates. Interesting conceptual problems arise from these striking homologies. Developmental cell death in C. e&am does not involve the selection of a small population of survivors from a much larger set doomed to die, as in the development of the mammalian nervous system or immune system. In all, there are only 131 cell death events in the lifetime of the organism, which possesses just over 1000 cells. Although these deaths are very precisely defined in space and time in nematode development, they are not used for extensive sculpting and moulding processes, and mutants that are constitutionally unable to activate death are not remarkably abnormal. Yet the death pathway in this simple invertebrate includes an ICE-family protease (ted 3) and a member of the B&2 family (ted 9). Invertebrate systems continue to provide new insights into the genetics of programmed death, and are discussed in the article by Hengartner (pp 34-38). , What then are the stimuli for death and how are they transduced? The first identified surface receptor whose activation induces apoptosis was a transmembrane receptor protein that appears on activated lymphocytes and several other cell types, called APO-l, fas or CD95. It is a member
2
Oncomma and cell pmlifwaUon
of the TNF receptor family, and its physiologicalligand (in the TNF family) has also been cloned. Advancescontinue to be made in the biology and downstream signalling mechanisms of both these receptors, and in this issue Tewari and Diit (pp 3e44) describe the TNF signalling pathway. Specific receptor-driven mechanisms such as CD95 and its ligand are not required for all apoptotic events, however, as mutant mouse strains deficient in each exist and are developmentally normal. There are also pathways that couple cell injury to the activation of apoptosis. The best known of these are sensitive to DNA breaks and activate apoptosis in a p53dependent manner, but there are certainly others. DNA-PK, a protein kinase complex activated through binding to DNA in the vicinity of a double strand break, may also contribute to signalling DNA damage. There is, at present, great uncertainty over why some cells proceed to apoptosis after DNA damage, whilst others successfully complete repair and survive. The nature of the cilluiar decisions to arrest proliferation, epter apoptosis or undertake repair is discussed in the reviews by Bates and Vousden(pp 12-18) and Jackson(pp 19-25). Further articles describe the use of powerful yeast systems to analyst this decision-making process (Lydall and Weinert [pp 4-101) and studies of the polyfunctional transcription complex TFIIH, which may lie at the heart of the decision-making process (Hoeijmakers, Egly and Vermeulen [pp 26-331).
Most studies to identify oncosuppressorgenes in human cancer, however, depend upon analyses of linkage with turnout susceptibility in cancer families, and positional cloning of common regions of allelic deletion on sporadic turnours. Both A!F,? (the gene responsible for the rarer type of familial neurofibromatosisreviewed by Gusella, Ramesh, MacCollin and Jacoby [pp 87-921) and the two breast cancer-susceptibilitygenesBRCAI and BRCAZ (reviewed by Stratton and Wooster [pp 93-971) were identified by these routes. Becauseno presuppositionsare made about the function of the oncosuppressorgene in this scenario, cloning of the gene represents only a first step towards the identification of the protein product and its biological roles, first in normal then in neoplastic cells. Whereas much of this still has to be explored in BRCAl and BRCA2, the NF2 protein is now known to be a member of the protein 4.1 family located at, or near, the cell membraneand is probably concernedwith connecting intra-membranous proteins to the cytoskeleton. Other proteins with these general characteristics have been shown to be significant in the organisationof cell shape, movement, attachment or communication.
A further approach, frequently adopted in conjunction with positional cloning based on loss of heterozygosity,is the testing of candidates that map to the deleted region. In view of the very widespread association of loss of p53 with tumour growth, transcriptional targets of p53 would appearto be good candidatesfor testing as potential oncosuppressorgenes.The review by Harper and Elledge (5664) summarisesrecent work on the family of cyclin New human kcee genes kinase inhibitors, of which one (pZZm/-‘) is known to be The articles in this section describerecent advancesin the transactivatedby wild-type ~53. The data show, however, detection of human cancer genes.The earliest method for that - of the cyclin kinase inhibitors tested - only detecting cancer genes depended upon transfer of tumour DNA into non-transformed cells in oitro and subsequent pl6 exerts a critical oncosuppressoractivity. Loss of function of the pl6 cyclin kinase inhibitor is evident in selection on the basis of dominant transforming activity. sporadic schwannomas,meningiomasand some malignant Most familial tumours showing a dominant pattern of mesotheliomas;as predicted for an oncosuppressorgene, inheritance of tumour susceptibility, however, do not arise through activation of such cytologically-dominant all the mutations result in truncated proteins which are oncogenes, but rather through loss of oncosuppressor presumably functionally inactive. genes, a phenomenon which cannot be detected by any simple DNA transfer method. It therefore came as some- A completely different mode of carcinogenesisis discussed thing of a surprise when the RET gene, responsible for in the article by Dunlop (pp 76-81). Defects in the transmission of the rare tumour-susceptibility syndrome genes responsible for DNA nucleotide m ismatch repair multiple endocrine neoplasia type 2, and some instances lead to a characteristic phenotype in which the incidence of medullary carcinoma of thyroid, turned out to be a of point mutations (usually by small frame shifts in transforming oncogene in the tyrosine kinase family, as m icrosatellite regions) are common. Patients inheriting reviewed here by Mak and Ponder (pp 82-86). such defective genes may be found as members of the familial syndromeof hereditary non-polyposiscolon cancer Another means of identifying dominant oncogenesis by (HNPCC). The full impact in human carcinogenesis searchingfor consistent translocationsin neoplasms.The of defects in m ismatch repair ,genes has not yet been translocation creates a dysregulated fusion gene which evaluated. Around l-5% of all colorectal cancers arise in can be identified by cloning the breakpoint region. The members of classic HNPCC kindreds, but between 10 review by Cooper (pp 71-75) summarisessome of the and 20% of all (apparently sporadic) colorectal cancers genetic loci at which such translocations occur in soft show the clonal appearanceof non-germline m icrosatellite tissue turnours. Surprisingly specific relationships emerge alleles, characteristicof m ismatchrepair deficiency. It is far between different translocationsand histologically-defined from clear what drives these cancers;this article explores turnout types. the interesting hypothesis that some of the patients may
EdItorId
be constitutionally mosaic for mutations in other colorectal cancer genes, as a result of failure of mismatch repair early in development. Finally, this section includes a critique (Williams and Jacks [pp 65-701) of the use of mouse strains bearing specific gene modifications for the analysisof mechanisms
ovotvlew
Marshall and Wyllii
3
of carcinogenicity.This has sometimes proved a definitive step in the dismissal or acceptance of a candidate cancer gene isolated initially on the basis of positional information. However, the technology now availableallows investigation of subtle interactions between different genes, and (in chimaeric animals) between cells of different genetic constitution.