Id Gene Expression As a Key Mediator of Tumor Cell Biology I

Home , Extramacrochaetae, ID2, ID3 (gene), Mediator (coactivator)

[CANCER RESEARCH (SUPPL.) 59, 1726s-1730s, April 1, 19991 Id Gene Expression as a Key Mediator of Tumor Cell Biology I

Mark A. Israel, z Maria-Clemencia Hernandez, Monica Florio, Pedro J. Andres-Barquin, Akio Mantani, John H. Carter, and Carol M. Julin Preuss Laboratory for Molecular Neuro-Oncology, Brain Tumor Research Center, Department of Neurological Surgery, School of Medicine, University of California San Francisco, San Francisco, California 94143

Abstract mechanisms by which normal cells mediate the expression of differentiated characteristics and how the expression of these characteristics Id genes encode members of the helix-loop-helix (HLH) family of is inhibited in tumor cells may provide important insights into the transcription factors that inhibit transcription by forming inactive het- pathological processes by which the malignant characteristics of can- erodimers with basic HLH (bHLH) proteins. There are four members of the Id gene family recognized in mammals, and the proteins they encode cer cells are regulated. share homology primarily in their HLH domain, bHLH proteins typically form heterodimers with other bHLH proteins, and their basic domain Id Genes binds to a DNA sequence element, the E-box, activating transcription. Products of Id genes lack the basic DNA binding domain of the bHLH Id proteins belong to a class of transcription factors known as HLH 3 transcription factors, and when they heterodimerize with bHLH proteins, proteins (1, 2). The distinguishing feature of this class is an amino the complexes are inactive. Generally, high levels of Id mRNA are de- acid motif predicted to form a HLH structure mediating the dimer- tected in proliferative undifferentiated, embryonal cells and lower levels ization of these proteins. Many members of the HLH family acting as are detected in well-differentiated, mature, adult tissues. In vitro, these transcription factors contain a domain of basic amino acids NH 2- genes are generally expressed at lower levels in cells after the induction of terminal to the HLH motif. The basic domain of such transcription differentiation. Recently, high levels of expression of Id genes have been factors (bHLH factors) mediates their binding to DNA when these identified in cell lines derived from a wide variety of different tumors and in tumor tissues as well. These findings suggest that not only the inappro- proteins dimerize (3). These proteins are structurally and biologically priate proliferation of tumors but also the anaplastic characteristics that distinct from other, related transcription factors such as bHLH-ZIP contribute to their malignant behavior may be regulated by ld gene factors, which, in addition to the HLH dimerization motif, also contain expression. a leucine zipper (1, 4). bHLH factors have been designated as class I and class II proteins based on their ubiquitous expression in many Introduction different cell types (class I) or a highly restricted pattern of expression An important challenge for cancer research is to understand how limited to cells of a single lineage (class II; Refs. 3 and 4). bHLH genetic alterations associated with tumorigenesis actually contribute transcription factors are known to play an important role in the to the complex cellular and biological alterations that can be recog- regulation of cell determination and differentiation in many lineages, nized as malignancy. Genes intimately involved in the regulation of and almost 70 different genes encoding lineage-specific bHLH tran- cellular proliferation are known to be altered in virtually all of the scription factors have been identified. commonly occurring tumors, including embryonal tumors. These Id proteins are distinct from bHLH factors in that they lack the genes typically belong to the family of cancer-related genes known as basic amino acid domain necessary for DNA binding. Id proteins tumor suppressor genes. However, the precise role of oncogenes, inhibit bHLH proteins from binding to DNA and inactivate transcrip- genes whose normal functions are enhanced in many different tumors, tion by heterodimerization with bHLH proteins (Refs. 1 and 4; Fig. 1; in mediating the key characteristics of malignancy is less clear. Tissue Table 1). This inhibition of DNA binding, in turn, is closely associated invasion, metastases, neoangiogenesis, immune escape, and therapeu- in vitro with the inhibition of induced differentiation in a number of tic resistance, all key characteristics of malignancy, seem to occur as different cell types in which tissue-specific differentiation has been a result of cellular transformation, but how this occurs is poorly studied, including neuronal cells (11), skeletal muscle (12), mammary understood. cells (13), adipocytes (14), and others (Table 2). Because Id proteins Virtually all cancer is characterized by two invariant characteristics, seem to bind class I bHLH proteins more avidly than those expressed inappropriate growth and a lack of the differentiated features that in a lineage-specific manner (4), the mechanism by which Id proteins characterize the tissue in which any given tumor arises. Enhanced inhibit cellular differentiation may be through the sequestration of proliferation and anaplasia are the cornerstones for diagnostic recog- class I bHLH proteins (Fig. 1). nition of malignant tissue. In tumors, this is frequently identified by Four Id genes, originally termed Id (Idl) (1), Id2 (4), HLH462 (Id3) the appearance of mitotic figures in tumor cells that are oftentimes so (20), and Id4 (21), have been cloned from the mouse. Human homo- undifferentiated that they cannot be recognized as one specific tissue logues of each of these have also been characterized (22-25). The four type or another. Interestingly, many of the biological features of Id genes map to different chromosomes in both man, in whom all four malignant cells, such as the ability to invade neighboring tissues, have been mapped, and mouse, in which Idl and Id2 have been migration, and their lack of recognition by the immune system, are mapped to the proximal region of chromosomes 2 and 12, respectively. also characteristics of embryonal cells. Understanding the molecular In man, the Idl, Id2, Id3, and Id4 genes map to chromosomes 20ql 1 (26), 2p25 (26), lp36 (23), and 6p21.3-22 (25), respectively. Despite

Received 9/18/98; accepted 2/1/99. their unlinked chromosomal locations, these genes share considerable 1 Presented at the "General Motors Cancer Research Foundation Twentieth Annual homology to one another and are likely to have been derived from a Scientific Conference: Developmental Biology and Cancer," June 9-10, 1998, Bethesda, common ancestral gene (2). The region of greatest conservation is MD. This work was supported in part by grants from the Pediatric Brain Tumor Foun- dation of the United States, the Preuss Foundation, the Betz Foundation, and the Nissen clearly in the HLH domain, which extends for approximately 40 family. amino acids and is located in the middle region of the Id molecule. 2 To whom requests for reprints should be addressed, at Preuss Laboratory for Molecular Neuro-Oncology, Brain Tumor Research Center, Department of Neurological Surgery, HSE 722, University of California San Francisco, 513 Parnassus Avenue, San 3 The abbreviations used are: HLH, helix-loop-helix; bHLH, basic HLH; pRb, retino- Francisco, CA 94143-0520. blastoma gene product. 1726s

TSF E47

ilIH4 H 4 IilII I LINEAGE-SPECIFIC GENE I >

[ LINEAGE-SPECIFIC GENE [ ~' IIIIH4 H IIIII -!-

TSF Id E47 J

Fig. 1. Id proteins function to inhibit tissue-specific gene expression by binding to bHLH transcription factors, bHLH transcription factors such as E47 are ubiquitously expressed, whereas other tissue-specific bHLH transcription factors (TSF) have a very limited distribution of expression and typically regulate the expression of other tissue-specific genes. Id proteins lack a basic DNA binding domain and, after dimerization,inhibit the binding of active, heterodimericcomplexes of bHLH factors to E-boxes (CANNTG) found in the upstream regulatory regions of many different genes. + and - indicate the likelihood of transcriptional activation when HLH-containing transcription factors are present in the molecular conformations indicated.

There is an approximately 85% amino acid identity in this region studies have been remarkable in demonstrating widespread expression among the four different Id genes, and several other very short regions of the ldl, Id2, and Id3 genes throughout the developing organisms of sequence similarity have been recognized among various members from very early times in gestation through birth. Some tissues clearly of this gene family. Interestingly, alternative open reading frames express multiple different Id genes, and data from cell lines in culture have been identified for both Idl (27) and Id3 (28), and a mRNA from indicate that a single, specific cell type can express multiple, different an Id2 pseudogene that could encode a peptide corresponding to the Id genes simultaneously. In sharp contrast to the other Id genes, the first 36 amino acids of Id2 has been identified (29). The functions of expression of Id4 seems to be limited largely to the peripheral and the distinctive NH2-terminal and COOH-terminal regions of the dif- central nervous systems (34). Also, at the level of resolution afforded ferent Id proteins are largely enigmatic. The COOH-terminal portion by published in situ hybridization studies, it appears that there is of Id3 is clearly important for the stabilization of this protein (30), considerable overlap in the pattern of expression of Idl and Id3, which although little is known of the mechanisms that regulate Id protein is distinguishable from the pattern of expression of Id2 throughout the levels in any cell type. Importantly, a role for the NH2-terminal region organism and from the patterns of Id2 and Id4 expression in the of Id2 in mediating the apoptotic response of cells to growth factor nervous system. Remarkably, the pattern of Id2 and Id4 expression in withdrawal has recently been identified, suggesting that multiple the nervous system is quite similar. These findings suggest that there different domains of these proteins will be of functional significance may be some functional redundancies of these different ld genes, a (see below). finding consistent with the extensive sequence similarity in the HLH Id Gene Expression regions of the Id proteins. Although it is clear that mRNA corresponding to several different Investigators using in situ hybridization have sought to examine the Id genes is expressed in some adult tissues, the precise cell types in expression of Id genes during mouse development (31-35). These which these genes are expressed and the relative levels of expression of these genes in mature tissues are unclear. It is generally thought that Table 1 Effect of Id gene expression on lineage-specific genes the highest levels of Id gene expression occur in undifferentiated Altered gene expressiona Species Tissue type Ref. tissues, whereas lower levels of expression occur in their differenti- Osteocalcin Rat Bone 5 ated counterparts. It is not known whether terminally differentiated Insulin Rat Pancreas 6 Cardiac c~-actin Human Heart 7 tissues still express Id proteins, although it seems likely that Id2 is lmmunoglobulin genes Mouse Lymphoid 8 expressed in some postmitotic neurons of the central nervous system Nerve growth factor receptor (p75rqcFR) Rat Nerve 9 a-Glycoprotein homaone Human Pituitary 10 (33). One possible interpretation of studies detailing the widespread c, The expression of each indicated gene is inhibited by Idl expression, except for the high-level expression of Id genes during early development is that repression of p75 N~FR, which is antagonized by Id2. these genes function to modulate the maturation of cells during 1727s

Table 2 Representative in vitro lineage-specific differentiation models associated with In mice, the constitutive expression of an Idl transgene under the changes in ld gene expression control of the immunoglobulin promoter results in inhibited B-cell Tissue type Cell line Ref. maturation (42). Skeletal muscle C2C12 12 A second well-characterized function of Id genes is enhancing cell Hematopoietic cells 32DC13 15 Bone MC3T3-E1 16 cycle progression. The hypothesis that Id proteins could have a role in Mammary tissues Scp2 13 cell proliferation and cell cycle regulation was initially proposed for Trophoblasts Rcho- 1 17 Id3, which was first described as HLH462 (20). This gene is induced Monocytes HL60 18 Adipose tissue 3T3-F442A 14 in mouse 3T3 cells as part of the immediate early transcription Sertoli cells Primary Sertoli cells 19 response to growth factors (20). Subsequently, the induction of other Id genes by serum and isolated growth factors was described, and the ability of Idl, Id2, or Id3 expression to enhance the progression of differentiation, maintaining them in a state that allows them to express cells from G~ to S phase was characterized. Antisense oligonucleo- a limited degree of differentiation without becoming terminally dif- tides that inhibit Id gene expression delay the reentry of growth- ferentiated and growth-arrested (35). While preventing differentiation, arrested 3T3 cells into the cell cycle elicited by stimulation with expression of Id proteins may also be responsible for keeping the cell serum (36). Interestingly, combined oligonucleotides against Idl, Id2, in a highly proliferative state (36-39). This is clearly true in vitro, and Id3 mRNAs were more effective than individual antisense oligo- where the induced differentiation of cell lines derived from several nucleotides in these experiments, suggesting that the activities of these different lineages is associated with varying degrees of decreased Id Id genes in enhancing cell cycle progression are not completely gene expression over the course of maturation and growth inhibition. redundant (36). Other data, however, suggest that the precise molecular events medi- The potential of Id genes to enhance cell cycle progression through ated by Id genes during development may be more complicated distinct molecular mechanisms is in accord with the finding that Id2, because it is clear that Id2 increases in association with myeloid but not Idl or Id3, can reverse the inhibition of cellular proliferation differentiation (18). In vitro, increased levels of Id gene expression mediated by pRb (37, 39). In vivo, a weak physical association commonly precede the decrease that typically accompanies induced between Id2 and pRb has been demonstrated, and in vitro unphos- differentiation. phorylated pRb but not the hyperphosphorylated form binds Id2. The HLH region of Id2 is important for binding to pRb, and wild-type pRb Id Gene Function is able to bind to a region of Id2 corresponding to only the HLH Roles for Id gene function in several different biological events domain. Id2 can also bind the pRb-related proteins p107 and p130 and important for both normal development and oncogenesis have been reverse their inhibition of cell cycle progression (39). In agreement identified (Fig. 2). The best characterized function of Id genes is to with these results, neither Idl nor Id3 affects the arrest of cell cycle inhibit lineage-specific gene expression. To date, virtually all data progression mediated by pRb. The growth-suppressive activity of supporting this view come from in vitro experiments in which the cyclin-dependent kinase inhibitors pl6 and p21 is also efficiently introduction of Id genes into various cell lines modulates lineage- antagonized by high levels of Id2 but not by Idl or Id3 (39). These specific gene expression (Table 1) and inhibits the induced differen- data provide evidence for Id2-mediated effects on physiological, tiation of cells as measured by either morphological examination or regulatory events important for cell cycle progression, and they iden- biochemical analysis (Table 2). Among the model systems that have tify distinctive roles for different Id gene products. This interpretation been examined are those mimicking differentiation of a considerable is consistent with recent work indicating that Idl overexpression in number of different tissues including those listed in Table 2. NIH 3T3 cells accelerates cell growth and inhibits p21 expression by Data from emc, a Drosophila homologue of Id, provided the first in antagonizing bHLH transcription factor-mediated p21 gene expres- vivo evidence for the inhibition of differentiation attributed to Id genes sion (43). Interestingly, the function of some Id genes to mediate cell (40, 41). The study of emc and a variety of its mutants indicates that cycle progression may be influenced by cyclin-dependent kinase- HLH proteins lacking a basic DNA binding region, such as emc and mediated phosphorylation (44). Id, can inhibit differentiation of the tissue in which they are expressed. A role for Id genes in mediating apoptosis has also been described

Nucleus i Cytoplasm

+,-oeoe N __ expression Differentiation

Proliferation

D-, Apoptosis

expression

Fig. 2. Id genes contribute to the regulation of cellular differentiation,proliferation, and apoptosis. Id proteins may require interaction with a bHLH protein before they can be transported to the nucleus, where they inhibit the binding of bHLH factors to DNA in upstream regulator regions. Similarly, bHLH proteins and pRb can inhibit Id activity. Because bHLH proteins can function as activators or repressors of gene expression, ld gene expression can function to either enhance or inhibit gene expression. Recent data indicatingthat the NHz-terminalregion (N) but not the HLH region of Id2 is required to enhance apoptosis in myeloid precursors suggest that Id effects may also be mediatedthrough interactions with other classes of proteins as well. 1728s

Downloaded from cancerres.aacrjournals.org on September 26, 2021. © 1999 American Association for Cancer Research. /d GENES IN CANCER previously (45). Several different genes important for cell cycle pro- development of lymphoma in vivo (42). Given these observations and gression, for example c-Myc and E2F1, have been recognized to the activities of Id genes to induce anaplasia and increase cellular induce apoptosis when ectopically expressed in cells deprived of proliferation, these genes are strong candidates for mediating onco- mitogenic factors (46, 47). These findings have been interpreted as genesis and the accompanying embryonal features of malignancy. supporting a model in which activation of the cell death pathway is due to a conflict between growth-promoting and growth-inhibitory Acknowledgments signals (48, 49). Transient transfection of Idl, Id2, or ld3 in rat We thank Lucy Avila for editorial assistance. We thank the Preuss Foun- embryo fibroblasts enhances the transition of cells into S phase, and dation and the Pediatric Brain Tumor Foundation of the United States for this same population of cells could be identified as containing cells support. We regret that we could not include all of the relevant citations due with an apoptotic morphology (45). The conditional expression of Id3 to space limitations. in stable transfectants of the same cell type also induces apoptosis in serum-deprived cells. In both cases, apoptosis was suppressed in the References presence of serum. Studying 32D.3 murine myeloid progenitors, our 1. Benezra, R., Davis, R. L., Lockshon, D., Turner, D. L., and Weintraub, H. The protein laboratory has found that Idl and Id2, but not Id3, can enhance ld: a negative regulator of helix-loop-helix DNA binding proteins. Cell, 61: 49-59, apoptosis that occurs after withdrawal of the survival factor interleu- 1990. kin 3. This activity of Id2 does not require the HLH region, although 2. Norton, J. D., Deed, R. W., Craggs, G., and Sablitzsky, F. Id helix-loop-helix proteins in cell growth and differentiation. Trends Cell. Biol., 8: 58-65, 1998. the NH2-terminal region is necessary (49). 3. Murre, C., Schonleber McCaw, P., Vaessin, H., Candy, M., Jan, L. Y., Jan, Y. N., Cabrera, C. V., Buskin, J. N., Hauschka, S. D., Lassar, A. B., Weintraub, H., and Id Gene Expression and Activity in Human Cancer Baltimore, D. interactions between heterologous helix-loop-helix proteins generate complexes that bind specifically to a common DNA sequence. Cell, 58: 537-544, 1989. Expression of mRNA encoded by the Idl, Id2, ld3, and Id4 genes 4. Sun, X-H., Copeland, N. G., Jenkins, N. A., and Baltimore, D. Id proteins Idl and Id2 has been examined in a variety of human tumor cell lines, including selectively inhibit DNA binding by one class of helix-toop-he|ix proteins. Mol. Cell. those from the lung, colon, and pancreas, and in both neuronal and Biol., 11:5603-5611, 1991. 5. Tamura, M., and Noda, M. identification of a DNA sequence involved in osteoblast- astrocytic tumors of the nervous system (23, 24, 33, 50, 51). In each specific gene expression via interaction with helix-loop-helix (HLH)-type transcrip- of these cell lines, one or more Id genes were found to be expressed. tion factors. J. Cell Biol., 126: 773-782, 1994. Interestingly, in nervous system cell lines in which Id4 is highly 6. Cordle, S. R., Henderson, E., Masuoka, H., Well, P. A., and Stein, R. Pancreatic /3-cell-type-specific transcription of the insulin gene is mediated by basic helix-loop- expressed, Id2 is also frequently expressed. Only a limited number of helix DNA-binding proteins. Mol. Cell. Biol., 11: 1734-1738, 1991. embryonal tumors have been examined to date, and these include only 7. Sartorelli, V., Hong, N. A., Bishopric, N. H., and Kedes, L. Myocardial activation of the human cardiac c~-actin promoter by helix-loop-helix proteins. Proc. Natl. Acad. nervous system tumors such as neuroblastoma and medulloblastoma. Sci. USA, 89: 4047-4051, 1992. Neuroblastoma-derived cell lines tend to express generally high levels 8. Kadesch, T. Consequences of heteromeric interactions among helix-loop-helix pro- of multiple Id genes, whereas very low levels of Id expression were teins. Cell Growth Differ., 4: 49-55, 1993. 9. Chiaramello, A., Neuman, K., Palm, K., Metsis, M., and Neuman, T. Helix-loop-helix observed in the cells derived from medulloblastomas, hnportantly, all transcription factors mediate activation and repression of the p75 LNc~FR gene. Mol. of these experiments have examined RNA from cells grown in the Cell. Biol., 15: 6036-6044, 1995. presence of serum, a known inducer of Id gene expression. In any 10. Jackson, S. M., Gutierrez-Hartmann, A., and Hoeffler, J. P. Upstream stimulatory factor, a basic-loop-helix-zipper protein, regulates the activity of the c~-glycoprotein case, it seems likely that in cell lines derived from most tumor types, hormone subunit gene in pituitary cells. Mol. Endocrinol., 9: 278-291, 1995. one or more ld genes will be highly expressed, although there may be 11. Lee, J. E. Basic helix-loop-helix genes in neural development. Curr. Opin. Neurobiol., 7: 13-20, 1997. occasional exceptions to that rule (33). 12. Jen, Y., Weintraub, H., and Benezra, R. Overexpression of ld protein inhibits the Experiments examining the morphological characteristics of muscle differentiation program: in vivo association of Id with E2A proteins. Genes NIH3T3 cells stably transfected with Id3 indicated that Id3 expression Dev., 6: 1466-1479, 1992. 13. Desprez, P. Y., Hara, E., Bissell, M. J., and Campisi, J. Suppression of mammary can induce a transformed morphology in these cells, although the cells epithelial cell differentiation by the helix-loop-helix protein ld-1. Mol. Cell. Biol., 15: did not acquire other characteristics of malignant transformation and 3398-3404, 1995. did not undergo apoptosis (52). This finding is consistent with the 14. Moldes, M., Lasnier, F., Feve, B., Pairault, J., and Djian, P. ld3 prevents differentiation of preadipose cells. Mol. Cell. Biol., 17:1796-1804, 1997. observation that primary rat embryo cells can be immortalized by 15. Kreider, B. L., Benezra, R., Rovera, G., and Kadesch, T. Inhibition of myeloid transfection of Id3 and either Bcl2 or BclX, but not by transfection of differentiation by the helix-loop-helix protein ld. Science (Washington DC), 255: 1700-1702, 1992. Id3 alone; it has been possible to induce stable foci formation of 16. Mun-ay, S. S., Glackin, C. A., Winters, K. A., Gazit, D., Kahn, A. I., and Murray, E. J. primary rat embryo cells with ld3 (45), and in unpublished studies, Expression of helix-loop-helix regulatory genes during differentiation of mouse Idl has been shown to immortalize primary human keratinocytes. 4 Id osteoblastic cells. J. Bone Miner. Res., 7:1131-1138, 1992. 17. Cross, J. C., Flannery, M. L., Blanar, M. A., Steingrimsson, E., Jenkins, N. A., genes are strong candidates to contribute to the malignant transfor- Copeland, N. G., Rutter, W. J., and Werb, Z. Hxt encodes a basic helix-loop-helix mation of some cell types and to regulate the expression of the transcription factor that regulates trophoblast cell development. Development malignant characteristics of human cancer. (Camb.), 121: 2513-2523, 1995. 18. lshiguro, A., Spirin, K. S., Shiohara, M., Tobler, A., Gombart, A. F., Israel, M. A., Norton, J. D., and Koeffler, H. P. Id2 expression increases with differentiation of Conclusion human myeloid cells. Blood, 87: 5225-5231, 1996. 19. Chaudhary, J., Cupp, A. S., and Skinner, M. K. Role of basic-helix-loop-helix Generally, high levels of Id mRNA are detected in proliferative, transcription factors in Sertoli cell differentiation: identification of an E-box response element in the transferrin promoter. Endocrinology, 138: 667-675, 1997. undifferentiated embryonal cells, and these genes are expressed at 20. Christy, B. A., Sanders, L. K., Lau, L. F., Copeland, N. G., Jenkins, N. A., and lower levels in cells induced to differentiate or in cells found in the Nathans, D. An Id-related helix-loop-helix protein encoded by a growth factor- mature tissues of adults. Recently, we have found high levels of ld inducible gene. Proc. Natl. Acad. Sci. USA, 88: 1815-1819, 1991. 21. Riechmann, V., van Criichten, I., and Sablitzky, F. The expression pattern of ld4, a gene expression in tumor cell lines derived from many different novel dominant negative helix-loop-helix protein, is distinct from Idl, Id2 and Id3. tissues (23, 24, 33, 50, 51). Overexpression of ld3 induces a morpho- Nucleic Acids Res., 22: 749-755, 1994. logically transformed phenotype in NIH3T3 cells, and Idl and Id3 can 22. Deed, R. W., Jasiok, M., and Norton, J. D. Nucleotide sequence of the cDNA encoding human helix-loop-helix Id-1 protein: identification of functionally con- contribute to the immortalization of primary cells (52). Also, the served residues common to ld proteins. Biochim. Biophys. Acta, 1219: 160-162, constitutive expression of Idl in developing B cells contributes to the 1994. 23. Biggs, J., Murphy, E. V., and Israel, M. A. A human Id-like helix-loop-helix protein expressed during early development. Proc. Natl. Acad. Sci. USA, 89: 1512-1516, 4 R. M. Alani, K. Munger, and M. A. Israel, unpublished data. 1992. 1729s

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M., and O'Brien, T. X. Expression of the helix-loop-helix factor Id during and accelerates apoptosis. Oncogene, 6." 1915-1922, 1991. mouse embryonic development. Dev. Biol., 159: 485-499, 1993. 47. Evan, G. I., Wytlie, A. H., Gilbert, C. S., Littlewood, T. D., Land, H., Brooks, M., 33. Zhu, W., Dahmen, J., Bulfone, A., Rigolet, M., Hernandez, M-C., Kuo, W-L., Puelles, Waters, C. M., Penn, L. Z., and Hancock, D. C. Induction of apoptosis in fibroblasts L., Rubenstein, J. L. R., and Israel, M. A. ld gene expression during development and by c-myc protein. Cell, 69: 119-128, 1992. molecular cloning of the human Id-1 gene. Mol. Brain Res., 30: 312-326, 1995. 48. Wu, X. W., and Levine, A. J. P53 and E2F-1 cooperate to mediate apoptosis. Proc. 34. Riectunann, V., mad Sablitzky, F. Mutually exclusive expression of two dominant- Natl. Acad. Sci. USA, 91: 3602-3606, 1994. negative helix-loop-helix (dnHLH) genes, Id4 and Id3, in the developing brain of the 49. Florio, M., Hernandez, M-C., Yang, H., Shu, H-K., Cleveland, J. L., and Israel, M. A. mouse suggests distinct regulatory roles of these dnHLH proteins during cellular prolif- Id2 promotes apoptosis by a novel mechanism independent of dimerization to basic eration and differentiation of the nervous system. Cell Growth Differ., 6: 837-843, 1995. helix-loop-helix factors. Mol. Cell. Biol., in press, 1999. 35. Jen, Y., Manova, K., and Benezra, R. Each member of the Id gene family exhibits a 50. Andres-Barquin, P. J., Hernandez, M-C., Hayes, T. E., McKay, R. D. G., and Israel, unique expression pattern in mouse gastrulation and neurogenesis. Dev. Dyn., 208: M. A. Id genes encoding inhibitors of transcription are expressed during in vitro 92-106, t997. astrocyte differentiation and in cell lines derived from astrocytic tumors. Cancer Res., 36. Barone, M. V., Pepperkok, R., Peverali, F. A., and Philipson, L. ld proteins control growth 57: 215-220, 1997. induction in mammalian cells. Proc. 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Discussion

Speaker: If Id2 is so important for tumorigenesis, wouldn't you Rb, 1 do you see a difference in the level of expression of Id genes in expect to see it occasionally genetically altered in translocation and Rb- tumors versus Rb+ tumors? overexpressed by amplification? That would be the real convincing Dr. Israel: That's a fair question. We haven't done that, so I evidence, wouldn't it? don't know the answer, but I would say that I personally think the primary function of the Id genes is to contribute to anaplasia, Dr. Israel: Well, that would be convincing evidence that Id2 was an which is invariably a characteristic of human cancer. If you ask a oncogene, but it's not clear to me that it is an oncogene. The other 10 pathologist how he recognizes that a tissue is malignant, he doesn't slides in my carousel described how overexpression of Id2 can also say that it's growing inappropriately. He will say that there is cause apoptosis, as do many genes that enhance proliferation. Our evidence of proliferation, an increased number of mitotic figures, feeling is that there is a sort of a finely tuned mechanism through and so forth and that it is anaplastic. So, my own feeling is that this which different ld genes in different environments can contribute to could be the mechanism by which Id contributes to the anaplastic malignant transformation. However, I would say that whereas it hasn't features of cancers and, as part of that, contributes to the malignant yet turned up and hasn't been found in translocations, I would be characteristics of tumors. reluctant at this point to say that there are no gain of function Dr. Sharon Murphy: Thank you. It's wonderful to see how the study mutations that exist. Data from other laboratories do suggest that Id of pediatric cancer provides models for understanding all forms of cancer. genes can contribute to transformation. Dr. Phillip Sharp: If the Id genes are working primarily through The abbreviation used is: Rb, retinoblastoma protein.

1730s

Mark A. Israel, Maria-Clemencia Hernandez, Monica Florio, et al.

Cancer Res 1999;59:1726s-1730s.

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