Recent Progress in Hormone Research 58:325-342 (2003)
© 2003 The Endocrine Society
Microarray Analysis and Identification of Novel Molecules Involved in Insulin-like Growth Factor-1 Receptor Signaling and Gene Expression
Joelle Dupont*,
Sandra E. Dunn
,
J. Carl Barrett
and
Derek LeRoith¶
* Physiologie de la Reproduction et des Comportements, Unite Mixte de Recherches 6073, Institut National de la Recherche Agronomique-Centre National de la Recherche Scientifique, Université F. Rabelais, Tours 37380 Nouzilly, France
British Columbia Research Institute for Children’s and Women’s Health, Vancouver, British Columbia, Canada
Center for Cancer Research, National Cancer Institute, Bethesda, Maryland 20892-2440
¶ Clinical Endocrinology Branch, National Institutes of Health, Bethesda, Maryland 20892-1758
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ABSTRACT
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The insulin receptor (IR) and the insulin-like growth factor-1 receptor (IGF-1R) are members of the same subfamily of receptor tyrosine kinases. The two receptors phosphorylate many of the same substrates and activate the same signaling modules, including the mitogen-activated protein kinase (MAPK) and phosphatidyl inositol 3' kinase (PI3K) signaling pathways. Although the IR and IGF-1R share some redundant functions in metabolism, cell growth, differentiation, and apoptosis, they also exhibit distinct physiological roles. Some of these may be due to differences in tissue distribution, receptor structure, formation of hybrid receptors, or mechanisms of ligand binding. However, the divergent effects of insulin and IGF-1 also may be explained by specificity in the intracellular signals generated by insulin and IGF-1. In particular, the IR and IGF-1R are capable of triggering their own biological responses by using specific or preferential substrates, molecular adapters, or signaling pathways. In a recent study, we used cDNA microarray analysis to identify genes differentially regulated by insulin and IGF-1. Mouse NIH-3T3 fibroblasts expressing either the wild-type human IGF-1R or IR were stimulated with either IGF-1 or insulin, respectively. We identified 39 genes differentially regulated by insulin and IGF-1. Most of these genes had not been reported previously to be responsive to insulin or IGF-1. The genes induced by IGF-1 generally were involved in mitogenesis or differentiation, while the genes found to be induced by insulin did not conform to any particular category. In a separate study, immortalized breast epithelial cells were stimulated with IGF-1 and a cDNA microarray analysis was used to generate a profile of IGF-1-regulated genes. A number of genes known to be involved in angiogenesis were found to be regulated by IGF-1. These results strongly suggest that this technology may be extremely useful in identifying groups of genes that are specifically regulated by different ligands and their activated receptors.
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I. Introduction
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Insulin and insulin-like growth factor-1 (IGF-1) are peptide hormones that are homologous in primary structure but differ in their physiological effects. Insulin, produced by the beta (ß) cells of the pancreas, stimulates the uptake of glucose and amino acids, inhibits gluconeogenesis, and promotes lipogenesis. IGF-1 is involved primarily in cell growth, survival, apoptosis, and differentiation. Insulin and IGF-1 mediate their biological effects by binding to their respective receptors, the insulin receptor (IR) and the IGF-1 receptor (IGF-1R). Although IR and IGF-1R are structurally and functionally similar, targeted gene knockouts in mice showed that they have both overlapping and distinct physiological roles (Nakae et al., 2001). One of the major areas of interest in this field is to understand how the specificity of IR and IGF-1R signaling is defined. In this review, we briefly compare and contrast the IR and IGF-1R signaling pathways and discuss various mechanisms that could explain the divergent physiological functions mediated by the two receptors. We then describe recent experiments using cDNA microarray analysis that have identified specific differences at the level of gene expression.
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II. Structure of the Insulin and IGF-1 Receptors
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The IR and IGF-1R are both comprised of two extracellular alpha (
) subunits containing ligand-binding sites and two transmembrane ß subunits transmitting the ligand-induced signal (Yarden and Ullrich, 1988). More specifically, IGF-1R and IR ß subunits consist of three domains: 1) a juxtamembrane domain, with motifs required for recruiting the major signaling adapter proteins; 2) a tyrosine kinase domain, essential for catalytic activity of the receptor; and 3) the carboxyl-terminal domain, which has several important residues for IGF-1R and IR signaling (Figure 1). As a consequence of this high level of homology, hybrid receptors, comprised of an insulin ß-hemireceptor and an IGF-1 ß-hemireceptor, can form in tissues and cultured cells expressing both the IR and the IGF-1R (Federici et al., 1997). Such hybrid receptors may play a role in the divergent actions of insulin and IGF-1.
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FIG. 1. The insulin-like growth factor (IGF) family of ligands, binding proteins (IGFBPs), and receptors (IGF-1R). The IGFs are bound by binding proteins in the circulation and in the extracellular matrix of the target cells. One mechanism for their release is specific protease effects on the IGFBPs, which release the IGFs to interact with their specific cell-surface receptors. IGFs may interact with IGF-1, insulin, and hybrid receptors to affect cell signaling within the target cell.
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A. EXTRACELLULAR (LIGAND-BINDING) DOMAIN
Despite the structural similarities between IGF-1 and insulin, the IR and IGF-1R have 100- to 1000-fold higher binding affinity for their cognate ligands. The subunits have been shown to confer ligand-binding specificity (Schumacher et al., 1991). Some studies using chimeric receptors have shown that the high affinity of the IR for insulin is determined by regions adjacent to the cysteine-rich domain (Gustafson and Rutter, 1990; Schumacher et al., 1991). On the other hand, the high-affinity IGF-1 binding by the IGF-1R is determined by its cysteine-rich domain within the subunit.
B. CYTOPLASMIC DOMAIN
Structural differences in the cytoplasmic domain of the ß subunits of the IR and IGF-1R may contribute to the divergence of these two signaling pathways. The highest degree of homology between the two receptors is found within the tyrosine kinase domain (about 84%), whereas the region of greatest divergence between the IR and IGF-1R is found within the juxtamembrane domain (about 61%) and the carboxyl-terminal domain (about 56%) (Ullrich et al., 1986; Ullrich and Schlessinger, 1990). Chimeric receptors consisting of the ligand-binding domain of IR and the cytoplasmic domain of IGF-1R functioned more like the IGF-1R than the IR (Lammers et al., 1989). Similarly, chimeric IGF-1R containing the carboxyl-terminal ß subunit domain of the IR more closely resembled the IR than the IGF-1R (Tartare et al., 1994). To eliminate interactions of ligands with endogenous receptors, other chimeras were generated in which the extracellular portion of the neurotrophin receptor was fused to the intracellular portions of IR or IGF-1R (Siddle et al., 2001). These chimeric molecules were stably expressed in 3T3-L1 fibroblasts (Kalloo-Hosein et al., 1997) or 3T3-L1 adipocytes (Urso et al., 1999,2001) at levels comparable to those of endogenous IR or IGF-1R and activated by nerve growth factor (NGF). The TrkC-IR chimeric receptor was more effective in stimulating physiologically relevant metabolic responses, whereas the TrkC-IGF-1R was more effective in promoting mitogenesis (Urso et al., 1999). Thus, the intracellular domains of the IR and IGF-1R are likely to mediate at least part of the observed receptor specificity.
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III. Signal Transduction via IR and IGF-1R
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A. COMMON SIGNALING PATHWAYS
Many of the intracellular signaling events mediated by activation of the IR and IGF-1R are remarkably similar (White, 1994; Cheatham and Kahn, 1995; LeRoith et al., 1995) (Figure 2). Some of the shared substrates that become phosphorylated by the IGF-1R and IR include members of the insulin receptor substrate (IRS) family of proteins (IRS-1, -2, -3, and -4) (Sun et al., 1991; Lavan and Lienhard, 1993; Patti et al., 1995; Fantin et al., 1998), Gab-1 (Winnay et al., 2000), and Shc (Pelicci et al., 1992). Upon stimulation by insulin or IGF-1, tyrosine-phosphorylated IRS and Shc proteins form signaling complexes between phosphotyrosine-containing binding motifs (YXXM) and Src homology 2 (SH2) domains found in molecules such as growth factor receptor binding-2 protein (Grb2) (Lowenstein et al., 1992; Skolnik et al., 1993) and the p85 regulatory subunit of the phosphatidyl inositol 3' kinase (PI3K) (Backer et al., 1992). The phosphotyrosine residues on IRS-1 also form docking sites for other signaling molecules, including Syp (SHPTP2) (Xiao et al., 1994), Fyn (Sun et al., 1996), Nck (Lee et al., 1993), and Crk (Beitner-Johnson et al., 1996).
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FIG. 2. Multiple signaling pathways for the IGF-1 receptors. These include the Ras/Raf/mitogen-activated protein (MAP) kinase pathways that lead to cell proliferation. For example, the phosphatidyl inositol 3' kinase (PI3K) pathway also has multiple effects and other pathways such as the p38 MAP kinase and Jun kinase (JNK) pathways also affect these biological outcomes. Abbreviations: CT, carboxy-terminal; ERK, extracellular signal-regulated kinase; GDP, guanosine diphosphate; GTP, guanosine triphosphate; IRS, insulin receptor substrate; MEK, mitogen extracellular kinase; PDK, phosphoinositide-dependent kinase; PH, pleckstrin homology domain; PI, phosphatidylinositol; PTEN, phosphatase and tensin homologue deleted on chromosome 10; SHC, Src homology collagen; SHP, Src homology phosphatase.
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By binding to Grb2, IRS proteins couple the IR and IGF-1R to the Ras/mitogen-activated protein kinase (MAPK) pathway. This pathway regulates cell growth, differentiation, and proliferation in response to insulin and IGF-1 (Blenis, 1993; Crews and Erikson, 1993). Various protein tyrosine phosphatases can regulate the activities of the IR and IGF-1R signaling systems.
B. SPECIFICITY
1. Proximal Substrates
To understand the mechanisms involved in the distinct physiological functions of insulin and IGF-1, some investigators searched for specific substrates for IR or IGF-1R. Najjar and coworkers identified pp120, a plasma membrane glycoprotein, which is a substrate for the IR but not for the IGF-1R (Najjar et al., 1997; Soni et al., 2000). Phosphorylation of pp120 is required for its function in insulin endocytosis (Formisano et al., 1995), bile acid transport (Sippel et al., 1994), tumor suppression (Kleinerman et al., 1995), and its inhibitory effect on the mitogenic actions of insulin (Soni et al., 2000). Interestingly, when the carboxyl-terminus of the IGF-1R is replaced by an equivalent region of the IR, the chimeric IGF-1R then can bind to and phosphorylate pp120, decreasing its effect on cell growth (Soni et al., 2000). Mutation of the tyr1316 in the IR, which is not conserved in the IGF-1R, abrogates the insulin-induced tyrosine phosphorylation of pp120 and its ability to suppress the mitogenic action of insulin (Soni et al., 2000).
Some of the other substrates of the IR and IGF-1R are differentially phosphorylated in response to IGF-1 or insulin, indicating that they may mediate specific effects for both ligands. The molecular adapter Grb14 binds specifically to the regulatory kinase loop of the IR and inhibits catalytic activity (Kasus-Jacobi et al., 1998). It recently was shown that Grb14 is three to 10 times less effective at inhibiting the catalytic activity of the IGF-1R than the IR (Bereziat et al., 2002). Rother and coworkers showed that the specificity of signaling may be explained by the preferential use of different substrates by the IR and IGF-1R (Rother et al., 1998). In particular, the IR was coupled preferentially to IRS-2, whereas the IGF-1R was coupled preferentially to IRS-1. This conclusion was confirmed by ablation of the IRS-1 and IRS-2 genes in mice (Araki et al., 1994; Tamemoto et al., 1994; Withers et al., 1998).
In Chinese hamster ovary (CHO) cells stably expressing either the human IR or IGF-1R, it was shown that there are differences in the complement of SH2-containing proteins recruited to IRS-1 by the two receptors (Amoui et al., 2001). In particular, the IGF-1R appears to couple IRS-1 preferentially to Grb2, whereas the IR appears to couple IRS-1 preferentially to the p85 subunit of PI3K (Amoui et al., 2001). In other recent studies, Olefsky and coworkers showed that both the IGF-1R and IR can function as G protein-coupled receptors and engage different G-protein partners. The IGF-1R utilizes Gi, whereas the IR does not (Dalle et al., 2001). In contrast, the IR signals through Gq/11, whereas the IGF-1R does not (Imamura et al., 1999; Dalle et al., 2001). Using the two-hybrid system, Grb10 was found to associate preferentially with the IR in mouse fibroblasts expressing either the IR or IGF-1R (Laviola et al., 1997). Using the same technique, the protein 14-3-3ß was found to bind to the IGF-1R but not to the IR (Furlanetto et al., 1997).
Recently, Ligensa and coworkers identified a new PDZ (postsynaptic density protein-95, disc large, zonula occlusions-1) domain-containing protein (IGF-1 receptor interacting protein-1, IIP-1) that interacts with the C-terminal tail of the IGF-1R but not the IR (Ligensa et al., 2001). Furthermore, the most distal three amino acids in the C-terminal tail of the IGF-1R appear to be crucial for the interaction of IIP-1 with IGF-1R. Indeed, a mutated IR tail carrying the terminal three amino acids of the IGF-1R is able to bind to IIP-1, whereas mutating any of the terminal three amino acids in the IGF-1R tail to the corresponding three amino acids in the IR abolishes the interaction with IIP-1 (Ligensa et al., 2001). Overexpression of IIP-1 in MCF-7 cells does not affect either IGF-1-dependent proliferation or IGF-1-mediated protection from apoptosis but significantly reduces cell motility (Ligensa et al., 2001). Hermanto and coworkers also identified a novel IGF-1R-interacting molecule called RACK1 (Hermanto et al., 2002). RACK1 associates specifically with the IGF-1R but not with the IR, both in yeast and in HEK293T and NIH-3T3 cells overexpressing either the IGF-1R or the IR (Hermanto et al., 2002). RACK-1 is involved in IGF-1R-mediated regulation of cell growth and transformation (Hermanto et al., 2002). However, the interaction of RACK1 with the IGF-1R but not the IR seems to be specific to certain cell types. Indeed, Kiely and coworkers showed that endogenous RACK-1 could interact with both the endogenous IGF-1R and IR in Chinese ovary siemens (COS) cells (Kiely et al., 2002). Nevertheless, these different receptor-specific adaptor proteins, particularly IIP-1 and RACK1, might contribute to the biological specificity of the two hormones.
2. Signaling Pathways
Some evidence suggests that the IR and IGF-1R may phosphorylate the same substrates but use different signaling pathways to mediate the same or different biological effects. For example, insulin induces the expression of vascular endothelial growth factor (VEGF) via the PI3K/Akt pathways in NIH-3T3 cells overexpressing the human IR, whereas IGF-1 induces VEGF expression via the mitogen extracellular kinase (MEK)/MAPK pathway in NIH-3T3 cells overexpressing human IGF-1R (Miele et al., 2000). In rat hepatic stellate cells, insulin and IGF-1 both stimulate cellular proliferation. However, both PI3K and extracellular signal-regulated kinase (ERK) are involved in IGF-1-induced mitogenesis, whereas insulin stimulated mitogenesis through a PI3K-dependent and ERK-independent pathway (Svegliati-Baroni et al., 1999). Interestingly, glycogen synthesis was more effectively stimulated by the IR than by the IGF-1R, although both receptors mediated similar activation of the Akt/protein kinase B (PKB) protein kinase in hepatocytes and in 3T3-L1 fibroblasts (Park et al., 1999). The insulin-specific stimulation of glycogen synthesis appears to involve a rapamycin-sensitive pathway in hepatocytes (Park et al., 1999).
To explain the specificities in the function of the IR and IGF-1R, some investigators suggested that downstream kinases may mediate the specific effects of the IR vs. the IGF-1R. For example, Nakae et al. reported that the transcription factor forkhead homologue to rhabdomyosarcoma (FKHR) is differentially regulated by insulin and IGF-1 in hepatocytes. The phosphorylation of one threonine residue in particular (Thr-24) appears to be induced by insulin but not by IGF-1. As this residue can be phosphorylated by PKB in vitro and PKB is also activated by IGF-1 in these cells, the authors proposed that a PKB-like kinase specifically activated by insulin may mediate this effect (Nakae et al., 2000). More recently, it has been shown that although both insulin and IGF-1 induce proliferation of murine skin keratinocytes, the action of insulin — but not IGF-1 — is mediated specifically via a protein kinase C delta (PKC
) and involves activation of the sodium/potassium (Na+/K+) pump (Shen et al., 2001). Thus, PKC is a multifunctional serine kinase that represents a divergence point in IR and IGF-1R signaling. In this same cell type, insulin and IGF-1 stimulate the translocation of different glucose transporters, although they both increase glucose uptake (Shen et al., 2001). Thus, insulin and IGF-1 can mediate the same or different biological responses by utilizing different signaling pathways or different intracellular mediators.
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IV. Induction of Specific Genes by the IR and IGF-1R
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Some reports have shown that insulin and IGF-1 can act on the same genes but with different outcomes. For example, in murine skin keratinocytes, insulin stimulates the expression of differentiation markers, whereas IGF-1 inhibits them (Wertheimer et al., 2000). Also, in the developing eye lens of the chicken, the level of delta-crystallin induced by IGF-1 is greater and occurs more quickly than that induced by insulin (Alemany et al., 1989). It has been shown that low concentrations of IGF-1 (10 nM) increase the expression of uncoupling protein 3 (UCP-3) by 2-fold, whereas much higher concentrations of insulin (860 nM) are necessary to obtain the same effect in human neuroblastoma SH-SYSY cells (Gustafsson et al., 2001).
cDNA microarray analysis recently has been established as a powerful tool to study the effects of hormones on cellular metabolism and gene regulation on a genomic scale. Until now, this technology was used to define the effects of IGF-1 on gene expression in different cell lines (Liu et al., 2001; Oh et al., 2002) but not to compare the different gene-expression profiles induced by insulin and IGF-1. We used cDNA microarray expression profiling to identify genes that are regulated differently by IGF-1 and insulin in mouse fibroblast NIH-3T3 cells (Dupont et al., 2001b) as a first step towards understanding the molecular basis for the different functions of the IGF-1R and the IR.
A. DIFFERENTIAL REGULATION OF GENE-EXPRESSION PATTERNS BY INSULIN AND IGF-1 IN NIH-3T3 FIBROBLASTS
The biological and physiological comparison of the IR and IGF-1R is complicated by the fact that each ligand can cross-react with the other receptor and hybrid receptors can form when both receptors are expressed in the same cells. To circumvent these problems, we have compared the effect of insulin and IGF-1 in NIH-3T3 fibroblasts overexpressing either human IR (IR cells) (Levy-Toledano et al., 1993) or human IGF-1R (NWTb3 cells) (Blakesley et al., 1995,1996). NWTb3 and IR cells were incubated in the presence or absence of IGF-1 (50 nM) or insulin (50 nM) for 90 minutes, respectively. Of the 2221 genes on the mouse cDNA microarrays, we found that the expression levels of 30 were significantly induced by IGF-1 but not by insulin. In contrast, only nine genes and one expressed sequence tag (EST) were upregulated specifically by insulin but not by IGF-1 (Tables I and II). We confirmed the IGF-1- and insulin-induced regulation for 10 of these genes by Northern analysis (Figure 3). The genes that were identified as regulated by IGF-1 and insulin are involved in various cellular functions, including proliferation, differentiation, apoptosis, cellular processes, and metabolism (Tables I and II). Interestingly, most of these genes were not known previously to be regulated by either IGF-1 or insulin. Indeed, only three genes — the Jun oncogene (Chiou and Chang, 1992; Monnier et al., 1994), 5 integrin (Palmade et al., 1994), and the early growth response-1 transcription factor (EGR-1) (Jhun et al., 1995) — had been reported to be induced by IGF-1. Furthermore, more than half of the genes upregulated by IGF-1 are associated with mitogenesis and differentiation, whereas none of the genes specifically upregulated by insulin are associated with these processes. IGF-1, but not insulin, induced the expression of two cytokine receptors (interleukin (IL) receptors 3 and 4) that have been reported to be involved in the regulation of cell growth (Keegan et al., 1994). IGF-1 also induced the expression of glial cell line-derived neurotrophic factor (GNDF), which is known to be crucial for the development and the maintenance of various neurons (Airaksinen and Saarma, 2002). IGF-1 increased the expression of the Wee-1-like kinase, which is involved in cell-cycle progression (Helmbrecht et al., 2000), and the EGR-1 transcription factor, which is known to enhance cell proliferation. These results suggest that IGF-1-induced cellular proliferation is a tightly regulated process.
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FIG. 3. Confirmation of the specific gene expression by IGF-1 or insulin using Northern blot analysis. Northern blot analysis was performed using RNA from cells expressing IGF-1 receptors (B3 and C43) or insulin receptors (IR), following stimulation. The specificity of stimulated gene expression correlated with the microarray results seen in Tables I and II. Abbreviations: CSF, colony-stimulating factor; EGR, early growth response; GDNF, glial cell line-derived neurotrophic factor; GLVR, gibbon ape leukemia virus receptor; PRLR, prolactin receptor; TDAG, T-cell death-associated gene. [Reprinted with permission from Dupont J, Khan J, Qu BH, Metzler P, Helman L, LeRoith D 2001 Insulin and IGF-1 induce different patterns of gene expression in mouse fibroblast NIH-3T3 cells: identification by cDNA microarray analysis. Endocrinology 142:4969–4975. Copyright The Endocrine Society.]
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Our study also suggested that insulin and IGF-1 are involved in the apoptosis process. IGF-1 treatment increased expression of the T-cell death-associated gene (TDAG)-51 and Daxx (Fas-binding) genes, whereas insulin increased expression of apoptotic protease-activating factor-1 (APAF-1) and seven in absentia homologue-1B (SIAH-1B) (Tables I and II). Importantly, IGF-1 is capable of increasing the expression of antiapoptotic genes such as Twist (Maestro et al., 1999). Thus, the induction of IGF-1- or insulin-specific genes could explain the specificity of the biological effects of these two hormones.
B. TWIST EXPRESSION IS SPECIFICALLY INDUCED BY IGF-1
In a separate study, we studied Twist, one of the genes that was specifically induced by the IGF-1-responsive gene (Dupont et al., 2001a). Twist belongs to the basic helix-loop-helix family of transcription factors, which play a central role in cell-type determination and differentiation in both vertebrates and invertebrates (Olson and Klein, 1994). IGF-1 treatment increased the abundance of Twist mRNA in NWTb3 cells, whereas insulin failed to increase Twist mRNA in IR cells. The IGF-1-induced increase in Twist expression requires activation of IGF-1R, since Twist mRNA expression was not induced in response to IGF-1 in parental NIH-3T3 cells, which express few IGF-1Rs, nor in the NKR (NIH-3T3 cells expressing an IGF-1 receptor with lysine-to-arginine substitution) cell line, which overexpresses the dominant-negative human IGF-1R (Kato et al., 1993). We also showed that injection of IGF-1 via the inferior vena cava increased Twist mRNA expression in muscle. We used various pharmacological inhibitors and a MEK-1 dominant-negative construct to investigate which IGF-1R signaling pathway was involved in the induction of Twist gene expression. These experiments demonstrated that the MEK/MAPK pathway plays a critical role in IGF-1-induced Twist expression. Using an antisense strategy, we also showed that Twist is positively involved in the antiapoptotic effects of the IGF-1R. These studies show that a gene that is regulated by IGF-1 receptor activation may, in turn, regulate the function of the IGF-1 receptor function.
C. IGF-1-INDUCED GENES AND CANCER PROGRESSION
IGF-1-regulated genes were studied in a preneoplastic, immortalized breast cell line, 184htert. The advantage of utilizing this cell line is that genes may be identified that are affected by IGF-1 and the IGF-1 receptor signaling pathways at an early stage in the progression of cancer. These cells were created by using retroviral technology to introduce the human telomerase reverse transcriptase gene into normal breast epithelial cells. Of the 
2000 known genes on the microarray chip, 156 (8%) were regulated by IGF-1. These genes exhibited various patterns of regulation; whereas some were either up- or downregulated at early time points, others were regulated in a biphasic manner. The IGF-1-responsive genes could be subdivided into various categories (e.g., transcription factors, cell cycle-related genes, genes involved in cancer progression, signaling-related genes, extracellular matrix genes, genes related to metabolism). Interestingly, IGF-1 regulated a large number of genes involved in angiogenesis. Many genes known to stimulate angiogenesis were upregulated by IGF-1, whereas inhibitors of angiogenesis such as plasminogen activator inhibitor-1 (PAI-1) and metalloproteases were inhibited by IGF-1 (Table III). The effects of IGF-1 on many of these genes — including c-fos, VEGF, Fas ligand, cyp1A1, cyp1B1, interleukin-1ß, and uPA — were validated by other techniques. Many of the genes that are regulated by IGF-1 are also responsive to the hypoxia-inducible factor-1alpha (HIF-1) and cAMP response binding protein (CREB) transcription factors. Indeed, IGF-1 induced nuclear translocation of HIF-1 and the phosphorylated form of CREB, thereby inducing gene expression.
Thus, this study demonstrated that IGF-1 regulates the expression of many genes involved in cancer progression. This new information may be helpful when considering gene targeting for therapeutic uses in the treatment of cancer.
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IV. Conclusion and Future Directions
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The purpose of this review is to bring the possible applications of this exciting new technology to the attention of researchers. While cDNA microarray analysis is associated with a number of technical hurdles and experimental flaws, its utility has enormous potential, if used carefully, with multiple controls and constant validation. This approach may prove to be useful as a rapid screening test to identify the many genes that are differentially regulated in different tissues and systems. In particular, it may lead to the identification of genes not previously known to be affected by a particular process. Furthermore, in studying the progression of disease states and variations in gene expression, in various models, it may prove to be the more rapid and economical method. Numerous other applications undoubtedly will emerge as more investigators utilize this technology.
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TOP
ABSTRACT
I. Introduction
