Alternative titles; symbols
HGNC Approved Gene Symbol: GRB10
Cytogenetic location: 7p12.1 Genomic coordinates (GRCh38): 7:50,590,068-50,793,453 (from NCBI)
Src homology region-2 (SH2) domain proteins bind to autophosphorylated growth factor receptors after activation of the receptors by ligand. By screening an NIH 3T3 library with epidermal growth factor receptor (EGFR; 131550) by use of the CORT technique (Skolnik et al., 1991), Ooi et al. (1995) cloned mouse Grb10.
By using the yeast 2-hybrid system to identify proteins that interact with the cytoplasmic tyrosine kinase domain of the insulin receptor (INSR; 147670), Liu and Roth (1995) isolated a GRB10 cDNA from HeLa cells. The cDNA, called GRBIR by them, encodes a predicted 548-amino acid protein containing an SH2 domain and an incomplete pleckstrin-homology (PH) domain. RT-PCR showed that the GRB10 gene is alternatively spliced, producing transcripts that encode proteins either with or without a 46-amino acid stretch that contains part of the PH domain. The SH2 and PH domains of the human GRB10 protein are 99% and 84% identical, respectively, to those of mouse Grb10. Northern blot analysis showed that the GRB10 gene is expressed as 2.2-, 5.0-, and 6.5-kb transcripts, predominantly in skeletal muscle and pancreas. Western blot analysis of HeLa cell lysates detected 50-, 65-, and 68-kD proteins.
By searching for ESTs that encode potential SH2 domains, Frantz et al. (1997) identified a human GRB10 cDNA, which they called GRB-IR-beta/GRB10, that was derived from a cerebellum cDNA library. This GRB10 cDNA encodes a predicted 536-amino acid protein that is identical to the GRB10 identified by Liu and Roth (1995), except that it contains a complete PH domain and is missing an N-terminal extension. These 2 proteins appeared to result from alternative mRNA splicing. By RT-PCR of skeletal muscle mRNA, Frantz et al. (1997) detected 3 GRB10 transcripts, including those representing the GRB10 cDNAs characterized by themselves and by Liu and Roth (1995). Frantz et al. (1997) found by Northern blot analysis that GRB10 is widely expressed as a 5.6-kb mRNA, with additional 3.1- and 4.8-kb transcripts in skeletal muscle.
Dong et al. (1997) cloned the gamma isoform of GRB10, which is identical to the alpha isoform except for a 138-nucleotide addition in the PH domain encoding an additional 46 amino acids, from a human muscle cDNA library using a GRB10-alpha cDNA as probe.
Monk et al. (2009) determined that the GRB10 gene contains 7 noncoding exons, which are located 5-prime of exon 1, and termed UN1A, UN1, UN2, UN3, UN3.1, UN3.2, and UN4. There are 19 coding exons, including exons 1, 1a, and 1b and exons 7 and 7a.
Using a human/rodent somatic cell hybrid panel, a radiation hybrid panel, and fluorescence in situ hybridization, Jerome et al. (1997) mapped the human GRB10 gene to chromosome 7p12-p11.2. By sequence analysis, Dong et al. (1997) confirmed the mapping of GRB10 to chromosome 7p12-p11.2.
Ooi et al. (1995) mapped the mouse Grb10 gene to chromosome 11.
Ooi et al. (1995) found that mouse Grb10 underwent serine but not tyrosine phosphorylation after EGF treatment and bound poorly to EGFR, suggesting that another protein binds EGFR in vivo.
Liu and Roth (1995) found that GRB10 bound with high affinity to autophosphorylated INSR in vitro. After treatment of cells with insulin (INS; 176730), GRB10 formed complexes with INSR. Formation of this complex inhibited the insulin-induced increase in phosphorylation of IRS1 (147545) and a 60-kD GTPase-activating protein (GAP)-associated protein, suggesting that GRB10 inhibits or redirects the INSR signaling pathway.
Frantz et al. (1997) found that, upon insulin stimulation, the 65-kD form of GRB10 translocated from the cytosol to the membrane. GRB10 also bound activated platelet-derived growth factor receptor (PDGFRB; 173410) and epidermal growth factor receptor, suggesting that GRB10 functions downstream from activated insulin and growth factor receptors.
GRB10 is closely related to GRB7 (601522) and GRB14 (601524); Jerome et al. (1997) stated that each contains a proline-rich N-terminal motif, a central pleckstrin-homology domain (see 173570), and a C-terminal SH2 domain.
Using a yeast 2-hybrid assay, Giovannone et al. (2003) showed that mouse Gigyf1 (612064) bound Grb10. Mutation analysis showed that the interaction required the GYF domain of Gigyf1 and at least 2 of the 3 proline-rich regions in the N terminus of Grb10. In mouse fibroblasts expressing Igf1r, a fragment of Gigyf1 containing the GYF domain bound Grb10 in the basal state. Stimulation with Igf1 resulted in increased binding of Gigyf1 to Grb10 and transient binding of Gigyf1 and Grb10 to Igf1r, presumably via the adaptor function of Grb10. At later time points, Gigyf1 dissociated, but Grb10 remained linked to Igf1r. Overexpression of the Grb10-binding fragment of Gigyf1 resulted in a significant increase in Igf1-stimulated Igf1r tyrosine phosphorylation. Giovannone et al. (2003) concluded that GRB10 and GIGYF1 may act cooperatively to regulate IGF1R signaling.
Yu et al. (2011) used large-scale quantitative phosphoproteomics experiments to define the signaling networks downstream of mammalian target of rapamycin (mTOR) complex-1 (mTORC1; see 601231) and mTORC2. Characterization of an mTORC1 substrate, Grb10, showed that mTORC1-mediated phosphorylation stabilized Grb10, leading to feedback inhibition of the phosphatidylinositol 3-kinase (PI3K; see 171834) and extracellular signal-regulated/mitogen-activated protein kinase (ERK/MAPK; see 176872) pathways. Grb10 expression is frequently downregulated in various cancers, and loss of Grb10 and loss of the well-established tumor suppressor phosphatase PTEN (601728) appear to be mutually exclusive events, suggesting that Grb10 might be a tumor suppressor regulated by mTORC1.
Imprinting
In a systematic screen for maternally expressed imprinted genes using subtraction hybridization with androgenetic and normal fertilized mouse embryos, Miyoshi et al. (1998) isolated 5 candidate maternally expressed genes (Megs) and 2 genes known to be maternally expressed, H19 (103280) and p57(Kip2) (CDKN1C; 600856). They demonstrated that one imprinted gene, Meg1, is apparently identical to Grb10, which is located on the proximal portion of mouse chromosome 11. GRB10 protein binds to the insulin receptor and the insulin-like growth factor I receptor (IGF1R; 147370) via its SH2 domain and inhibits the associated tyrosine kinase activity that is involved in the growth-promoting activities of insulin and insulin-like growth factors I (IGF1; 147440) and II (IGF2; 147470). Thus, it is probable that Meg1/Grb10 is responsible for the imprinted effects of prenatal growth retardation or growth promotion caused by maternal or paternal duplication of proximal chromosome 11 with reciprocal deficiencies, respectively.
Using allele-specific transcription analysis in various fetal tissues, Blagitko et al. (2000) found that human GRB10 is imprinted in a highly isoform- and tissue-specific manner. In fetal skeletal muscle, GRB10 isoform gamma-1 is expressed from the maternal allele alone, whereas in numerous other fetal tissues, all GRB10 splice variants are transcribed from both parental alleles. A remarkable finding was the paternal-specific expression of GRB10 in human fetal brain, since in the mouse, the gene is transcribed exclusively from the maternal allele. Imprinted expression in human fetal brain is not accompanied by allele-specific methylation of the most 5-prime CpG island. This appeared to be the first example of a gene that is oppositely imprinted in mouse and human.
To investigate the discrepant imprinting between mouse and human GRB10, Arnaud et al. (2003) compared the sequence organization of their upstream regions and examined their allelic methylation patterns and the splice variant organization of the mouse locus. Both maternal and paternal expression of mouse Grb10 was detected. Expression of the paternal allele arose from a different promoter region than the maternal allele and, as in human, was restricted to the brain. The upstream regions were well conserved, especially the presence of 2 CpG islands. Both genes had a similar imprinted methylation pattern, and the second CpG island (CGI2) was a differentially methylated region (DMR) with maternal methylation in both species. Analysis of 24 patients with Silver-Russell syndrome without maternal uniparental isodisomy of chromosome 7 (see SRS2, 618905) (see MOLECULAR GENETICS) did not reveal methylation anomalies in the DMR. Arnaud et al. (2003) suggested that the difference in imprinted expression in mouse and human is not due to acquisition of an imprint mark, but rather in differences in reading of this mark.
Monk et al. (2009) explored the conservation of reciprocal imprinting of the GRB10 gene in human fetal tissues. As in mice, human GRB10 was paternally expressed in brain and spinal cord. Maternal allele-specific expression was conserved only in placental villous trophoblasts, an essential part of the placenta involved in nutrient transfer. All other fetal tissues, including lung, limb, umbilical cord, skin, kidney, adrenal gland, pancreas, liver and heart showed equal biallelic expression. The authors suggested that maternal GRB10 expression in placenta may be evolutionarily important, presumably in the control of fetal growth. Maternal transcripts originated from exons UN1 or UN1A located several kilobases upstream of CGI2, which is the imprinting control region (ICR), and brain-specific paternal expression originated from exon UN2 within the ICR. Both maternal and paternal expression in humans showed mechanistic similarities with the mouse. The conserved CpG island, CGI2, was DNA methylated on the maternal allele and was marked on the paternal allele by developmentally regulated bivalent chromatin, with the presence of both H3K4 and H3K27 methylation. The strong conservation of the opposite allelic expression in placenta versus brain supports the hypothesis that GRB10 imprinting may have evolved to mediate diverse roles in mammalian growth and behavior.
In the mouse, Garfield et al. (2011) demonstrated that within the brain Grb10 is expressed from the paternal allele from fetal life into adulthood and that ablation of this expression engenders increased social dominance specifically among other aspects of social behavior, a finding supported by the observed increase in allogrooming by paternal Grb10-deficient animals. Grb10 was, therefore, the first example of an imprinted gene that regulates social behavior. It was also alone in exhibiting imprinted expression from each of the paternal alleles in a tissue-specific manner, as loss of the peripherally expressed maternal allele leads to significant fetal and placental overgrowth. Thus, Grb10 was at that time a unique imprinted gene, able to influence distinct physiologic processes, fetal growth, and adult behavior, owing to actions of the 2 paternal alleles in different tissues.
Mapping of the GRB gene to 7p made it a candidate gene for Silver-Russell syndrome (SRS2; 618905). In humans, maternal uniparental disomy 7 is responsible for approximately 10% of cases of SRS, which has effects including pre- and postnatal growth retardation and other dysmorphologies. GRB10 has a suppressive effect on growth through its interaction with either the IGF-I receptor (IGF1R; 147370) or the growth hormone receptor. Yoshihashi et al. (2000) demonstrated that the GRB10 gene is monoallelically expressed in the human fetal brain tissues and is transcribed from the maternally derived allele in somatic cell hybrids. Hence, human GRB10 is imprinted. By mutation analysis of GRB10 in 58 unrelated patients with SRS, Yoshihashi et al. (2000) identified a pro95-to-ser substitution within the N-terminal domain of the protein in 2 of the patients. In these 2 cases, the mutant allele was inherited from the mother. The fact that monoallelic GRB10 expression was observed from the maternal allele suggests that the maternally transmitted mutant allele contributed to the SRS phenotype.
The role of GRB10 in Silver-Russell syndrome was cast in doubt by the reports of Hannula et al. (2001), Hitchins et al. (2001), and McCann et al. (2001). Hannula et al. (2001) reported a patient with SRS and maternal uniparental disomy of a narrow segment of chromosome 7, 7q31-qter, and biparental inheritance of the rest of chromosome 7. Two imprinted genes residing in the uniparental region were MEST/PEG1 (601029) and COPG2 (604355). GRB10 at 7p12-p11.2 was located within the region of biparental inheritance. Hitchins et al. (2001) showed repression of the maternal allele in human fetal brain and spinal cord, with biallelic expression in a wide range of other organs and peripheral tissues. No mutations were found in a screening of all 16 exons of the GRB10 gene by sequencing in 18 classic SRS patients, where major structural chromosomal abnormalities and matUPD7 had been excluded. These findings suggested to the authors that this gene is unlikely to contribute to SRS in a significant number of patients or to be responsible for the full disease spectrum. Using RT-PCR, McCann et al. (2001) confirmed that GRB10 imprinting in brain and muscle is isoform specific, and they demonstrated absence of imprinting in growth plate cartilage, the tissue most directly involved in linear growth. Thus they considered it unlikely that GRB10 is the gene responsible for Silver-Russell syndrome.
To investigate the function of the Grb10 adaptor protein, Charalambous et al. (2003) generated mice in which the Grb10 gene was disrupted by a gene-trap insertion. The experiments confirmed that Grb10 is subject to genomic imprinting with most of Grb10 expression arising from the maternally inherited allele. Consistent with this, disruption of the maternal allele resulted in overgrowth of both the embryo and the placenta such that mutant mice were approximately 30% larger than normal at birth. This observation established Grb10 as a potent growth inhibitor. Charalambous et al. (2003) suggested that, in at least some cases of RSS, changes in GRB10 dosage account for the severe growth retardation that is characteristic of the disorder. Because Grb10 is a signaling protein capable of interacting with tyrosine kinase receptors, the authors used genetic crosses to test whether Grb10 acts downstream of Igf2, a paternally expressed growth-promoting gene. The result indicated that Grb10 action is essentially independent of Igf2, providing evidence that imprinting acts on at least 2 major fetal growth axes in a manner consistent with parent-offspring conflict theory.
Mice with maternal duplication of proximal chromosome 11 (MatDp(prox11)), where Meg1/Grb10 is located, exhibit pre- and postnatal growth retardation. Shiura et al. (2009) generated model mice mimicking the pattern of imprinted gene expression observed in the MatDp(prox11) by deleting the differentially methylated region of Meg1/Grb10 (Meg1-DMR). Neighboring genes of Meg1/Grb10, such as COBL (610317) and DDC (107930), also comprised the imprinted region. Paternal deletion of the Meg1-DMR (+/delta-DMR) caused both upregulation of the maternally expressed Meg1/Grb10 type I transcript in the whole body and Cobl in the yolk sac and loss of paternally expressed Meg1/Grb10 type II transcript and Ddc in the neonatal brain and heart, respectively, demonstrating maternalization of the entire Meg1/Grb10 imprinted region. The +/delta-DMR mice exhibited the same growth abnormalities as the MatDp(prox11) mice. Fetal and neonatal growth were very sensitive to the expression level of Meg1/Grb10 type I transcript, indicating that the 2-fold increment of the Meg1/Grb10 type I transcript is one of the major causes of the growth retardation observed in the MatDp(prox11) and +/delta-DMR mice. Shiura et al. (2009) suggested that the corresponding human GRB10 type I transcript may play a role in the etiology of Silver-Russell syndrome caused by partial trisomy of chromosome 7p13-p11.
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Blagitko, N., Mergenthaler, S., Schulz, U., Wollmann, H. A., Craigen, W., Eggermann, T., Ropers, H.-H., Kalscheuer, V. M. Human GRB10 is imprinted and expressed from the paternal and maternal allele in a highly tissue- and isoform-specific fashion. Hum. Molec. Genet. 9: 1587-1595, 2000. [PubMed: 10861285] [Full Text: https://doi.org/10.1093/hmg/9.11.1587]
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Yu, Y., Yoon, S.-O., Poulogiannis, G., Yang, Q., Ma, X. M., Villen, J., Kubica, N., Hoffman, G. R., Cantley, L. C., Gygi, S. P., Blenis, J. Phosphoproteomic analysis identifies Grb10 as an mTORC1 substrate that negatively regulates insulin signaling. Science 332: 1322-1326, 2011. [PubMed: 21659605] [Full Text: https://doi.org/10.1126/science.1199484]