Alternative titles; symbols
HGNC Approved Gene Symbol: CHTOP
Cytogenetic location: 1q21.3 Genomic coordinates (GRCh38): 1:153,634,066-153,646,306 (from NCBI)
By PCR of a HeLa cell cDNA library, Zullo et al. (2009) cloned 3 splice variants of CHTOP, which they called SRAG, SRAG3, and SRAG5. SRAG encodes the full-length 248-amino acid protein, which contains a central arginine- and glycine-rich domain similar to that found in nucleolin (NCL; 164035) and fibrillarin (FBL; 134795). Compared with the full-length SRAG protein, SRAG3 contains an N-terminal deletion, and SRAG5 encodes a protein with a partial deletion within the arginine- and glycine-rich domain. Western blot analysis of mouse tissues revealed widespread Srag expression, with highest levels in thymus, spleen, and lymph node. Srag3 was expressed in lung only, and Srag5 was expressed in stomach only. Endogenous HEK293 cell SRAG protein migrated as a doublet with an apparent molecular mass of about 28 kD. Cell fractionation and extraction experiments revealed that SRAG was predominantly a nuclear protein, with some nucleolar localization. SRAG5, but not SRAG3, was also detected in nucleoli. However, immunofluorescence microscopy of transfected HeLa cells revealed that fluorescence-tagged SRAG and SRAG5 were nucleolar in 25 to 35% of cells, whereas SRAG3 was nucleolar in nearly 100% of cells. Database analysis revealed SRAG orthologs in vertebrates, but not invertebrates.
Van Dijk et al. (2010) cloned mouse Chtop, which they designated Fop, and by database analysis, they identified full-length human FOP. The mouse and human FOP proteins both contain 249 amino acids. A second FOP isoform may be translated from a conserved internal methionine at residue 26. Full-length FOP has a central glycine- and arginine-rich (GAR) domain and 2 copies of a 9-amino acid sequence near the C terminus. Immunohistochemical analysis of mouse erythroleukemia (MEL) cells revealed a punctate nuclear distribution. In day-16.5 mouse embryos, Fop protein was expressed in heart, lung, gut, kidney, submandibular gland, thymus, follicles of the vibrissae, muscle, brown fat, and neuronal cells of brain, olfactory epithelium, and dorsal root ganglia. Fop tightly associated with facultative heterochromatin in MEL cells. Time-lapse imaging showed that fluorescence-tagged Fop released from condensed chromosomes during mitosis and relocalized to chromatin after cell division.
Using mutation analysis, Zullo et al. (2009) showed that the N-terminal 40 amino acids of full-length SRAG functioned as a nucleolar exclusion signal. Treatment with an RNase or disruption of nucleolar ribosomal RNA reduced nucleolar SRAG localization, suggesting that nucleolar SRAG depends upon RNA. Srag protein expression was reduced following activation of mouse thymocytes and during the G2/M phase of the cell cycle in HeLa cells. Overexpression of SRAG in HeLa cells slowed cell growth by reducing entry into the G2/M phase and by enhancing cell death.
Using fetal mouse liver cells expressing the entire human beta-globin cluster (see HBB; 141900), van Dijk et al. (2010) found that knockdown of Fop via lentivirus infection upregulated the expression of human fetal gamma-globin (see HBG1; 142200) within the beta-globin cluster, in addition to upregulating expression of the embryonic mouse beta-globin genes, epsilon-Y and beta-H1. Silencing of Fop did not alter expression of the adult mouse globin genes beta-major and alpha-globin (see HBA1; 141800). Depletion of FOP from adult human erythroid progenitor cells increased expression of embryonic epsilon-globin (HBE1; 142100) and gamma-globin and decreased expression of adult beta-globin mRNA. The embryonic zeta-globin gene (HBZ; 142310) within the alpha-globin cluster was also reactivated following FOP depletion. FOP appeared to modulate SOX6 (607257)-dependent silencing of gamma-globin in the adult erythroid progenitor cells. Van Dijk et al. (2010) concluded that FOP is a critical regulator of fetal globin gene expression.
Using yeast 2-hybrid and coimmunoprecipitation analyses, van Dijk et al. (2010) found that mouse Fop interacted with the methyltransferase Prmt1 (602950). The GAR domain of Fop was heavily methylated by Prmt1 and Prmt5 (604045) in vitro and in vivo, and both methyltransferases competed for Fop binding. Knockdown of FOP in estrogen-responsive MCF7 human breast cancer cells via small interfering RNA almost completely blocked estrogen-induced promoter occupancy by ER-alpha (ESR1; 133430) and reduced expression of the estrogen-responsive genes PS2 (TFF1; 113710), lactoferrin (LTF; 150210), and TGF-alpha (TGFA; 190170).
Hartz (2011) mapped the CHTOP gene to chromosome 1q21.3 based on an alignment of the CHTOP sequence (GenBank AL050260) with the genomic sequence (GRCh37).
Zullo et al. (2009) found that Srag knockout in mice was embryonic lethal and incompatible with viability in mouse embryonic fibroblasts and stem cells.
In a study of 1,751 knockout alleles created by the International Mouse Phenotyping Consortium (IMPC), Dickinson et al. (2016) found that knockout of the mouse homolog of human CHTOP is homozygous-lethal (defined as absence of homozygous mice after screening of at least 28 pups before weaning). Chtop homozygous knockout embryos showed obvious developmental delay, neural tube defects, craniofacial dysmorphology, abnormal eye development, and subcutaneous edema. High-resolution transmission electron microscopy (HREM) at E14.5 further showed major abnormalities in ribs and vertebrae, cardiovascular system, and nervous system.
Dickinson, M. E., Flenniken, A. M., Ji, X., Teboul, L., Wong, M. D., White, J. K., Meehan, T. F., Weninger, W. J., Westerberg, H., Adissu, H., Baker, C. N., Bower, L., and 73 others. High-throughput discovery of novel developmental phenotypes. Nature 537: 508-514, 2016. Note: Erratum: Nature 551: 398 only, 2017. [PubMed: 27626380] [Full Text: https://doi.org/10.1038/nature19356]
Hartz, P. A. Personal Communication. Baltimore, Md. 6/15/2011.
van Dijk, T. B., Gillemans, N., Pourfarzad, F., van Lom, K., von Lindern, M., Grosveld, F., Philipsen, S. Fetal globin expression is regulated by Friend of Prmt1. Blood 116: 4349-4352, 2010. [PubMed: 20688955] [Full Text: https://doi.org/10.1182/blood-2010-03-274399]
van Dijk, T. B., Gillemans, N., Stein, C., Fanis, P., Demmers, J., van de Corput, M., Essers, J., Grosveld, F., Bauer, U.-M., Philipsen, S. Friend of Prmt1, a novel chromatin target of protein arginine methyltransferases. Molec. Cell. Biol. 30: 260-272, 2010. [PubMed: 19858291] [Full Text: https://doi.org/10.1128/MCB.00645-09]
Zullo, A. J., Michaud, M., Zhang, W., Grusby, M. J. Identification of the small protein rich in arginine and glycine (SRAG): a newly identified nucleolar protein that can regulate cell proliferation. J. Biol. Chem. 284: 12504-12511, 2009. [PubMed: 19254951] [Full Text: https://doi.org/10.1074/jbc.M809436200]