Abstract: The rate-limiting step in ribosome biogenesis is the transcription of ribosomal RNA, which is controlled by environmental conditions. The JmjC enzyme KDM2A/JHDM1A/FbxL11 demethylates mono- and dimethylated Lys 36 of histone H3, but its function is unclear. Here, we show that KDM2A represses the transcription of ribosomal RNA. KDM2A was localized in nucleoli and bound to the ribosomal RNA gene promoter. Overexpression of KDM2A repressed the transcription of ribosomal RNA in a demethylase activity-dependent manner. When ribosomal RNA transcription was reduced under starvation, a cell-permeable succinate that inhibited the demethylase activity of KDM2A prevented the reduction of ribosomal RNA transcription. Starvation reduced the levels of mono- and dimethylated Lys 36 of histone H3 marks on the rDNA promoter, and treatment with the cell-permeable succinate suppressed the reduction of the marks during starvation. The knockdown of KDM2A increased mono- and dimethylated Lys 36 of histone H3 marks, and suppressed the reduction of ribosomal RNA transcription under starvation. These results show a novel mechanism by which KDM2A activity is stimulated by starvation to reduce ribosomal RNA transcription.
Abstract: Transcription factor binding sites are short DNA sequences that interact with transcription factors and the proper control of gene expression appears to require the mechanisms including the regulation through the genome context around the transcription factor binding sites. The MYC proteins are central regulators of cell growth. Many genes have been reported to be regulated by MYC through E-box sites. However, the characters of E-box that Myc selects to function are not clear and identification of additional genes controlled by MYC will provide information to completely understand the functions of MYC. Here we report that MYC directly induces TAF4b expression. We mapped the transcription start site and characterized functional promoter elements for MYC response in the TAF4b promoter. There are several E-box sequences near the transcription start site, including canonical (CACGTG) and non-canonical (CGCGTG) ones. We found that c-MYC induces TAF4b expression through one of the non-canonical E-box sites, which is in a highly conserved region of TAF4b promoters in mammals, suggesting the importance of the genome context around the target E-box. When the non-canonical E-box in the TAF4b promoter was mutated to a canonical one, MYC functioned on both E-boxes, while another E-box-binding transcription factor, USF, did so on only the canonical E-box. These results suggest that in addition to the context where the target E-box exists, a sequence within an E-box is involved in the mechanisms by which specific E-box sites are selected by Myc.
Abstract: Mina53 (mina) was identified as a gene, which is directly induced by the oncogene c-myc. Elevated expression of Mina53 protein was found in >80% of colon cancer and esophageal squamous cell carcinoma (ESCC). Patients with high expression of Mina53 had shorter survival, suggesting the prognostic usefulness of Mina53. We studied Mina53 expression in lymphoma subtypes to examine its diagnostic significance and its possible role in lymphoma-genesis. Surgical cases of 28 lymphoma and 4 non-neoplastic tissues were stained immunochemically using anti-Mina53 monoclonal antibody. Mina53 expression correlated well with c-Myc expression in lymphoma, suggesting that c-Myc is a controlling factor for mina53 expression also in lymphomas. Although the expression of Mina53 as well as c-Myc was less frequent in lymphoma compared with those of colon and ESCC, increased expression of Mina53 was found in Burkitt-like lymphoma (1/1), Hodgkin's lymphoma (3/5), diffuse large B cell lymphoma (DLBCL) (5/13), lymphomas with a transition from follicular to DLBCL (1/2), with none in follicular (0/4) and T cell lymphoma (0/3). Analyses of the data suggested that Mina53 was frequently expressed in aggressive types of B cell lymphoma. To get more information about the expression of Mina53 in DLBCL, which most frequently occurs among lymphomas, we analyzed the expression of Mina53 in another 21 DLBCL specimens, which were in more advanced stages than those described above. The expression level of Mina53 correlated to the international prognostic index (IPI) values with statistical significance (r=0.477, P=0.0275). Notably, in this group, Mina53 expression did not correlate with c-Myc expression, suggesting that other factor(s) besides c-Myc largely affect the expression of Mina53 in advanced DLBCL. These results suggest that although Mina53 expression is not prominent in lymphoma in general, it may be related to tumor progression of B cell lymphoma.
Abstract: Recently we have identified a novel gene mina53 (mina), which is a direct transcriptional target of oncoprotein Myc. Mina53 protein was shown to be highly expressed in tumour cells and to play a role in cell proliferation. Here we report the expression of Mina53 in mouse testis, which contains proliferating cells and expresses many cancer-related genes. Immunohistochemical studies by using newly produced monoclonal antibody to Mina53 showed that Mina53 was expressed in the nuclei of spermatogonia. Mina53 was also expressed in meiotic prophase cells such as preleptotene, leptotene and zygotene, and weakly in early pachytene spermatocytes, but was absent in late pachytene spermatocytes, spermatids and mature sperm. The expression pattern of Mina53 was quite similar to that of proliferation cell nuclear antigen (PCNA). Using experimental cryptorchid testis, it was found that Mina53 was highly expressed in undifferentiated spermatogonia, which were PCNA-positive. These results suggest that Mina53 is prominently expressed in proliferating, undifferentiated spermatogonia, and plays a role in cell proliferation from the spermatogonial stage to the meiotic prophase in spermatogenesis, but not in meiotic divisions per se.
Abstract: We investigated the distribution of haptoglobin (HP) alleles and haplotypes among Africans (Ghanaians), Europeans (from South Africa), and Chinese. HP*1F was present only in Africans and Europeans, whereas HP*del was unique to Chinese. Six base substitutions at the promoter region were population specific. Only 3 out of 18 haplotypes were shared among the populations. A probable application of HP in human population genetics appears legitimate.
Abstract: Myc is a ubiquitous mediator of cell proliferation that transactivates the expression of various genes through E-box sites. Here we report a novel gene, mimitin (Myc-induced mitochondrial protein), that encodes a mitochondrial protein with a molecular mass of 20 kDa. We demonstrated that the transcription of mimitin is directly stimulated by c-Myc. To investigate the role of Mimitin, its expression was suppressed by the RNA interference (RNAi) technique. Whereas specific inhibition of mimitin expression did not affect cell proliferation in human cervical carcinoma, colon adenocarcinoma, and hepatocarcinoma cell lines, it did suppress cell proliferation in human glioblastoma, esophageal squamous cell carcinoma (ESCC), and embryonic lung fibroblastic cells, with the greatest suppression efficiency in ESCC cells. To investigate whether mimitin is related to tumorigenesis in ESCC in vivo, the expression of Mimitin protein in ESCC tissues was studied. Mimitin was highly expressed in 80% (28 of 35) of ESCC tumors, suggesting that high expression of Mimitin is a characteristic feature of ESCC. The expression level of Mimitin was found to be correlated with that of c-Myc and cell proliferation, but not with the histopathological grade, stage of cancer, or age of patients. Taken together, these results suggest that the novel gene mimitin is a direct transcriptional target of c-Myc, and is involved in Myc-dependent cell proliferation at least in ESCC cells.
Abstract: Mina53 is a novel Myc target gene that we previously demonstrated to be involved in cell proliferation. We studied, here, the expression of Mina53 in colon cancer to examine its possible role in carcinogenesis. We generated a specific monoclonal anti-human Mina53 antibody and found that colon tumor cell lines expressed Mina53 highly. We also found that expression of Mina53 was elevated in colon tumor tissues by immunoblotting analysis. Tissue sections of 23 surgical cases of adenocarcinoma and 1 case of adenoma were stained immunohistochemically, and the expression of Mina53 was found to be elevated in all of the adenocarcinomas compared to adjacent nonneoplastic tissues, which showed little staining. Deeply invading tumors as well as tumors that have invaded lymphatic vessels showed strong immunoreactivity against anti-Mina53 antibody. Mina53 was expressed in all pathological grades of cancer as well as in the adenoma. Staining patterns of Ki-67, a biomarker for cell proliferation, were similar to those of Mina53 in most cases, but the percentage of tumor cells stained by anti-Mina53 was higher. Although anti-Ki-67 antibody strongly stained some well-proliferating nonneoplastic cells including cells in the deeper part of the crypts and in lymphoid germinal centers, antibody to Mina53 rarely stained those cells. Suppression of mina53 expression severely suppressed proliferation of colon tumor cells in vitro. Together, our results indicate that the elevated expression of Mina53 is a characteristic feature in colon cancer, one that may have therapeutic applications.
Abstract: PURPOSE: We previously identified mina53, a novel Myc target gene. Here we investigated whether mina53 is related to esophageal squamous cell carcinoma (ESCC), a disease with poor prognosis. EXPERIMENTAL DESIGN: Mina53 expression was suppressed in ESCC cell lines by a RNA interference method to investigate whether Mina53 is involved in cell proliferation. Expression of Mina53 was investigated by Western blotting in tissue sections from patients with ESCC. Immunohistochemical analysis of Mina53 was carried out and compared with that using anti-Ki-67 antibody. Finally, the level of Mina53 expression was compared with the length of survival of patients with ESCC. RESULTS: Reduction of mina53 expression by RNA interference suppressed cell proliferation in ESCC cell lines. Western blot analysis of surgically resected ESCC specimens indicated that the expression of Mina53 in tumors was increased compared with that in adjacent nonneoplastic tissues in all four specimens examined. When formalin-fixed specimens from 52 patients with ESCC were stained immunohistochemically, it was found that Mina53 was highly expressed in 83% of specimens. Anti-Mina53 antibody stained tumors more efficiently than antibody against Ki-67, a cell proliferation biomarker, in some cancer specimens. Patients with high expression of Mina53 had shorter survival periods, whereas the expression level of Ki-67 in ESCC showed no relationship to patient outcome. CONCLUSIONS: Taken together, our results indicate that expression of Mina53 is a characteristic feature of ESCC and suggest that immunostaining by anti-Mina53 antibody may be useful as a potential prognostic indicator.
Abstract: We have identified a novel base substitution at codon 247 in the beta-chain of the haptoglobin 2 ( Hp(2)) allele in a Ghanaian with the Hp0 (ahaptoglobinemic) phenotype. The heterozygous T-->C substitution caused reduced expression of the protein when the mutant was transfected into COS7 cells. The base substitution resulted in a missense change of the non-polar amino acid isoleucine to the polar amino acid threonine at a position in the beta-chain that is highly conserved among several species. We had previously identified a mutation in the Hp gene promoter region for the same individual, which gives her genotype as -61C Hp(2)/-61C Hp(2)(I247T). Since the -61C mutation also leads to low Hp expression, the genotype represents the first and most definitive ahaptoglobinemic case reported in Africa.
Abstract: We have investigated the genetic basis for the Hp0 phenotype amongst 123 randomly selected Ghanaians. A total of 17 individuals were determined to be Hp0 phenotype, based on the classical method for Hp phenotyping of Hb-supplemented plasma. Out of the 17 Hp0 individuals, nine subjects were further classified as ahaptoglobinaemic and eight as hypohaptoglobinaemic by Western blots and double immunodiffusion. We identified three previously known base substitutions (A-55G, A-61C and T-104A) and three new ones (C-101G, T-191G and C-242T) within the 5' flanking region of the Hp gene. The A-61C base substitution significantly decreased transcriptional activity and was associated strongly with Hp2 allele and ahaptoglobinaemia. The C-101G substitution was similar in transcriptional activity to the wild-type and was associated with Hp1S allele and hypohaptoglobinaemia. The Hpdel allele seen in Asian populations was absent. We conclude that the Hp0 phenotype in Ghana has a genetic basis that differs significantly from that seen in Asia.
Abstract: Myc is a ubiquitous mediator of cell proliferation and can transactivate the expression of various genes through E-box sites. Here we report a novel gene, mina53 (Myc-induced nuclear antigen with a molecular mass of 53 kDa). The mina53 gene encodes a protein with a molecular weight of 53 kDa, which is localized in the nucleus and with part of the protein concentrated in the nucleolus. When serum-starved cells were activated by serum, the level of c-myc mRNA was elevated, and an increase in mina53 mRNA followed the elevation of c-myc mRNA. When expression of c-myc was reduced in human promyelocytic leukemia HL60 cells by phorbol 12-myristate 13-acetate, the expression of mina53 mRNA and protein was reduced. The expression of mina53 mRNA and Mina53 protein was induced by ectopic introduction of wild type c-Myc but not by a mutant c-Myc lacking the transactivation domain. When c-Myc in the c-MycER chimeric protein was activated, mina53 mRNA was increased, even in the presence of an inhibitor for protein synthesis. E-box sites are present in a region proximal to the transcription initiation sites of the mina53 gene. The gene expression from the mina53 promoter was elevated by c-Myc through E-box sites. c-Myc protein bound to the mina53 promoter region in vivo in HL60 cells in the proliferating phase but not after treatment of cells with phorbol 12-myristate 13-acetate. Specific inhibition of mina53 expression by an RNA interference method severely suppressed cell proliferation. Taken together, these results indicate that mina53 is a direct target gene of Myc, suggesting that mina53 is involved in mammalian cell proliferation.
Abstract: The basis for managing diabetes mellitus with aqueous extract of Ocimum canum Sim (Lamiaceae), in Ghana was investigated in diabetic and normoglycemic mice. In the diabetic mice, fasting blood glucose decreased by 60% compared to 10% in control mice after 13 weeks of extract administration. Body weight, serum total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) decreased while serum high-density lipoprotein cholesterol (HDL-C) increased in the extract-treated group. In vitro hydroxyl (OH) and superoxide (O2) radical formation, and lipid peroxidation of isolated human LDL and mouse liver homogenates decreased in extract-treated experimental systems. These findings justify the use of O. canum extract as an antidiabetic folk medicine.
