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Manfredo Quadroni - One of the best experts on this subject based on the ideXlab platform.
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ribonuclease inhibitor 1 regulates erythropoiesis by controlling gata1 translation
Journal of Clinical Investigation, 2018Co-Authors: Vijaykumar Chennupati, Kendle M Maslowski, Nicola Andina, Eric Chiwang Yu, Martina Stilinovic, Diogo F T Veiga, Michel A. Duchosal, Cedric Simillion, Aubry Tardivel, Manfredo QuadroniAbstract:Ribosomal proteins (RP) regulate specific gene expression by selectively translating subsets of mRNAs. Indeed, in Diamond-Blackfan anemia and 5q- Syndrome, mutations in RP genes lead to a specific defect in erythroid gene translation and cause anemia. Little is known about the molecular mechanisms of selective mRNA translation and involvement of ribosomal-associated factors in this process. Ribonuclease inhibitor 1 (RNH1) is a ubiquitously expressed protein that binds to and inhibits pancreatic-type ribonucleases. Here, we report that RNH1 binds to ribosomes and regulates erythropoiesis by controlling translation of the erythroid transcription factor GATA1. Rnh1-deficient mice die between embryonic days E8.5 and E10 due to impaired production of mature erythroid cells from progenitor cells. In Rnh1-deficient embryos, mRNA levels of Gata1 are normal, but GATA1 protein levels are decreased. At the molecular level, we found that RNH1 binds to the 40S subunit of ribosomes and facilitates polysome formation on Gata1 mRNA to confer transcript-specific translation. Further, RNH1 knockdown in human CD34+ progenitor cells decreased erythroid differentiation without affecting myelopoiesis. Our results reveal an unsuspected role for RNH1 in the control of GATA1 mRNA translation and erythropoiesis.
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an unexpected role for ribonuclease inhibitor rnh1 in erythropoiesis
Blood, 2014Co-Authors: Ramanjaneyulu Allam, Vijaykumar Chennupati, Kendle M Maslowski, Diogo F T Veiga, Michel A. Duchosal, Manfredo Quadroni, Aubry Tardivel, Robson H Macdonald, Nicolas Fasel, Anne AngelilloscherrerAbstract:Ribonuclease Inhibitor (RNH1) is a ubiquitously expressed leucine-rich repeat protein. The human RNH1 gene evolved via gene duplication and is conserved among mammalian species. RNH1 binds to and inhibits pancreatic type ribonucleases. Further, RNH1 contains numerous cysteine residues whose sulfhydryl groups might play key structural roles and protect from oxidative damage (Dickson et al Prog. Nucleic Acid Res. Mol. Biol 2005). Despite of all these observations, the precise biological role of RNH1in vivo remains unexplored. Here, we describe an essential role for Rnh1 in the regulation of erythropoiesis by controlling erythroid differentiation. To understand the biological function of Rnh1, Rnh1-deficient (Rnh1-/-) mice were generated. Rnh1-/- embryos die between embryonic days E8.5 to E10 due to severe decrease in erythroid cells. Similar percentages of c-Kit+CD41+ cells (Hematopoietic stem/progenitor cells) were present in Rnh1-/- yolk sacs compared to control genotypes, however differentiation of mature erythroid cells was impaired. Rnh1 is expressed in erythroid cells and its expression coincides with the site of primitive erythropoiesis in the yolk sac. Gene expression studies revealed that levels of hematopoietic transcription factors (TF) in Rnh1-deficient yolk sacs were normal, but their target genes were down-regulated. These results indicate that a post-transcriptional mechanism that affects TF gene function. Supporting this, protein levels of the erythroid transcription factor GATA1 and PPARγ, previously shown to control the proliferation and differentiation of erythroid progenitors, were selectively impaired. Whereas myeloid transcription factors C/EBPa and C/EBPb were not affected in Rnh1-/- embryos, suggesting that Rnh1 deficiency specifically affects the translation of erythroid transcription factors. At the molecular level, using the human erythroid K562 cell line, we show that RNH1 is recruited to the ribosome complex and binds to the ribosomal proteins. RNH1-deficiency decreased polysome formation and conversely its overexpression increased polysome formation. Increased expression of RNH1 also increased globin gene expression in K562 cells. These results suggest that RNH1 associates with ribosomes and regulates the translation of erythroid-specific genes, which are necessary for erythroid differentiation. Furthermore, Rnh1 haploinsufficiency leads to decreased erythropoiesis in the spleen of adult mice. Ribosomal haploinsufficiency in several ribosomal genes is known to impair ribosome function and cause macrocytic anemia in Diamond–Blackfan anemia (DBA), a congenital bone marrow failure Syndrome, and the 5q- Syndrome, a subtype of myelodysplastic Syndrome (Narla et al Int. J. Hematol 2011). Recently it has been shown that ribosomal haploinsufficiency can specifically cause a decrease in GATA1 mRNA translation (Ludwig et al Nature Med 2014). Similar to these ribosomal genes, we demonstrate that Rnh1 associates with ribosomes and its deficiency impairs the translation of Gata1 and other erythroid-specific transcription factors, which leads to arrest in erythroid maturation. Collectively our results unravel the important biological function of Rnh1 in the regulation of erythropoiesis, and point to novel therapeutic targets for disorders of erythropoiesis involving ribosomal defects. Summary Figure:RNH1 is recruited to ribosomal complex and is involved in translation of erythroid specific transcription factors (TF) e.g.GATA1. These TFs are necessary for differentiation of progenitor cells in to erythroid cells. RNH1 deficiency impairs the translation of GATA1 and other erythroid-specific transcription factors, which leads to arrest in erythroid maturation. Disclosures No relevant conflicts of interest to declare.
James S. Wainscoat - One of the best experts on this subject based on the ideXlab platform.
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targeted re sequencing analysis of 25 genes commonly mutated in myeloid disorders in del 5q myelodysplastic Syndromes
Haematologica, 2013Co-Authors: Marta Fernandezmercado, James S. Wainscoat, Aristoteles Giagounidis, Sally Killick, Andrea Pellagatti, Ulrich Germing, Adam Burns, Xabier Agirre, Felipe Prosper, Anna SchuhAbstract:Interstitial deletion of chromosome 5q is the most common chromosomal abnormality in myelodysplastic Syndromes. The catalogue of genes involved in the molecular pathogenesis of myelodysplastic Syndromes is rapidly expanding and next-generation sequencing technology allows detection of these mutations at great depth. Here we describe the design, validation and application of a targeted next-generation sequencing approach to simultaneously screen 25 genes mutated in myeloid malignancies. We used this method alongside single nucleotide polymorphism-array technology to characterize the mutational and cytogenetic profile of 43 cases of early or advanced del(5q) myelodysplastic Syndromes. A total of 29 mutations were detected in our cohort. Overall, 45% of early and 66.7% of advanced cases had at least one mutation. Genes with the highest mutation frequency among advanced cases were TP53 and ASXL1 (25% of patients each). These showed a lower mutation frequency in cases of 5q- Syndrome (4.5% and 13.6%, respectively), suggesting a role in disease progression in del(5q) myelodysplastic Syndromes. Fifty-two percent of mutations identified were in genes involved in epigenetic regulation (ASXL1, TET2, DNMT3A and JAK2). Six mutations had allele frequencies <20%, likely below the detection limit of traditional sequencing methods. Genomic array data showed that cases of advanced del(5q) myelodysplastic Syndrome had a complex background of cytogenetic aberrations, often encompassing genes involved in myeloid disorders. Our study is the first to investigate the molecular pathogenesis of early and advanced del(5q) myelodysplastic Syndromes using next-generation sequencing technology on a large panel of genes frequently mutated in myeloid malignancies, further illuminating the molecular landscape of del(5q) myelodysplastic Syndromes.
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activation of the mtor pathway by the amino acid l leucine in the 5q Syndrome and other ribosomopathies
Advances in biological regulation, 2013Co-Authors: Jacqueline Boultwood, Andrea Pellagatti, Bon Ham Yip, Chaitanya Vuppusetty, James S. WainscoatAbstract:Patients with the 5q- Syndrome and Diamond-Blackfan anemia (DBA) suffer from a severe macrocytic anemia. The 5q- Syndrome and DBA are disorders of aberrant ribosome biogenesis (ribosomopathies) and haploinsufficiency of the ribosomal protein genes RPS14 and RPS19, respectively, underlies the anemia found in these disorders. Erythroblasts obtained from patients with the 5q- Syndrome and DBA show impaired mRNA translation and this defect in translation may represent a potential therapeutic target in these ribosomopathies. There are some indications that the amino acid l-leucine, a translation enhancer, may have some efficacy in this group of disorders. Recent studies have shown that l-leucine treatment of zebrafish and murine models of the 5q- Syndrome and DBA results in a marked improvement in the anemia. l-leucine treatment of RPS14-deficient and RPS19-deficient erythroblasts and erythroblasts from patients with the 5q- Syndrome has been shown to result in an increase in cell proliferation, erythroid differentiation and mRNA translation in culture. l-leucine has been shown to improve hemoglobin levels and transfusion independence in a patient with DBA. l-leucine activates the mTOR (mammalian target of rapamycin) signaling pathway that controls cell growth and mRNA translation. There is evidence to suggest that the promotion of translation via the mTOR pathway by l-leucine is the mechanism that underlies the enhanced erythroid progenitor cell growth and differentiation observed in animal and cellular models of the 5q- Syndrome and DBA treated with this amino acid. These data support the rationale for clinical trials of l-leucine as a therapeutic agent for the 5q- Syndrome and DBA.
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Hematological features of patients with myelodysplastic Syndromes associated with a chromosome 5q deletion.
American journal of hematology, 1995Co-Authors: S Lewis, Jacqueline Boultwood, Veronica J Buckle, David Oscier, Fiona M. Ross, M. Fitchett, Katrina Rack, Gail Abrahamson, James S. WainscoatAbstract:The hematological and clinical features of 26 patients with myelodysplasia and a chromosome 5q deletion in the bone marrow are presented. We have examined the relationship of French-American-British Co-operative Group (FAB) 1982 classification and bone marrow karyotype at diagnosis with patient outcome and the presence or absence of the classical features of the 5q-Syndrome. Those patients classified as refractory anemia (RA) with no additional karyotypic abnormalities have the typical features of the 5q-Syndrome and a good prognosis. None of the patients in this group transformed to acute leukemia during the period of follow-up. Patients with either refractory anemia and excess blasts (RAEB) or additional karyotypic abnormalities show many of the hematologic features of the 5q-Syndrome but do not share the good prognosis. We conclude that the 5q-Syndrome may be best defined as primary MDS of the FAB type RA with a 5q deletion as the sole karyotypic abnormality. This simple definition will distinguish patients with a good prognosis and all the classical features of the 5q-Syndrome.
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allelic loss of irf1 in myelodysplasia and acute myeloid leukemia retention of irf1 on the 5q chromosome in some patients with the 5q Syndrome
Blood, 1993Co-Authors: Jacqueline Boultwood, Carrie Fidler, S Lewis, A Maccarthy, H Sheridan, S Kelly, D Oscier, Veronica J Buckle, James S. WainscoatAbstract:: Acquired interstitial deletions of the long arm of chromosome 5 occur frequently in the myelodysplastic Syndrome (MDS) and acute myeloid leukemia (AML). Recently IRF1, a putative tumor suppressor gene localized to the long arm of chromosome 5, has been shown to be deleted from the 5q- chromosome in a group of patients with MDS and AML. It has been suggested that the loss of IRF1 may be critical to the development of the 5q- Syndrome. We have investigated the allelic loss of IRF1 in a group of 12 patients with MDS and a 5q deletion and 2 patients with AML and a 5q deletion. Gene dosage experiments demonstrated that 12 of 14 patients had loss of one allele of the IRF1 gene but no evidence of homozygous loss and that 2 patients with 5q- Syndrome retained both copies of the gene. The retention of IRF1 on the 5q- chromosome in these two cases has been confirmed by fluorescent in situ hybridization localization using an IRF1 cosmid. Pulsed field gel electrophoresis was used to determine whether there was any evidence for structural rearrangement in the region encompassing the IRF1 gene in these two patients. No aberrant bands were detected with a range of rare cutter enzyme digests. We conclude that IRF1 maps outside the commonly deleted segment of the 5q- chromosome and that loss of IRF1 is not solely responsible for the development of the 5q- Syndrome.
Jacqueline Boultwood - One of the best experts on this subject based on the ideXlab platform.
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association between downregulation of pot1 expression and chromosome 7 deletion response to hypomethylation agent treatment and patient survival in myelodysplastic Syndromes
Blood, 2014Co-Authors: Yue Wei, Amit Verma, Monica Cabrero, Yu Jia, Hong Zheng, Zhihong Fang, Simona Colla, Zach Bohannan, Teresa V Bowman, Jacqueline BoultwoodAbstract:The POT1 gene is located in chromosome 7 and encodes a key component of the shelterin complex, which is essential for the maintenance of telomere and chromosome integrity. Somatic mutations of POT1 have been identified in chronic lymphocytic leukemia, which indicates that POT1 dysfunction is involved in the pathogenesis of hematological neoplasms. At the same time, abnormal telomere shortening has been observed in MDS/AML and a spectrum of bone marrow failure Syndromes. We therefore sought to study the potential role of POT1 in MDS by sequencing the gene and characterizing its expression in primary bone marrow specimens of patients with MDS. We first sequenced all POT1 coding regions that are known to have mutations in CLL. PCR-Sanger sequencing was performed in bone marrow mononuclear cells (BM-MNNC) of a cohort of 30 patients with MDS (15 with RAEB/RAEBT, 11 with RA/RARS/RCMD/MDS-U, 2 with CMML, and 2 with 5q- Syndromes). No genetic mutations in the POT1 gene were detected. This result suggests that genetic alteration of POT1 is rare in MDS. We then evaluated the expression of POT1 using cDNA arrays (n=183) or RT-PCR (n=58) in a cohort of 241 patients with MDS from two centers. The median age of our patients was 71 years (32-95). Diagnoses included RAEB in 108 (45%), 5q- Syndrome in 18 (8%), and other Syndromes (RA, RCMD, and MDS-U) in 115 (47%) cases. In this cohort, 140 (58%) patients were diploid, 22 (9%) had chromosome 7 alterations, 21 (9%) had 5q deletion, and 58 (24%) had other cytogenetic abnormalities. Results indicate that POT1 was underexpressed (less than 50% of the POT1 level in normal controls) in the bone marrow CD34+ hematopoietic progenitor cell population in 138 patients (57%). However, no significant difference was observed between the whole MDS cohort and control BM CD34+ cells from healthy donors (n=25). Further subset analysis based on karyotypes revealed that 81% of patients with chromosome 7 alterations (7- and 7q-) had lower expression of POT1 versus 38% of diploid patients, 35% of 5q patients, and 42% of patients with other cytogenetic alterations (p=0.001). ANOVA testing indicated that expression of POT1 was significantly downregulated (less than 50% of control) only in patients with chromosome 7 alteration (p When we compared the survival of patients with POT1 downregulation to other groups, we observed a strong tendency toward shorter overall survival in patients with POT1 downregulation (median OS of 37 months [95% CI: 21-52] vs 53 months [95%CI: 30-75]; p=0.139). This tendency toward poorer OS was also observed when we excluded cases with chromosome 7 alterations (37 months [95% CI: 17-57] vs 53 months [95%CI: 25-80]; p=0.186). Next, we evaluated the potential impact of POT1 expression on responses to therapies. In the subgroup of patients with available treatment records for analysis (n=58), a total of 42 patients received hypomethylating agents (HMA), and 47% of them achieved responses. When comparing POT1 expression levels to HMA response, we observed significantly lower POT1 expression in HMA non-responders than in responders (U Mann-Whitney test p= 0.028). In a regression model for response to HMA, we also observed that downregulation of POT1 was associated with a poorer response to HMA (OR 4.96 [1.01-24.37]; p=0.049). However, when we introduced chromosome 7 alterations into the model, POT1 expression lost its effect, which suggests that the impact of POT1 on response to HMA is due to its interaction with chromosome 7 alterations. Taken together, the results of this study indicate that the downregulation of POT1 gene expression, which is related to chromosome 7 deletions, may play a role in the pathogenesis and prognosis of MDS, including response to HMA-based therapies. Disclosures No relevant conflicts of interest to declare.
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activation of the mtor pathway by the amino acid l leucine in the 5q Syndrome and other ribosomopathies
Advances in biological regulation, 2013Co-Authors: Jacqueline Boultwood, Andrea Pellagatti, Bon Ham Yip, Chaitanya Vuppusetty, James S. WainscoatAbstract:Patients with the 5q- Syndrome and Diamond-Blackfan anemia (DBA) suffer from a severe macrocytic anemia. The 5q- Syndrome and DBA are disorders of aberrant ribosome biogenesis (ribosomopathies) and haploinsufficiency of the ribosomal protein genes RPS14 and RPS19, respectively, underlies the anemia found in these disorders. Erythroblasts obtained from patients with the 5q- Syndrome and DBA show impaired mRNA translation and this defect in translation may represent a potential therapeutic target in these ribosomopathies. There are some indications that the amino acid l-leucine, a translation enhancer, may have some efficacy in this group of disorders. Recent studies have shown that l-leucine treatment of zebrafish and murine models of the 5q- Syndrome and DBA results in a marked improvement in the anemia. l-leucine treatment of RPS14-deficient and RPS19-deficient erythroblasts and erythroblasts from patients with the 5q- Syndrome has been shown to result in an increase in cell proliferation, erythroid differentiation and mRNA translation in culture. l-leucine has been shown to improve hemoglobin levels and transfusion independence in a patient with DBA. l-leucine activates the mTOR (mammalian target of rapamycin) signaling pathway that controls cell growth and mRNA translation. There is evidence to suggest that the promotion of translation via the mTOR pathway by l-leucine is the mechanism that underlies the enhanced erythroid progenitor cell growth and differentiation observed in animal and cellular models of the 5q- Syndrome and DBA treated with this amino acid. These data support the rationale for clinical trials of l-leucine as a therapeutic agent for the 5q- Syndrome and DBA.
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Genome-wide analysis of copy number changes and loss of heterozygosity in myelodysplastic Syndrome with del(5q) using high-density single nucleotide polymorphism arrays
Haematologica, 2008Co-Authors: Li Wang, Carrie Fidler, Nandita Nadig, Aristoteles Giagounidis, Matteo Giovanni Della Porta, Luca Malcovati, Sally Killick, Norbert Gattermann, Carlo Aul, Jacqueline BoultwoodAbstract:Background We undertook a genome wide single nucleotide polymorphism analysis of a spectrum of patients with myelodysplastic Syndrome del(5q) in order to investigate whether additional genomic abnormalities occur. Single nucleotide polymorphism array analysis has been shown to detect not only gene deletions but also regions of uniparental disomy that can pinpoint particular regions for mutation analysis.Design and Methods We studied 42 cases of myelodysplastic Syndrome with del(5q), comprising 21 patients with 5q- Syndrome and 21 with del(5q) (not 5q- Syndrome), and 45 healthy controls by genome wide single nucleotide polymorphism analysis with the 50K Affymetrix single nucleotide polymorphism arrays.Results The del(5q) was characterized in all cases. The commonly deleted region of the 5q- Syndrome extends between the genes SH3TC2 (proximal boundary) and GLRA1 (distal boundary) and measures 2.9 Mb. Copy number changes in addition to the del(5q) were observed in 10 of 21 patients with del(5q) myelodysplastic Syndrome but in none of the patients with the 5q- Syndrome. A total of 63 regions of uniparental disomy greater than 2 Mb were detected in 40 of 42 patients, dispersed on 18/23 chromosomes. In the 5q- Syndrome group 31 regions of uniparental disomy were identified in 19 of 21 patients, the largest one being 7.6 Mb. All 21 patients with del(5q) myelodysplastic Syndrome had uniparental disomy; in total 32 regions of uniparental disomy were identified in the 21 patients, including six regions of uniparental disomy > 10 Mb. Eight recurrent regions of uniparental disomy were observed among the 42 patients. For eight patients we had T-cell DNA as a germline control and four recurrent regions of uniparental disomy were identified that were present only in the neutrophil and not T-cell DNA. One small region of uniparental disomy at 10p12.31-p12.2 was observed in four patients with the 5q- Syndrome.Conclusions This study shows that regions of uniparental disomy greater than 2 Mb are found in the 5q-Syndrome and del(5q) myelodysplastic Syndrome, although large regions of uniparental disomy (>10 Mb) are only found in the latter group. The recurrent regions of uniparental disomy may indicate the position of novel leukemia-associated genes.
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Hematological features of patients with myelodysplastic Syndromes associated with a chromosome 5q deletion.
American journal of hematology, 1995Co-Authors: S Lewis, Jacqueline Boultwood, Veronica J Buckle, David Oscier, Fiona M. Ross, M. Fitchett, Katrina Rack, Gail Abrahamson, James S. WainscoatAbstract:The hematological and clinical features of 26 patients with myelodysplasia and a chromosome 5q deletion in the bone marrow are presented. We have examined the relationship of French-American-British Co-operative Group (FAB) 1982 classification and bone marrow karyotype at diagnosis with patient outcome and the presence or absence of the classical features of the 5q-Syndrome. Those patients classified as refractory anemia (RA) with no additional karyotypic abnormalities have the typical features of the 5q-Syndrome and a good prognosis. None of the patients in this group transformed to acute leukemia during the period of follow-up. Patients with either refractory anemia and excess blasts (RAEB) or additional karyotypic abnormalities show many of the hematologic features of the 5q-Syndrome but do not share the good prognosis. We conclude that the 5q-Syndrome may be best defined as primary MDS of the FAB type RA with a 5q deletion as the sole karyotypic abnormality. This simple definition will distinguish patients with a good prognosis and all the classical features of the 5q-Syndrome.
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allelic loss of irf1 in myelodysplasia and acute myeloid leukemia retention of irf1 on the 5q chromosome in some patients with the 5q Syndrome
Blood, 1993Co-Authors: Jacqueline Boultwood, Carrie Fidler, S Lewis, A Maccarthy, H Sheridan, S Kelly, D Oscier, Veronica J Buckle, James S. WainscoatAbstract:: Acquired interstitial deletions of the long arm of chromosome 5 occur frequently in the myelodysplastic Syndrome (MDS) and acute myeloid leukemia (AML). Recently IRF1, a putative tumor suppressor gene localized to the long arm of chromosome 5, has been shown to be deleted from the 5q- chromosome in a group of patients with MDS and AML. It has been suggested that the loss of IRF1 may be critical to the development of the 5q- Syndrome. We have investigated the allelic loss of IRF1 in a group of 12 patients with MDS and a 5q deletion and 2 patients with AML and a 5q deletion. Gene dosage experiments demonstrated that 12 of 14 patients had loss of one allele of the IRF1 gene but no evidence of homozygous loss and that 2 patients with 5q- Syndrome retained both copies of the gene. The retention of IRF1 on the 5q- chromosome in these two cases has been confirmed by fluorescent in situ hybridization localization using an IRF1 cosmid. Pulsed field gel electrophoresis was used to determine whether there was any evidence for structural rearrangement in the region encompassing the IRF1 gene in these two patients. No aberrant bands were detected with a range of rare cutter enzyme digests. We conclude that IRF1 maps outside the commonly deleted segment of the 5q- chromosome and that loss of IRF1 is not solely responsible for the development of the 5q- Syndrome.
Aubry Tardivel - One of the best experts on this subject based on the ideXlab platform.
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2030 ribonuclease inhibitor rnh1 regulates hematopoietic cell specific translation
Experimental Hematology, 2019Co-Authors: Martina Stilinovic, Nicola Andina, Aubry Tardivel, Irene Keller, Ramanjaneyulu AllamAbstract:Regulation of gene expression is important for normal development and is mainly controlled at the level of transcription. However, recent studies show that ribosomal proteins (RPs) regulate specific gene expression by selectively facilitating translation of specific mRNAs. Indeed, in Diamond- Blackfan anemia (DBA) and 5q– Syndrome, mutations in RP genes lead to a specific defect in erythroid gene translation and cause anemia. How mutations in RP genes leads to hematopoietic specific defects is largely unknown. Similar to transcription factors the existence of cell type specific translation regulators remain elusive. Here, we report that Ribonuclease inhibitor (RNH1) regulates hematopoietic cell specific translation. Recently, we published that RNH1 is a ribosomal associated protein and regulates erythropoiesis by regulating GATA1 mRNA translation. In this study, we found that RNH1-deficiency in human hematopoietic origin cells such as erythroid leukemia cells, monocytic cells and T lymphocytes decreased polysome formation but not in non-hematopoietic origin cells such as HEK293, HaCat, HeLa cells. Similarly, OP-Puro incorporation experiments in mice revealed that RNH1-deficency leads to translation defect in hematopoietic cells but not in non- hematopoietic cells. At molecular level, we found that RNH1 binds to ribosomes and regulates RPs gene expression at translation level independent of mTOR signaling. Interestingly, it has been shown that RNH1 expression is translationally down regulated in RPS19 knockdown cells, which is frequently mutated in DBA patients. Supporting RNH1 role in translation, over expression of RNH1 rescues erythroid and translation defects in RPS19 knockdown cells. Collectively, our result unravels the existence of cell type specific translation regulators and may partially explain cell type specific defects caused by mutations in RP genes.
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ribonuclease inhibitor 1 regulates erythropoiesis by controlling gata1 translation
Journal of Clinical Investigation, 2018Co-Authors: Vijaykumar Chennupati, Kendle M Maslowski, Nicola Andina, Eric Chiwang Yu, Martina Stilinovic, Diogo F T Veiga, Michel A. Duchosal, Cedric Simillion, Aubry Tardivel, Manfredo QuadroniAbstract:Ribosomal proteins (RP) regulate specific gene expression by selectively translating subsets of mRNAs. Indeed, in Diamond-Blackfan anemia and 5q- Syndrome, mutations in RP genes lead to a specific defect in erythroid gene translation and cause anemia. Little is known about the molecular mechanisms of selective mRNA translation and involvement of ribosomal-associated factors in this process. Ribonuclease inhibitor 1 (RNH1) is a ubiquitously expressed protein that binds to and inhibits pancreatic-type ribonucleases. Here, we report that RNH1 binds to ribosomes and regulates erythropoiesis by controlling translation of the erythroid transcription factor GATA1. Rnh1-deficient mice die between embryonic days E8.5 and E10 due to impaired production of mature erythroid cells from progenitor cells. In Rnh1-deficient embryos, mRNA levels of Gata1 are normal, but GATA1 protein levels are decreased. At the molecular level, we found that RNH1 binds to the 40S subunit of ribosomes and facilitates polysome formation on Gata1 mRNA to confer transcript-specific translation. Further, RNH1 knockdown in human CD34+ progenitor cells decreased erythroid differentiation without affecting myelopoiesis. Our results reveal an unsuspected role for RNH1 in the control of GATA1 mRNA translation and erythropoiesis.
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an unexpected role for ribonuclease inhibitor rnh1 in erythropoiesis
Blood, 2014Co-Authors: Ramanjaneyulu Allam, Vijaykumar Chennupati, Kendle M Maslowski, Diogo F T Veiga, Michel A. Duchosal, Manfredo Quadroni, Aubry Tardivel, Robson H Macdonald, Nicolas Fasel, Anne AngelilloscherrerAbstract:Ribonuclease Inhibitor (RNH1) is a ubiquitously expressed leucine-rich repeat protein. The human RNH1 gene evolved via gene duplication and is conserved among mammalian species. RNH1 binds to and inhibits pancreatic type ribonucleases. Further, RNH1 contains numerous cysteine residues whose sulfhydryl groups might play key structural roles and protect from oxidative damage (Dickson et al Prog. Nucleic Acid Res. Mol. Biol 2005). Despite of all these observations, the precise biological role of RNH1in vivo remains unexplored. Here, we describe an essential role for Rnh1 in the regulation of erythropoiesis by controlling erythroid differentiation. To understand the biological function of Rnh1, Rnh1-deficient (Rnh1-/-) mice were generated. Rnh1-/- embryos die between embryonic days E8.5 to E10 due to severe decrease in erythroid cells. Similar percentages of c-Kit+CD41+ cells (Hematopoietic stem/progenitor cells) were present in Rnh1-/- yolk sacs compared to control genotypes, however differentiation of mature erythroid cells was impaired. Rnh1 is expressed in erythroid cells and its expression coincides with the site of primitive erythropoiesis in the yolk sac. Gene expression studies revealed that levels of hematopoietic transcription factors (TF) in Rnh1-deficient yolk sacs were normal, but their target genes were down-regulated. These results indicate that a post-transcriptional mechanism that affects TF gene function. Supporting this, protein levels of the erythroid transcription factor GATA1 and PPARγ, previously shown to control the proliferation and differentiation of erythroid progenitors, were selectively impaired. Whereas myeloid transcription factors C/EBPa and C/EBPb were not affected in Rnh1-/- embryos, suggesting that Rnh1 deficiency specifically affects the translation of erythroid transcription factors. At the molecular level, using the human erythroid K562 cell line, we show that RNH1 is recruited to the ribosome complex and binds to the ribosomal proteins. RNH1-deficiency decreased polysome formation and conversely its overexpression increased polysome formation. Increased expression of RNH1 also increased globin gene expression in K562 cells. These results suggest that RNH1 associates with ribosomes and regulates the translation of erythroid-specific genes, which are necessary for erythroid differentiation. Furthermore, Rnh1 haploinsufficiency leads to decreased erythropoiesis in the spleen of adult mice. Ribosomal haploinsufficiency in several ribosomal genes is known to impair ribosome function and cause macrocytic anemia in Diamond–Blackfan anemia (DBA), a congenital bone marrow failure Syndrome, and the 5q- Syndrome, a subtype of myelodysplastic Syndrome (Narla et al Int. J. Hematol 2011). Recently it has been shown that ribosomal haploinsufficiency can specifically cause a decrease in GATA1 mRNA translation (Ludwig et al Nature Med 2014). Similar to these ribosomal genes, we demonstrate that Rnh1 associates with ribosomes and its deficiency impairs the translation of Gata1 and other erythroid-specific transcription factors, which leads to arrest in erythroid maturation. Collectively our results unravel the important biological function of Rnh1 in the regulation of erythropoiesis, and point to novel therapeutic targets for disorders of erythropoiesis involving ribosomal defects. Summary Figure:RNH1 is recruited to ribosomal complex and is involved in translation of erythroid specific transcription factors (TF) e.g.GATA1. These TFs are necessary for differentiation of progenitor cells in to erythroid cells. RNH1 deficiency impairs the translation of GATA1 and other erythroid-specific transcription factors, which leads to arrest in erythroid maturation. Disclosures No relevant conflicts of interest to declare.
Martina Stilinovic - One of the best experts on this subject based on the ideXlab platform.
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2030 ribonuclease inhibitor rnh1 regulates hematopoietic cell specific translation
Experimental Hematology, 2019Co-Authors: Martina Stilinovic, Nicola Andina, Aubry Tardivel, Irene Keller, Ramanjaneyulu AllamAbstract:Regulation of gene expression is important for normal development and is mainly controlled at the level of transcription. However, recent studies show that ribosomal proteins (RPs) regulate specific gene expression by selectively facilitating translation of specific mRNAs. Indeed, in Diamond- Blackfan anemia (DBA) and 5q– Syndrome, mutations in RP genes lead to a specific defect in erythroid gene translation and cause anemia. How mutations in RP genes leads to hematopoietic specific defects is largely unknown. Similar to transcription factors the existence of cell type specific translation regulators remain elusive. Here, we report that Ribonuclease inhibitor (RNH1) regulates hematopoietic cell specific translation. Recently, we published that RNH1 is a ribosomal associated protein and regulates erythropoiesis by regulating GATA1 mRNA translation. In this study, we found that RNH1-deficiency in human hematopoietic origin cells such as erythroid leukemia cells, monocytic cells and T lymphocytes decreased polysome formation but not in non-hematopoietic origin cells such as HEK293, HaCat, HeLa cells. Similarly, OP-Puro incorporation experiments in mice revealed that RNH1-deficency leads to translation defect in hematopoietic cells but not in non- hematopoietic cells. At molecular level, we found that RNH1 binds to ribosomes and regulates RPs gene expression at translation level independent of mTOR signaling. Interestingly, it has been shown that RNH1 expression is translationally down regulated in RPS19 knockdown cells, which is frequently mutated in DBA patients. Supporting RNH1 role in translation, over expression of RNH1 rescues erythroid and translation defects in RPS19 knockdown cells. Collectively, our result unravels the existence of cell type specific translation regulators and may partially explain cell type specific defects caused by mutations in RP genes.
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ribonuclease inhibitor 1 regulates erythropoiesis by controlling gata1 translation
Journal of Clinical Investigation, 2018Co-Authors: Vijaykumar Chennupati, Kendle M Maslowski, Nicola Andina, Eric Chiwang Yu, Martina Stilinovic, Diogo F T Veiga, Michel A. Duchosal, Cedric Simillion, Aubry Tardivel, Manfredo QuadroniAbstract:Ribosomal proteins (RP) regulate specific gene expression by selectively translating subsets of mRNAs. Indeed, in Diamond-Blackfan anemia and 5q- Syndrome, mutations in RP genes lead to a specific defect in erythroid gene translation and cause anemia. Little is known about the molecular mechanisms of selective mRNA translation and involvement of ribosomal-associated factors in this process. Ribonuclease inhibitor 1 (RNH1) is a ubiquitously expressed protein that binds to and inhibits pancreatic-type ribonucleases. Here, we report that RNH1 binds to ribosomes and regulates erythropoiesis by controlling translation of the erythroid transcription factor GATA1. Rnh1-deficient mice die between embryonic days E8.5 and E10 due to impaired production of mature erythroid cells from progenitor cells. In Rnh1-deficient embryos, mRNA levels of Gata1 are normal, but GATA1 protein levels are decreased. At the molecular level, we found that RNH1 binds to the 40S subunit of ribosomes and facilitates polysome formation on Gata1 mRNA to confer transcript-specific translation. Further, RNH1 knockdown in human CD34+ progenitor cells decreased erythroid differentiation without affecting myelopoiesis. Our results reveal an unsuspected role for RNH1 in the control of GATA1 mRNA translation and erythropoiesis.
