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Akiko Shimamura - One of the best experts on this subject based on the ideXlab platform.
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Clinical spectrum and molecular pathophysiology of Shwachman-Diamond syndrome.
Current Opinion in Hematology, 2020Co-Authors: James N. Huang, Akiko ShimamuraAbstract:PURPOSE OF REVIEW Shwachman Diamond syndrome (SDS) is an inherited bone marrow failure and cancer predisposition syndrome that affects multiple organ systems. Mutations in the SBDS gene are found in the majority of patients, but the molecular function of the SBDS protein product remains unclear. Here, we review recent progress in the clinical and molecular characterization of SDS.
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Impaired ribosomal subunit association in Shwachman-Diamond syndrome
Blood, 2012Co-Authors: Nicholas Burwick, Tomoka Nakamura, Scott A Coats, Akiko ShimamuraAbstract:Shwachman-Diamond syndrome (SDS) is an autosomal-recessive marrow failure syndrome with a predisposition to leukemia. SDS patients harbor biallelic mutations in the SBDS gene, resulting in low levels of SBDS protein. Data from nonhuman models demonstrate that the SBDS protein facilitates the release of eIF6, a factor that prevents ribosome joining. The complete abrogation of SBDS expression in these models results in severe cellular and lethal physiologic abnormalities that differ from the human disease phenotype. Because human SDS cells are characterized by partial rather than complete loss of SBDS expression, we interrogated SDS patient cells for defects in ribosomal assembly. SDS patient cells exhibit altered ribosomal profiles and impaired association of the 40S and 60S subunits. Introduction of a wild-type SBDS cDNA into SDS patient cells corrected the ribosomal association defect, while patient-derived SBDS point mutants only partially improved subunit association. Knockdown of eIF6 expression improved ribosomal subunit association but did not correct the hematopoietic defect of SBDS-deficient cells. In summary, we demonstrate an SBDS-dependent ribosome maturation defect in SDS patient cells. The role of ribosomal subunit joining in marrow failure warrants further investigation.
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clinical features of shwachman diamond syndrome patients lacking biallelic SBDS mutation
Blood, 2011Co-Authors: Akiko Shimamura, Audrey Anna Bolyard, Joan Moore, Satabdi Chakrabarti, Jordan M Bond, Theresa Cole, Laurence A Boxer, Peter E Newburger, Blanche P Alter, Richard E. HarrisAbstract:Abstract 4367 Shwachman-Diamond syndrome (SDS) is an autosomal recessively inherited disorder defined clinically by marrow failure and exocrine pancreatic dysfunction. Previous research estimates that 90% of patients harbor biallelic mutations in the SBDS gene. The clinical course of patients lacking SBDS mutations has not been examined previously. To address this question, we examined 102 patients referred to the North American Shwachman-Diamond syndrome registry (SDSR) or the Severe Chronic Neutropenia International Registry (SCNIR). Seventy-nine subjects were 18 years of age (median 23.6, range 18.2–61.9), with a male:female ratio of 1.6:1. SBDS genetic analyses were available for 75 patients; 48 of 75 have biallelic SBDS mutations. Twenty-seven patients were phenotypically consistent with SDS, as demonstrated by exocrine pancreatic dysfunction and marrow failure, but either lacked SBDS mutations (24 patients) or harbored only one mutant SBDS allele (3 patients). The remaining 27 subjects were indeterminate for SDS or lacked sufficient data. In this study, we compared the hematologic complications of the 75 SDS patients presenting with or without SBDS mutations. Fifty-two subjects had complete hematological data. Of the 28 patients with SBDS mutations, neutropenia was noted in 21 (6 severe with ANC Bone marrow reports were available for 47 patients. Of the 21 patients with SBDS mutations, 17 had marrow hypoplasia, 5 had marrow dysplasia. Nine showed clonal abnormalities including del(20q), iso(7q), monosomy 7, trisomy 8, and trisomy 7q21. Of the 26 patients without biallelic mutations, 16 had marrow hypoplasia, 3 had marrow dysplasia and 8 showed clonal abnormalities including del(20q), monosomy 7, del(3q), del (21q), del (7q), and iso(7q). Three subjects (2 with biallelic SBDS mutations, 1 lacking SBDS mutations) developed MDS. One patient without SBDS mutations developed AML. Three deaths have been reported. All deaths involved patients with clinical SDS lacking SBDS mutations. Causes of death were AML, failure to engraft during bone marrow transplant, and sepsis. Eight patients (4 with biallelic SBDS mutations and 4 lacking SBDS mutations) underwent bone marrow transplant. These data suggest that patients without mutations in SBDS may be more common than previous estimates. The hematological complications for patients with and without SBDS mutations appear to be similar. These Registries offer important opportunities to study the genetic and pathophysiological mechanisms for SDS. Disclosures: Boxer:Amgen: Equity Ownership. Dale:Amgen: Consultancy, Honoraria, Membership on an entity’s Board of Directors or advisory committees, Research Funding.
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SBDS and eif6 modulate ribosome subunit joining in shwachman diamond syndrome
Blood, 2011Co-Authors: Nicholas Burwick, Scott A Coats, Akiko ShimamuraAbstract:Abstract 3438 Shwachman Diamond syndrome (SDS) is an autosomal recessive marrow failure syndrome with a predisposition to leukemia. Over 90% of SDS patients harbor biallelic mutations in the SBDS gene. SBDS has been implicated in several cellular functions including ribosome biogenesis and mitotic spindle stabilization. Deletion of SBDS orthologues in yeast results in a severe slow growth phenotype and depressed polysomes. Homozygous deletion of SBDS in murine models results in early embryonic lethality, while conditional deletion of SBDS in mouse liver demonstrates accumulation of 40S and 60S subunits and halfmer formation consistent with impaired ribosome joining. SBDS facilitates the release of eIF6, a factor that prevents ribosome joining. The dramatic phenotypic and polysome changes noted in these experimental models were not observed in cells derived from SDS patients. SDS patient cells have only a mildly reduced growth rate compared to heatlhy controls, and polysome profiles do not demonstrate depressed polysomes or halfmer formation. Since complete abrogation of SBDS expression is lethal and biallelic null mutations in SBDS have not been reported, we examined the role of SBDS and eIF6 in SDS patients and human cell models. We first investigated whether ribosome subunit homeostasis is impaired in SDS patient cells. We find that the 60S:40S ribosomal subunit ratio is consistently reduced in bone marrow stromal cells from SDS patients of different genotypes (n=4). This impairment in 60S:40S ratio is demonstrated in both SDS patient stromal cells and patient lymphoblasts. Stable lentiviral knockdown of SDS in normal marrow stromal cells recapitulates the reduction in 60S:40S ratio. SBDS and eIF6 co-sediment in polysome gradients of human SDS cells. This co-sedimentation is specific for the 60S ribosomal subunit. Since eIF6 has a role as an anti-joining factor, we next developed an in vitro assay to test for ribosome subunit joining in human cells. In this assay, we validate that over-expression of eIF6 results in reduced ribosome joining, and eIF6 knockdown promotes ribosome joining. Moreover, we find that SDS patient stromal cells and patient lymphoblasts both demonstrate impaired ribosome subunit joining, compared with healthy controls. Importantly, the addition of wild type SBDS or depletion of eIF6 improve ribosome joining in SDS patient cells. We demonstrate that the amino terminal sequences of SBDS are necessary but not sufficient for the association of SBDS with the 60S ribosomal subunit. Insertion of a patient-derived N-terminal SBDS point mutation also results in decreased association of SBDS with the 60S ribosomal subunit. These structure-function studies may help to inform genotype:phenotype correlations in SDS. The role of defective ribosome joining in promoting the SDS hematopoietic phenotype is of particular interest. Ongoing studies are interrogating the role of eIF6 modulation on the hematopoietic phenotype in SBDS- depleted cells. Insights garnered from these experiments will help inform the development of novel agents to improve the hematopoetic defect in human SDS. Disclosures: No relevant conflicts of interest to declare.
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SBDS protein expression patterns in the bone marrow
Pediatric Blood & Cancer, 2010Co-Authors: Akiko Shimamura, Trisha E Wong, Monica L Calicchio, Mark D Fleming, Marian H HarrisAbstract:Shwachman–Diamond syndrome (SDS) is an inherited bone marrow failure syndrome caused by biallelic SBDS gene mutations. Here we examined SBDS protein levels in human bone marrow. SBDS protein expression was high in neutrophil progenitors, megakaryocytes, plasma cells, and osteoblasts. In contrast, SBDS protein levels were low in all hematopoietic cell lineages from patients harboring the common SBDS mutations. We conclude that SBDS protein levels vary widely between specific marrow lineages. Uniformly low SBDS protein expression levels distinguish the majority of SDS patients from controls or other marrow failure syndromes. Pediatr Blood Cancer. 2010;55:546–549. © 2010 Wiley-Liss, Inc.
Johanna M Rommens - One of the best experts on this subject based on the ideXlab platform.
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deficiency of the ribosome biogenesis gene SBDS in hematopoietic stem and progenitor cells causes neutropenia in mice by attenuating lineage progression in myelocytes
Haematologica, 2015Co-Authors: Noemi A Zambetti, Johanna M Rommens, Paulina M H Van Strien, Marijke Valkhof, Maria N Adisty, Eric M J Bindels, Remco Hoogenboezem, Mathijs A Sanders, Ivo P Touw, Marc H G P RaaijmakersAbstract:Shwachman-Diamond syndrome is a congenital bone marrow failure disorder characterized by debilitating neutropenia. The disease is associated with loss-of-function mutations in the SBDS gene, implicated in ribosome biogenesis, but the cellular and molecular events driving cell specific phenotypes in ribosomopathies remain poorly defined. Here, we established what is to our knowledge the first mammalian model of neutropenia in Shwachman-Diamond syndrome through targeted downregulation of SBDS in hematopoietic stem and progenitor cells expressing the myeloid transcription factor CCAAT/enhancer binding protein α (Cebpa). SBDS deficiency in the myeloid lineage specifically affected myelocytes and their downstream progeny while, unexpectedly, it was well tolerated by rapidly cycling hematopoietic progenitor cells. Molecular insights provided by massive parallel sequencing supported cellular observations of impaired cell cycle exit and formation of secondary granules associated with the defect of myeloid lineage progression in myelocytes. Mechanistically, SBDS deficiency activated the p53 tumor suppressor pathway and induced apoptosis in these cells. Collectively, the data reveal a previously unanticipated, selective dependency of myelocytes and downstream progeny, but not rapidly cycling progenitors, on this ubiquitous ribosome biogenesis protein, thus providing a cellular basis for the understanding of myeloid lineage biased defects in Shwachman-Diamond syndrome.
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deletion of SBDS from hematopoietic progenitors causes neutropenia in a mouse model of shwachman diamond syndrome by specifically blocking myeloid lineage progression at late differentiation stages
Blood, 2014Co-Authors: Noemi A Zambetti, Johanna M Rommens, Paulina M H Van Strien, Marijke Valkhof, Maria N Adisty, Eric M J Bindels, Remco Hoogenboezem, Mathijs A Sanders, Ivo P Touw, Marc H G P RaaijmakersAbstract:Shwachman-Diamond Syndrome (SDS) is a congenital bone marrow failure disorder characterized by neutropenia and predisposition to leukemia. SDS is associated with loss-of-function mutations in the SBDS gene, involved in ribosome biogenesis, but the cellular and molecular events driving neutropenia in SDS remain poorly defined, largely due to a lack of mammalian disease models recapitulating the hematopoietic features of SDS. To achieve deletion of SBDS in early hematopoietic/myeloid progenitors, we generated Cebpa cre/+ SBDS f/f R26 EYFP/+ mice ( SBDS Δ/Δ), which were non-viable. Analysis of E14.5 embryos demonstrated global conservation of the hematopoietic hierarchy in SBDS Δ/Δ embryos compared to Cebpa cre/+ R26 EYFP/+ controls ( SBDS +/+). Functional relevance of SBDS deletion in Cebpa+ progenitors was tested by transplanting fetal liver cells from E14.5 SBDS Δ/Δ or SBDS +/+ embryos into lethally irradiated B6.SJL mice. Deficiency of SBDS in different EYFP+ hematopoietic subsets was confirmed by qPCR (log2 fold change: LKS -2.25, CMP -9.42, GMP -9.1 and neutrophils -7.29). Mice transplanted with SBDS Δ/Δ cells developed profound neutropenia (log2FC: -2.56; p =0.002, n=7), which was stable during the time of analysis (16 weeks). FACS and morphological studies of bone marrow EYFP+ cells demonstrated increased frequencies of early progenitor populations, recapitulating the left-shifted hematopoiesis observed in human SDS patients (Dror et al. , Ann N Y Acad Sci 2011), with a pronounced accumulation of cKitint Gr1low EYFP+ myelocytes-metamyelocytes (MC-MMs) (frequency of EYFP+ cells: SBDS +/+ 15.3±1.7 %, SBDS Δ/Δ 39.4±1.1 %; p = 8.7x10-8) and a marked decrease in Gr1+ Mac1+ cells ( SBDS +/+: 54.5±7.5 %, SBDS Δ/Δ: 22.3±2.8 %; p = 1.6x10-4), indicating that neutropenia was caused by disrupted lineage progression from MC-MMs to neutrophils. In line with this, whole transcriptome analysis of prospectively isolated EYFP+ MC-MMs (RNA-seq, n=4) revealed enrichment for hematopoietic stem and progenitor cell signatures in SBDS Δ/Δ recipients, while myeloid signatures were enriched in SBDS +/+ mice (GSEA). Transcript analysis further showed reduced expression of granule components produced at the MC-MM stage, such as Ltf , Mmp8 , Mmp9 . As expected, reduced expression of SBDS (FDR=6.6x10-9, log2FC=-2.67) and dysregulation of ribosome proteins (RP) production were observed, with increased expression of over 60 RP genes in SBDS Δ/Δ MC-MMs. To investigate the molecular events underlying impaired lineage progression, we focused on p53 activation, a common mechanism of tissue failure in ribosomopathies. P53 protein was significantly and selectively accumulated in EYFP+ MC-MMs of SBDS Δ/Δ recipients as measured by FACS (Mean Fluorescence Intensity FC: 1.8; p= 0.01), with increased expression of p53 targets such as Cdkn1a (p21), Bbc3 (PUMA) and Bax . This was accompanied by increased apoptosis (annexin V FACS analysis) in both MC-MMs and mature neutrophils (FC: 1.36; p=0.02 and FC=2.44; p=1.6x10-4, respectively). However, pharmacological inhibition of the p53 pathway by pifithrin-α (2 months, i.p.) failed to rescue neutropenia, suggesting that alternative mechanisms, such as disrupted expression of transcription factors, may contribute to neutropenia in SDS. Transcript analysis of SBDS -deficient MC-MMs revealed downregulation of Rara (Retinoic acid receptor α) and its target Cdkn1b (p27), implicated in cell cycle arrest and terminal granulopoiesis (Walkley et al ., Blood 2004). Congruent with this finding, Ki67 staining showed that end-stage granulocytes from SBDS Δ/Δ recipients fail to exit cell cycle. In conclusion, we established a mouse model of neutropenia caused by SBDS deficiency in the hematopoietic system, providing experimental support for a direct causative link between SBDS deficiency and neutropenia in mammals. The data reveal a previously unanticipated, selective dependency of late myeloid cells on this ubiquitous protein, while its deficiency spares the function of rapid cycling hematopoietic progenitors. Mechanistically, disrupted expression transcription factors governing terminal myeloid differentiation may be implicated. We anticipate the mouse model to be a valuable tool in further dissecting the molecular pathogenesis of SDS and enabling preclinical studies for disease modulation. Disclosures No relevant conflicts of interest to declare.
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deficiency of SBDS in the mouse pancreas leads to features of shwachman diamond syndrome with loss of zymogen granules
Gastroenterology, 2012Co-Authors: Marina E Tourlakis, Peter R. Durie, Jian Zhong, Rikesh Gandhi, Siyi Zhang, Lingling Chen, Johanna M RommensAbstract:Background & Aims Shwachman–Diamond syndrome (SDS) is the second leading cause of hereditary exocrine pancreatic dysfunction. More than 90% of patients with SDS have biallelic loss-of-function mutations in the Shwachman–Bodian Diamond syndrome ( SBDS ) gene, which encodes a factor involved in ribosome function. We investigated whether mutations in SBDS lead to similar pancreatic defects in mice. Methods Pancreas-specific knock-out mice were generated using a floxed SBDS allele and bred with mice carrying a null or disease-associated missense SBDS allele. Cre recombinase, regulated by the pancreatic transcription factor 1a promoter, was used to disrupt SBDS specifically in the pancreas. Models were assessed for pancreatic dysfunction and growth impairment. Results Disruption of SBDS in the mouse pancreas was sufficient to recapitulate SDS phenotypes. Pancreata of mice with SBDS mutations had decreased mass, fat infiltration, but general preservation of ductal and endocrine compartments. Pancreatic extracts from mutant mice had defects in formation of the 80S ribosomal complex. The exocrine compartment of mutant mice was hypoplastic and individual acini produced few zymogen granules. The null SBDS allele resulted in an earlier onset of phenotypes as well as endocrine impairment. Mutant mice had reduced serum levels of digestive enzymes and overall growth impairment. Conclusions We developed a mouse model of SDS with pancreatic phenotypes similar to those of the human disease. This model could be used to investigate organ-specific consequences of SBDS -associated ribosomopathy. SBDS genotypes correlated with phenotypes. Defects developed specifically in the pancreata of mice, reducing growth of mice and production of digestive enzymes. SBDS therefore appears to be required for normal pancreatic development and function.
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Deficiency of SBDS in the Mouse Pancreas Leads to Features of Shwachman–Diamond Syndrome, With Loss of Zymogen Granules
Gastroenterology, 2012Co-Authors: Marina E Tourlakis, Peter R. Durie, Jian Zhong, Rikesh Gandhi, Siyi Zhang, Lingling Chen, Johanna M RommensAbstract:Background & Aims Shwachman–Diamond syndrome (SDS) is the second leading cause of hereditary exocrine pancreatic dysfunction. More than 90% of patients with SDS have biallelic loss-of-function mutations in the Shwachman–Bodian Diamond syndrome ( SBDS ) gene, which encodes a factor involved in ribosome function. We investigated whether mutations in SBDS lead to similar pancreatic defects in mice. Methods Pancreas-specific knock-out mice were generated using a floxed SBDS allele and bred with mice carrying a null or disease-associated missense SBDS allele. Cre recombinase, regulated by the pancreatic transcription factor 1a promoter, was used to disrupt SBDS specifically in the pancreas. Models were assessed for pancreatic dysfunction and growth impairment. Results Disruption of SBDS in the mouse pancreas was sufficient to recapitulate SDS phenotypes. Pancreata of mice with SBDS mutations had decreased mass, fat infiltration, but general preservation of ductal and endocrine compartments. Pancreatic extracts from mutant mice had defects in formation of the 80S ribosomal complex. The exocrine compartment of mutant mice was hypoplastic and individual acini produced few zymogen granules. The null SBDS allele resulted in an earlier onset of phenotypes as well as endocrine impairment. Mutant mice had reduced serum levels of digestive enzymes and overall growth impairment. Conclusions We developed a mouse model of SDS with pancreatic phenotypes similar to those of the human disease. This model could be used to investigate organ-specific consequences of SBDS -associated ribosomopathy. SBDS genotypes correlated with phenotypes. Defects developed specifically in the pancreata of mice, reducing growth of mice and production of digestive enzymes. SBDS therefore appears to be required for normal pancreatic development and function.
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SBDS deficient neutrophils exhibit normal numbers chemotaxis and phagocytic functions but impaired nadph oxidase activity
Blood, 2006Co-Authors: Siyi Zhang, Jian Zhong, Michael Glogauer, Johanna M RommensAbstract:Shwachman-Diamond syndrome (SDS) is a recessive disease with multi-system involvement, and is characterized by hematological dysfunction, exocrine pancreatic insufficiency and skeletal abnormalities. Recurrent infections are a major cause of morbidity for SDS patients, due to neutropenia and proposed defects in neutrophil functions. SDS is caused by mutations in SBDS , a highly conserved gene that has been suggested to be involved in RNA metabolism and/or ribosome biogenesis. A conditional Cre-Lox SBDS allele was generated in the mouse using gene targeting technology, as complete ablation of SBDS has been shown to lead to early embryonic lethality. To investigate neutrophil functions and their specific sensitivity to loss of SBDS, we elected to delete the gene by breeding mice with the conditional allele to mice with Cre recombinase under the control of the lysozyme M promoter. While SBDS was confirmed to be ablated in mature neutrophils in adult offspring mice, their numbers and morphology were found to be normal in peripheral blood. SBDS -ablated neutrophils were also found to have normal chemotatic and chemokinetic abilities, and they demonstrated phagocytic function comparable to neutrophils of control mice. In contrast, NADPH oxidase activity was observed to be greatly reduced in response to phorbol-12-myristate 13-acetate stimulation, while the expression levels and phosphorylation of the NADPH oxidase complex components remained intact. To evaluate the net effect of the loss of SBDS in vivo , clearance of Pseudomonas aeruginosa from infected lungs was investigated, and found to be significantly reduced in mice with SBDS -abated neutrophils compared their wild type littermates. In summary, loss of SBDS does not to affect total peripheral neutrophil numbers directly, or chemotaxis and phagocytic abilities, but was essential for a robust NADPH oxidase response. These findings suggest that neutrophils are compromised with loss of SBDS , but that at least some of the neutrophil deficiencies of SDS patients are likely due to microenvironment deficiencies, or early hematopoiesis abnormalities.
Bronwyn Kerr - One of the best experts on this subject based on the ideXlab platform.
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The Shwachman–Bodian–Diamond syndrome gene mutations cause a neonatal form of spondylometaphysial dysplasia (SMD) resembling SMD Sedaghatian type
Journal of Medical Genetics, 2007Co-Authors: Nishimura, Phillip Cox, Y. Miyamoto, Trevor Cole, Yuichiro Hirose, David L. Rimoin, Daniel H. Cohn, Eiji Nakashima, Ralph S Lachman, Bronwyn KerrAbstract:The Shwachman–Bodian–Diamond syndrome (SBDS) gene is a causative gene for Shwachman–Diamond syndrome, an autosomal recessive disorder with exocrine pancreatic insufficiency, bone marrow dysfunction and skeletal dysplasia. We report here on two patients with skeletal manifestations at the severest end of the phenotypic spectrum of SBDS mutations. An 11‐year‐old Japanese girl presented with neonatal respiratory failure necessitating lifelong ventilation support, severe short stature and severe developmental delay. She developed neutropenia in infancy, and decreased serum amylase was noted in childhood. A British boy was a stillbirth with pulmonary hypoplasia and hepatic fibrosis found on autopsy. Both cases had neonatal skeletal manifestations that included platyspondyly, lacy iliac crests and severe metaphysial dysplasia, and thus did not fall in the range of the known Shwachman–Diamond syndrome skeletal phenotype but resembled spondylometaphysial dysplasia (SMD) Sedaghatian type. The girl harboured a recurrent mutation (183TA→CT) and a novel missense mutation (79T→C), whereas the boy carried two recurrent mutations (183TA→CT and 258+2T→C). We also examined SBDS in one typical case with SMD Sedaghantian type and eight additional cases with neonatal SMD, but failed to discover SBDS mutations. Our experience expands the phenotypic spectrum of SBDS mutations, which, at its severest end, results in severe neonatal SMD.
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The Shwachman-Bodian-Diamond syndrome gene mutations cause a neonatal form of spondylometaphysial dysplasia (SMD) resembling SMD Sedaghatian type.
Journal of medical genetics, 2007Co-Authors: Gen Nishimura, Phillip Cox, Y. Miyamoto, Trevor Cole, Yuichiro Hirose, David L. Rimoin, Daniel H. Cohn, Eiji Nakashima, Ralph S Lachman, Bronwyn KerrAbstract:The Shwachman-Bodian-Diamond syndrome (SBDS) gene is a causative gene for Shwachman-Diamond syndrome, an autosomal recessive disorder with exocrine pancreatic insufficiency, bone marrow dysfunction and skeletal dysplasia. We report here on two patients with skeletal manifestations at the severest end of the phenotypic spectrum of SBDS mutations. An 11-year-old Japanese girl presented with neonatal respiratory failure necessitating lifelong ventilation support, severe short stature and severe developmental delay. She developed neutropenia in infancy, and decreased serum amylase was noted in childhood. A British boy was a stillbirth with pulmonary hypoplasia and hepatic fibrosis found on autopsy. Both cases had neonatal skeletal manifestations that included platyspondyly, lacy iliac crests and severe metaphysial dysplasia, and thus did not fall in the range of the known Shwachman-Diamond syndrome skeletal phenotype but resembled spondylometaphysial dysplasia (SMD) Sedaghatian type. The girl harboured a recurrent mutation (183TA-->CT) and a novel missense mutation (79T-->C), whereas the boy carried two recurrent mutations (183TA-->CT and 258+2T-->C). We also examined SBDS in one typical case with SMD Sedaghantian type and eight additional cases with neonatal SMD, but failed to discover SBDS mutations. Our experience expands the phenotypic spectrum of SBDS mutations, which, at its severest end, results in severe neonatal SMD.
Yigal Dror - One of the best experts on this subject based on the ideXlab platform.
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the ribosome related protein SBDS is critical for normal erythropoiesis
Blood, 2011Co-Authors: Hanming Wang, Kim Zhou, Chi Lan Nghiem, Chetankumar S Tailor, Meredith S Irwin, Yigal DrorAbstract:Although anemia is common in Shwachman- Diamond syndrome (SDS), the underlying mechanism remains unclear. We asked whether SBDS , which is mutated in most SDS patients, is critical for erythroid development. We found that SBDS expression is high early during erythroid differentiation. Inhibition of SBDS in CD34 + hematopoietic stem cells and early progenitors (HSC/Ps) and K562 cells led to slow cell expansion during erythroid differentiation. Induction of erythroid differentiation resulted in markedly accelerated apoptosis in the knockdown cells; however, proliferation was only mildly reduced. The percentage of cells entering differentiation was not reduced. Differentiation also increased the oxidative stress in SBDS -knockdown K562 cells, and antioxidants enhanced the expansion capability of differentiating SBDS -knockdown K562 cells and colony production of SDS patient HSC/Ps. Erythroid differentiation also resulted in reduction of all ribosomal subunits and global translation. Furthermore, stimulation of global translation with leucine improved the erythroid cell expansion of SBDS -knockdown cells and colony production of SDS patient HSC/Ps. Leucine did not reduce the oxidative stress in SBDS-deficient K562 cells. These results demonstrate that SBDS is critical for normal erythropoiesis. Erythropoietic failure caused by SBDS deficiency is at least in part related to elevated ROS levels and translation insufficiency because antioxidants and leucine improved cell expansion.
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SBDS deficiency results in deregulation of reactive oxygen species leading to increased cell death and decreased cell growth
Pediatric Blood & Cancer, 2010Co-Authors: Chhaya Ambekar, Herman Yeger, Yigal DrorAbstract:Background Shwachman–Diamond syndrome (SDS) is characterized by reduced hematopoietic and exocrine pancreatic cell numbers and a marked propensity for leukemia. Most patients have mutations in the SBDS gene. We previously reported that SBDS-deficient cells overexpress Fas, undergo accelerated spontaneous and Fas-mediated apoptosis and grow slowly. However the mechanism of how SBDS regulates apoptosis remains unknown. Several studies have shown that reactive oxygen species (ROS) regulate cell growth and spontaneous and Fas-mediated cell death. Therefore, we hypothesized that SBDS-deficiency disrupts ROS regulation and subsequently increases sensitivity to Fas stimulation and reduced cell growth. Procedure SBDS was knocked down in HeLa cervical cancer cells and TF-1 myeloid cells using short hairpin RNA. ROS levels were evaluated by oxidation of 2′,7′-dichlorodihydrofluorescein diacetate. Apoptosis and cell growth were evaluated with and without antioxidants by annexin V/propidium iodide and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays, respectively. Results We found that shRNA mediated SBDS-knockdown resulted in a significant increase in ROS levels compared to control cells. Fas stimulation further increased ROS levels in the SBDS-knockdown HeLa cells more than in the controls. Importantly, balancing ROS levels by antioxidants rescued SBDS-deficient cells from spontaneous and Fas-mediated apoptosis and reduced cell growth. Conclusions ROS levels are increased in SBDS-deficient cells, which leads to increased apoptosis and decreased cell growth. Increased baseline and Fas-mediated ROS levels in SBDS-deficient cells can enhance the sensitivity to Fas stimulation. By balancing ROS levels, antioxidants can improve cell growth and survival in SBDS-deficient cells. Pediatr Blood Cancer. 2010;55:1138–1144. © 2010 Wiley-Liss, Inc.
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mechanisms of erythropoietic failure in shwachman diamond syndrome caused by loss of ribosome related protein SBDS
Blood, 2009Co-Authors: Hanming Wang, Sallylin Adams, Kim Zhou, Yigal DrorAbstract:Abstract 3203 Poster Board III-140 Anemia occurs in 60% of patients with Shwachman Diamond Syndrome (SDS). Although bi-allelic mutations in SBDS cause SDS, it is unclear whether SBDS is critical for erythropoiesis and what the pathogenesis of anemia is in SDS. We hypothesize that SBDS protects early erythroid progenitors from apoptosis by promoting ribosome biosynthesis and translation. During early erythroid differentiation of human K562 cells and primary CD133+ cells, a prominent upregulation of SBDS by RT-qPCR was found. SBDS deficiency by vector-based shRNA led to impaired cell expansion of differentiating K562 cells due to accelerated apoptosis and a mild reduction in proliferation. Furthermore, the cells showed general reduction of 40S, 60S, 80S ribosomal subunits, loss of polysomes and impaired global translation during differentiation. Both cell expansion and translation defects were rescued upon re-introduction of SBDS in K562 cells. Interestingly, leucine partly corrected the cell expansion and translational defects of non-differentiating SBDS-deficient K562 cells, while differentiating SBDS-deficient K562 cells showed improved cell expansion in the presence of additional translation stimulators such as IGF-1. SBDS -knockdown CD133+ cells showed increased BFU-E colony formation under conditions with leucine and a combination of leucine and IGF-1 treatment. Although the erythroid cell expansion defect in K562 cells is independent of p53 as these cells do not express the gene, an upregulation of TAp73, was found in resting SBDS deficient K562 cells. However expression of TAp73 was lost during differentiation. DNp63 was also not upregulated in SBDS-deficient K562 erythroid cells. These results demonstrate that the role of SBDS in non-differentiated cells versus differentiated cells represents two dynamic scenarios and that SBDS plays a critical role in erythroid expansion by promoting survival of early erythroid progenitors and in maintaining ribosome biogenesis during erythroid maturation through a pathway independent of p53 family members. Disclosures No relevant conflicts of interest to declare.
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SBDS deficiency results in specific hypersensitivity to fas stimulation and accumulation of fas at the plasma membrane
Apoptosis, 2009Co-Authors: Kenichiro Watanabe, Hanming Wang, Chhaya Ambekar, Aaron D Schimmer, Amanda Ciccolini, Yigal DrorAbstract:Shwachman–Diamond syndrome (SDS) is an inherited disorder characterized by reduced cellularity in the bone marrow and exocrine pancreas. Most patients have mutations in the SBDS gene, whose functions are unknown. We previously showed that cells deficient in the SBDS protein are characterized by accelerated apoptosis and Fas hypersensitivity, suggesting that the protein might play an important role in Fas-mediated apoptosis. To study the mechanism of Fas hypersensitivity, we compared shRNA-mediated SBDS-knockdown HeLa cells and SDS marrow CD34+ cells for their sensitivity to several groups of apoptosis inducers. Marked hypersensitivity was noticed in response to Fas stimulation, but not to tumor necrosis factor-α, DNA-damaging agents, transcription inhibition or protein synthesis inhibition. To identify the Fas signaling factors that cause hypersensitivity, we analyzed the expression of the pathway’s proteins. We found that Fas accumulated at the plasma membrane in SBDS-knockdown cells with corresponding expression of Fas transcript 1, the main Fas transcript which contains both the transmembrane domain and the death domain. However, the total levels of Fas protein and mRNA were comparable to controls, and Fas internalization occurred normally. Expression of FADD, caspase-8 and -3 were not elevated and the pathway inhibitors: ERK, c-FLIP and XIAP were not decreased. These results suggest that SBDS loss results in abnormal accumulation of Fas at the plasma membrane, where it sensitizes the cells to stimulation by Fas ligand.
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reactive oxygen species regulate spontaneous and fas mediated apoptosis in SBDS deficient cells
Blood, 2008Co-Authors: Chhaya Ambekar, Herman Yeger, Yigal DrorAbstract:Background and hypotheses : Shwachman-Diamond syndrome (SDS) is an autosomal recessive disorder characterized by bone marrow failure, pancreatic insufficiency, and a marked propensity for myelodysplastic syndrome and leukemia. Approximately 90% of the patients have mutations in the SBDS gene, but the function of the gene is unknown. We previously showed that marrow cells from SDS patients and SBDS-deficient HeLa Cells are characterized by accelerated apoptosis, overexpression of Fas and hypersensitive to Fas stimulation. Involvement of reactive oxygen species (ROS; oxidative stress) have been shown to be related to Fas hypersensitivity and overexpression in a variety of cell types. Therefore, we hypothesized that functional deficiency in SBDS in cells that express Fas could lead to impaired ROS generation and a subsequent increase in spontaneous and Fas-mediated apoptosis and decrease in cell growth. Methods : We used shRNA-mediated SBDS-knockdown HeLa cells as a model. We investigated whether SBDS-deficiency increases ROS levels and if antioxidants can rescue the cell growth and apoptosis phenotype. To measure ROS formation cells were incubated with DCFH-DA and fluorescence measured in Gemini Spectra MAX microplate reader. Staining with annexin V and propidium iodide was done to determine apoptosis and necrotic cell death. MTT assay was used to measure cell viability. Results : ROS levels in SBDS knockdown cells were significantly increased compared to control. Apoptosis analysis by annexin V and propidium iodide showed a marked decrease in cell viability in the SBDS- knockdown cells. NAC treatment decreased ROS levels, enhanced ERK phosphorylation (pERK), improved cell viability, and decreased apoptotic and necrotic cell death. Stimulation of the Fas signaling pathway by CH-11 (activating anti-Fas antibody) and Fas ligand showed increased ROS production in SBDS Knockdown cells. CH-11 treatment showed a marked increase in apoptotic and necrotic cell death after 24 and 48hrs incubation. Cell viability decreased by 40% and 80% after 24 and 48hrs incubation with CH-11. Treatment with NAC lowered ROS levels, enhanced pERK expression, protected the cells from Fas-mediated early apoptosis and improved cell survival. Conclusion : We have demonstrated that stable loss of SBDS results in increased ROS levels, leading to apoptotic and necrotic cell death. Thus, increased baseline and Fas-stimulated ROS could result in increased sensitivity to apoptosis and necrotic cell death. NAC appeared to reverse the ROS-mediated decrease in cell survival and apoptotic cell death. Our data support the novel concept that SBDS may be a homeostatic regulator of oxidative stress
Eiji Nakashima - One of the best experts on this subject based on the ideXlab platform.
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The Shwachman–Bodian–Diamond syndrome gene mutations cause a neonatal form of spondylometaphysial dysplasia (SMD) resembling SMD Sedaghatian type
Journal of Medical Genetics, 2007Co-Authors: Nishimura, Phillip Cox, Y. Miyamoto, Trevor Cole, Yuichiro Hirose, David L. Rimoin, Daniel H. Cohn, Eiji Nakashima, Ralph S Lachman, Bronwyn KerrAbstract:The Shwachman–Bodian–Diamond syndrome (SBDS) gene is a causative gene for Shwachman–Diamond syndrome, an autosomal recessive disorder with exocrine pancreatic insufficiency, bone marrow dysfunction and skeletal dysplasia. We report here on two patients with skeletal manifestations at the severest end of the phenotypic spectrum of SBDS mutations. An 11‐year‐old Japanese girl presented with neonatal respiratory failure necessitating lifelong ventilation support, severe short stature and severe developmental delay. She developed neutropenia in infancy, and decreased serum amylase was noted in childhood. A British boy was a stillbirth with pulmonary hypoplasia and hepatic fibrosis found on autopsy. Both cases had neonatal skeletal manifestations that included platyspondyly, lacy iliac crests and severe metaphysial dysplasia, and thus did not fall in the range of the known Shwachman–Diamond syndrome skeletal phenotype but resembled spondylometaphysial dysplasia (SMD) Sedaghatian type. The girl harboured a recurrent mutation (183TA→CT) and a novel missense mutation (79T→C), whereas the boy carried two recurrent mutations (183TA→CT and 258+2T→C). We also examined SBDS in one typical case with SMD Sedaghantian type and eight additional cases with neonatal SMD, but failed to discover SBDS mutations. Our experience expands the phenotypic spectrum of SBDS mutations, which, at its severest end, results in severe neonatal SMD.
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The Shwachman-Bodian-Diamond syndrome gene mutations cause a neonatal form of spondylometaphysial dysplasia (SMD) resembling SMD Sedaghatian type.
Journal of medical genetics, 2007Co-Authors: Gen Nishimura, Phillip Cox, Y. Miyamoto, Trevor Cole, Yuichiro Hirose, David L. Rimoin, Daniel H. Cohn, Eiji Nakashima, Ralph S Lachman, Bronwyn KerrAbstract:The Shwachman-Bodian-Diamond syndrome (SBDS) gene is a causative gene for Shwachman-Diamond syndrome, an autosomal recessive disorder with exocrine pancreatic insufficiency, bone marrow dysfunction and skeletal dysplasia. We report here on two patients with skeletal manifestations at the severest end of the phenotypic spectrum of SBDS mutations. An 11-year-old Japanese girl presented with neonatal respiratory failure necessitating lifelong ventilation support, severe short stature and severe developmental delay. She developed neutropenia in infancy, and decreased serum amylase was noted in childhood. A British boy was a stillbirth with pulmonary hypoplasia and hepatic fibrosis found on autopsy. Both cases had neonatal skeletal manifestations that included platyspondyly, lacy iliac crests and severe metaphysial dysplasia, and thus did not fall in the range of the known Shwachman-Diamond syndrome skeletal phenotype but resembled spondylometaphysial dysplasia (SMD) Sedaghatian type. The girl harboured a recurrent mutation (183TA-->CT) and a novel missense mutation (79T-->C), whereas the boy carried two recurrent mutations (183TA-->CT and 258+2T-->C). We also examined SBDS in one typical case with SMD Sedaghantian type and eight additional cases with neonatal SMD, but failed to discover SBDS mutations. Our experience expands the phenotypic spectrum of SBDS mutations, which, at its severest end, results in severe neonatal SMD.
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Novel SBDS mutations caused by gene conversion in Japanese patients with Shwachman-Diamond syndrome
Human Genetics, 2004Co-Authors: Eiji Nakashima, Mitsuo Masuno, Akihiko Mabuchi, Yoshio Makita, Gen Nishimura, Hirofumi Ohashi, Shiro IkegawaAbstract:Shwachman-Diamond syndrome (SDS; OMIM 260400) is an autosomal recessive disorder characterized by exocrine pancreatic insufficiency, bone marrow dysfunction and metaphyseal chondrodysplasia. SDS is caused by mutations in SBDS , an uncharacterized gene. A previous study in SDS patients largely of European ancestry found that most SBDS mutations occurred within a ~240-bp region of exon 2 and resulted from gene conversion due to recombination with a pseudogene, SBDSP . It is unknown, however, whether these findings are applicable to other ethnic groups. To address this question, we examined SBDS mutations in six Japanese families with SDS by direct sequencing. We identified compound heterozygous mutations in four families: two were recurrent (96–97insA, 258+2T>C), and three were novel [292–295delAAAG, (183–184TA>CT +201A>G), (141C>T+183–184TA>CT+201A>G)] mutations. Most of these mutations also appear to result from gene conversion, but the conversion events occurred at various sites between intron 1 and exon 3. Thus, gene conversion mutations in SBDS are common to different ethnic groups, but they are not confined to a limited region of the gene.
