EFL1 mutations impair eIF6 release to cause Shwachman-Diamond syndrome

Abstract Shwachman-Diamond syndrome (SDS) is a recessive disorder typified by bone marrow failure and predisposition to hematological malignancies. SDS is predominantly caused by deficiency of the allosteric regulator Shwachman-Bodian-Diamond syndrome that cooperates with elongation factor-like GTPase 1 (EFL1) to catalyze release of the ribosome antiassociation factor eIF6 and activate translation. Here, we report biallelic mutations in EFL1 in 3 unrelated individuals with clinical features of SDS. Cellular defects in these individuals include impaired ribosomal subunit joining and attenuated global protein translation as a consequence of defective eIF6 eviction. In mice, Efl1 deficiency recapitulates key aspects of the SDS phenotype. By identifying biallelic EFL1 mutations in SDS, we define this leukemia predisposition disorder as a ribosomopathy that is caused by corruption of a fundamental, conserved mechanism, which licenses entry of the large ribosomal subunit into translation.

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Attenuated Protein Synthesis Drives the Hematopoietic Defects in Shwachman-Diamond Syndrome

Blood ◽

10.1182/blood.v130.suppl_1.876.876 ◽

2017 ◽

Vol 130 (Suppl_1) ◽

pp. 876-876

Author(s):

Pekka Jaako ◽

Chi C Wong ◽

David Adams ◽

Alan J. Warren

Keyword(s):

Bone Marrow ◽

Protein Synthesis ◽

Progenitor Cells ◽

Bone Marrow Failure ◽

Ribosomal Subunit ◽

Hematopoietic Progenitor ◽

Assembly Factor ◽

Shwachman Diamond Syndrome ◽

Subunit Joining

Abstract Shwachman-Diamond syndrome (SDS) is an autosomal recessive disorder characterized by bone marrow failure and a striking propensity to develop poor prognosis myelodysplastic syndrome and acute myeloid leukemia. In 90 % of cases the disease is caused by biallelic mutations in the gene encoding SBDS. We have shown previously that SBDS is a cytoplasmic ribosome assembly factor that catalyzes the release of the eukaryotic initiation factor 6 (eIF6) from the subunit joining interface of 60S ribosomal subunit (Menne et al, 2007; Finch et al, 2011). Deficiency of SBDS therefore results in aberrant retention of eIF6 on the 60S subunits that in turn perturbs ribosomal subunit joining and the formation of translation-competent 80S ribosomes. However, the mechanism linking defective ribosome assembly to marrow failure and leukemia in SDS remain poorly understood. Lack of viable mouse models presents a barrier to progress in understanding SDS disease pathophysiology and to evaluate novel therapies. We hypothesized that induced overexpression of eIF6 would mimic the consequences of SBDS deficiency by reducing the cytoplasmic pool of free 60S subunits and impairing translation. To test this hypothesis we have generated a novel transgenic eIF6 mouse model for SDS using KH2 embryonic stem cells that constitutively express the M2-reverse tetracycline transactivator at the Rosa26 locus with the EIF6 gene targeted downstream of the Col1a1 locus. This strategy permits systemic doxycycline-inducible and graded overexpression of eIF6 through control of the transgene copy number. We have validated that eIF6 overexpression promotes an increase in eIF6-bound cytoplasmic 60S subunits with a concomitant reduction in 80S ribosomes and polysomes in c-kit+ hematopoietic progenitor cells isolated from the transgenic eIF6 mice, thereby recapitulating the ribosomal subunit joining defect observed in patients with SDS. In vitro, the hematopoietic progenitor cells exhibit a strict eIF6 dose-dependent expansion defect. In vivo, mice with graded eIF6 overexpression are viable but develop macrocytic anemia with reticulocytopenia, thrombocytosis and mild leukopenia. Bone marrow transfer experiments demonstrate that the phenotype is autonomous to the hematopoietic system. Longitudinal phenotypic analyses in primary and transplanted animals are ongoing. Flow cytometric analysis of the bone marrow from transgenic eIF6 mice reveals a significant increase in the frequencies of preCFU-E and CFU-E erythroid progenitor cells and erythroblasts, but a significant reduction in the frequency of reticulocytes. Furthermore, we observe a striking accumulation of abnormal orthochromatic erythroblast-like cells that appear to have failed to enucleate, comprising approximately 1.5 % of the total bone marrow cells. Amnis ImageStream analysis, which combines flow cytometry with fluorescent microscopy, reveals a significant decrease in the frequency of erythroblasts that are able to complete the enucleation process. To address the underlying mechanism, we hypothesized that by impairing the formation of translation-competent 80S ribosomes, eIF6 overexpression would reduce the global rate of protein synthesis. Indeed, O-propargyl-puromycin incorporation assays established that the erythroblasts from the transgenic eIF6 mice have an approximately 3-fold reduction in global protein synthesis rate. Furthermore, our preliminary data suggest that the erythroid phenotype is p53-independent. Finally, erythroblasts from the transgenic eIF6 mice show a significant increase in levels of reactive oxygen species, but the functional significance of this finding remains unclear. We conclude that reduced rates of global translation drive defective hematopoiesis in the transgenic eIF6 mice. Importantly, eIF6 overexpression in vivo phenocopies SBDS depletion in human CD34+ cells (Sen et al, 2011). Together with the recent discovery of DNAJC21 (the human homologue of the 60S ribosomal assembly factor JJJ1 in yeast) as an SDS disease gene, our data support the hypothesis that deregulated cytoplasmic 60S subunit maturation and reduced translation are the primary drivers of the hematopoietic defect in SDS. Our viable transgenic eIF6 mouse model provides a unique tool to further dissect the mechanisms that underlie bone marrow failure and malignant transformation in SDS and for the development of novel therapeutics. Disclosures No relevant conflicts of interest to declare.

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Defective ribosome assembly in Shwachman-Diamond syndrome

Blood ◽

10.1182/blood-2011-06-353938 ◽

2011 ◽

Vol 118 (16) ◽

pp. 4305-4312 ◽

Cited By ~ 96

Author(s):

Chi C. Wong ◽

David Traynor ◽

Nicolas Basse ◽

Robert R. Kay ◽

Alan J. Warren

Keyword(s):

Ribosome Biogenesis ◽

Bone Marrow Failure ◽

Pancreatic Insufficiency ◽

Ribosomal Subunit ◽

Initiation Factor ◽

Ribosome Assembly ◽

Restrictive Temperature ◽

Poor Growth ◽

Shwachman Diamond Syndrome ◽

Subunit Joining

AbstractShwachman-Diamond syndrome (SDS), a recessive leukemia predisposition disorder characterized by bone marrow failure, exocrine pancreatic insufficiency, skeletal abnormalities and poor growth, is caused by mutations in the highly conserved SBDS gene. Here, we test the hypothesis that defective ribosome biogenesis underlies the pathogenesis of SDS. We create conditional mutants in the essential SBDS ortholog of the ancient eukaryote Dictyostelium discoideum using temperature-sensitive, self-splicing inteins, showing that mutant cells fail to grow at the restrictive temperature because ribosomal subunit joining is markedly impaired. Remarkably, wild type human SBDS complements the growth and ribosome assembly defects in mutant Dictyostelium cells, but disease-associated human SBDS variants are defective. SBDS directly interacts with the GTPase elongation factor-like 1 (EFL1) on nascent 60S subunits in vivo and together they catalyze eviction of the ribosome antiassociation factor eukaryotic initiation factor 6 (eIF6), a prerequisite for the translational activation of ribosomes. Importantly, lymphoblasts from SDS patients harbor a striking defect in ribosomal subunit joining whose magnitude is inversely proportional to the level of SBDS protein. These findings in Dictyostelium and SDS patient cells provide compelling support for the hypothesis that SDS is a ribosomopathy caused by corruption of an essential cytoplasmic step in 60S subunit maturation.

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Impaired ribosomal subunit association in Shwachman-Diamond syndrome

Blood ◽

10.1182/blood-2012-04-420166 ◽

2012 ◽

Vol 120 (26) ◽

pp. 5143-5152 ◽

Cited By ~ 53

Author(s):

Nicholas Burwick ◽

Scott A. Coats ◽

Tomoka Nakamura ◽

Akiko Shimamura

Keyword(s):

Autosomal Recessive ◽

Ribosomal Subunit ◽

Wild Type ◽

Ribosomal Assembly ◽

Shwachman Diamond Syndrome ◽

Low Levels ◽

Maturation Defect ◽

Biallelic Mutations ◽

Subunit Joining

Abstract 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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New Bone Marrow Failure Genes: DNAJC21 and ERCC6L2

Blood ◽

10.1182/blood.v130.suppl_1.sci-22.sci-22 ◽

2017 ◽

Vol 130 (Suppl_1) ◽

pp. SCI-22-SCI-22

Author(s):

Inderjeet Dokal

Keyword(s):

Bone Marrow ◽

Dna Damage ◽

Nuclear Export ◽

Conflicts Of Interest ◽

Excision Repair ◽

Bone Marrow Failure ◽

Retinal Dystrophy ◽

Ribosomal Subunit ◽

Genotoxic Stress ◽

Biallelic Mutations

A significant number of cases with bone marrow failure present with one or more extra-hematopoietic abnormality. This suggests a constitutional or genetic basis, and yet many of them remain uncharacterized. Through exome sequencing, we have recently identified two sub groups of these cases, one defined by germline biallelic mutations in DNAJC21 (DNAJ homolog subfamily C member 21) and the other in ERCC6L2 (excision repair cross complementing 6 like-2). Patients with DNAJC21 mutations are characterized by global bone marrow failure in early childhood. They can also have a variable number of extra-hematopoietic abnormalities such as short stature and retinal dystrophy. The encoded protein associates with ribosomal RNA (rRNA) and plays a highly conserved role in the maturation of the 60S ribosomal subunit. Lymphoblastoid patient cells exhibit increased sensitivity to the transcriptional inhibitor actinomycin D and reduced levels of rRNA. Characterisation of mutations has revealed impairment in interactions with cofactors (PA2G4, HSPA8 and ZNF622) involved in 60S maturation. DNAJC21 deficiency results in cytoplasmic accumulation of the 60S nuclear export factor PA2G4, aberrant ribosome profiles and increased cell death. Collectively these findings demonstrate that biallelic mutations in DNAJC21 cause disease due to defects in early nuclear rRNA biogenesis and late cytoplasmic maturation of the 60S subunit. Patients harbouring biallelic ERCC6L 2 mutations are characterized by bone marrow failure (in childhood or early adulthood) and one or more extra-hematopoietic abnormality such as microcephaly. Knockdown of ERCC6L2 in human cells significantly reduces their viability upon exposure to the DNA damaging agent irofulven but not etoposide and camptothecin suggesting a role in nucleotide excision repair. ERCC6L2 knockdown cells and patient cells harbouring biallelic ERCC6L2 mutations also display H2AX phosphorylation that significantly increases upon genotoxic stress, suggesting an early DNA damage response. ERCC6L2 is seen to translocate to mitochondria as well as the nucleus in response to DNA damage and its knockdown induces intracellular reactive oxygen species (ROS). Treatment with the ROS scavenger, N-acetyl-cysteine, attenuates the irofulven-induced cytotoxicity in ERCC6L2 knockdown cells and abolishes its traffic to mitochondria and nucleus in response to this DNA damaging agent. Collectively, these observations suggest that ERCC6L2has a pivotal rolein DNA repair and mitochondrial function. In conclusion, ERCC6L2 and DNAJC21 have an important role in maintaining genomic stability and ribosome biogenesis, respectively. They bring into focus new biological connections/pathways whose constitutional disruption is associated with defective hematopoiesis since patients harbouring germline biallelic mutations in these genes uniformly have bone marrow failure. Disclosures No relevant conflicts of interest to declare.

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SBDS Protein Plays a Role in Ribosome Biogenesis.

Blood ◽

10.1182/blood.v108.11.185.185 ◽

2006 ◽

Vol 108 (11) ◽

pp. 185-185

Author(s):

Karthik A. Ganapathi ◽

Karyn M. Austin ◽

Maggie Malsch ◽

Akiko Shimamura

Keyword(s):

Ribosome Biogenesis ◽

Actinomycin D ◽

Bone Marrow Failure ◽

Pancreatic Insufficiency ◽

Protein Translation ◽

Exocrine Pancreatic Insufficiency ◽

Nucleolar Localization ◽

28S Rrna ◽

Wild Type ◽

Shwachman Diamond Syndrome

Abstract Shwachman-Diamond syndrome is an autosomal recessive disorder characterized by exocrine pancreatic insufficiency, bone marrow failure, and leukemia predisposition. The majority of patients with Shwachman-Diamond syndrome harbor mutations in the SBDS gene. SBDS is a novel gene of unknown function and is highly conserved throughout evolution. Studies of the yeast orthologue, YLR022c/SDO1, suggest that SBDS may play a role in ribosome biogenesis. In support of this hypothesis, we have found that the SBDS protein shuttles in and out of the nucleolus. Previously we have shown that SBDS nucleolar localization is regulated in a cell cycle-dependant manner. We now find that SBDS nucleolar localization is also lost following exposure to actinomycin D, suggesting that SBDS nucleolar localization is dependent on active ribosomal RNA (rRNA) transcription. In cell survival assays, SBDS−/− patient-derived cells are sensitive to actinomycin D treatment relative to normal control cells. Introduction of the wild-type SBDS cDNA into SBDS−/− cells corrects their actinomycin D sensitivity, confirming that the observed sensitivity is SBDS-dependent. In contrast, SBDS−/− cells do not exhibit increased sensitivity to cyclohexamide, a protein translation inhibitor. Consistent with this result, SBDS protein co-localizes with ribosomal precursor subunits but not with mature polysomes upon sucrose gradient sedimentation. No differences in polysome profiles are observed between SBDS−/− cells and wild type control cells. Gel filtration studies suggest that SBDS associates into a complex with other proteins. SBDS co-immunoprecipitates with other nucleolar proteins involved in rRNA biogenesis. RNA immunoprecipitation studies reveal that SBDS also associates with the 28S rRNA but not the 18S rRNA. These findings support the hypothesis that SBDS plays a role in ribosome biogenesis

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Depletion of the Shwachman-Diamond Syndrome Protein in Hematopoietic Progenitor Cells Impairs Growth, Colony Formation, and Ribosome Function.

Blood ◽

10.1182/blood.v112.11.2046.2046 ◽

2008 ◽

Vol 112 (11) ◽

pp. 2046-2046

Author(s):

Gulay Sezgin ◽

Abdallah Nihrane ◽

Steven Ellis ◽

Johnson M. Liu

Keyword(s):

Cell Cycle ◽

Fluorescent Protein ◽

Bone Marrow Failure ◽

Gradient Centrifugation ◽

Flow Cytometric Analysis ◽

Ribosomal Subunit ◽

Autosomal Recessive Disorder ◽

Cycle Phase ◽

Cell Extracts ◽

Shwachman Diamond Syndrome

Abstract Shwachman-Diamond syndrome (SDS) is an autosomal recessive disorder characterized by pancreatic exocrine dysfunction, skeletal abnormalities, and bone marrow failure, which can evolve to leukemia. Mutations in SBDS have been shown to cause SDS. We and other investigators have suggested that SBDS orthologs in yeast play a role in biogenesis and function of the 60S ribosomal subunit. To clarify the unknown function of SBDS in hematopoiesis, human erythroleukemia TF-1 cells were transduced with lentiviral vectors expressing the green fluorescent protein (GFP), neomycin phosphotransferase, and small interfering RNA (siRNA) against SBDS. After transduction, cells were selected for neomycin resistance and then sorted by flow cytometry. To probe for SBDS, an antibody against the carboxyl-terminus of human SBDS was generated, and individual TF-1 cell clones expressing different siRNAs were confirmed to knock down SBDS expression by Western blot analysis. Our experiments were aimed at analyzing the cellular effects of SBDS knockdown. The growth and hematopoietic colony forming potential of TF-1 knockdown cells were markedly hindered when compared to cells stably transduced with siRNA against a scrambled SBDS sequence. Using propidium iodide staining and flow cytometric analysis, we found an increased percentage of knockdown cells retained at the G0/G1 cell cycle phase. To address whether TF-1 cells expressing siRNA against SBDS have a selective deficiency of 60S ribosomal subunits, cell extracts were prepared and polysome profiles examined after sucrose gradient centrifugation. In preliminary experiments, TF-1 cells expressing siRNA against SBDS appeared to show a reduction in free 60S subunits and 80S subunits with a shift toward smaller polysomes, compared to cells expressing the scrambled sequence siRNA. We conclude that depletion of SBDS results in a significant growth and clonogenic defect in TF-1 hematopoietic cells. Our preliminary results also suggest defects in ribosome function and cell cycle transit, which may provide an integrated molecular explanation for the hematopoietic defect in SDS since nucleolar stress has been linked to cell cycle arrest and p53 stabilization.

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Impaired Ribosome Maturation In Human Cells Depleted of Shwachman-Diamond Syndrome Protein SBDS

Blood ◽

10.1182/blood.v116.21.2242.2242 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2242-2242

Author(s):

Gulay Sezgin ◽

Abdallah Nihrane ◽

Adrianna Henson ◽

Max Wattenberg ◽

Steven Ellis ◽

...

Keyword(s):

Conflicts Of Interest ◽

Bone Marrow Failure ◽

Ribosomal Subunit ◽

A549 Cells ◽

Autosomal Recessive Disorder ◽

Human Cells ◽

Yeast Cells ◽

Cycle Phase ◽

Shwachman Diamond Syndrome ◽

Pancreatic Exocrine Dysfunction

Abstract Abstract 2242 Background: Shwachman-Diamond syndrome (SDS) is an autosomal recessive disorder characterized by pancreatic exocrine dysfunction, neurocognitive and skeletal abnormalities, and bone marrow failure. Mutations in SBDS have been shown to cause SDS. From experiments on its yeast ortholog (Haematologica 2010. 95:57-64), SBDS has been implicated in maturation and function of the 60S ribosomal subunit. In particular, subunit maturation in the SDS yeast model was associated with delayed export and accumulation of 60S-like particles in the nucleoplasm. Methods and Results: To clarify its role in human cells, erythroleukemia TF-1 cells were transduced with lentiviral vectors expressing short hairpin RNA (shRNA) against SBDS. Immunoblot assays confirmed approximately 60% knockdown in individual TF-1 cell clones expressing different shRNAs. The growth and hematopoietic colony forming potential of TF-1 knockdown cells were markedly hindered when compared to cells stably transduced with shRNA against a scrambled SBDS sequence. Using Hoechst 33342/Pyronin Y staining and flow cytometry, we also found an increased percentage of knockdown cells retained at the G0/G1 cell cycle phase. To address whether near-complete knockdown of SBDS affected ribosome synthesis as it does in yeast cells, we silenced SBDS in A549 cells. Our data revealed a reduction in polysomes but in contrast to what was observed in yeast, there was no evidence of half-mer polysomes indicative of decreased 60S subunits participating in translation. The absence of half-mers is not unusual in mammalian systems, so to better analyze the effect of SBDS on 60S subunit maturation subunit localization was assessed by co-transfection with a vector expressing a fusion between human RPL29 and enhanced GFP. Preliminary studies indicated a higher percentage of SBDS-depleted cells with nuclear localization of 60S subunits, when compared with normal controls (Fig. 1). Conclusions: Disclosures: No relevant conflicts of interest to declare.

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SBDS and eIF6 Modulate Ribosome Subunit Joining in Shwachman Diamond Syndrome,

Blood ◽

10.1182/blood.v118.21.3438.3438 ◽

2011 ◽

Vol 118 (21) ◽

pp. 3438-3438

Author(s):

Nicholas Burwick ◽

Scott Coats ◽

Akiko Shimamura

Keyword(s):

Stromal Cells ◽

Ribosome Biogenesis ◽

Conflicts Of Interest ◽

Ribosomal Subunit ◽

Marrow Stromal Cells ◽

Vitro Assay ◽

Amino Terminal ◽

Shwachman Diamond Syndrome ◽

Subunit Joining

Abstract 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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Linking Defective Ribosome Maturation to Shwachman-Diamond Syndrome

Blood ◽

10.1182/blood.v122.21.sci-36.sci-36 ◽

2013 ◽

Vol 122 (21) ◽

pp. SCI-36-SCI-36

Author(s):

Alan John Warren

Keyword(s):

Ribosome Biogenesis ◽

Conflicts Of Interest ◽

Ribosomal Subunit ◽

Protein Translation ◽

Ribosome Assembly ◽

Genetic Dissection ◽

Gtp Hydrolysis ◽

Shwachman Diamond Syndrome ◽

Novel Therapeutic Strategies ◽

Syndrome Protein

Abstract Ribosomes are RNA-protein machines that translate the genetic information encoded by the mRNA template in all living cells. Recent high-resolution structures of the ribosome have revolutionized our understanding of protein translation. However, the mechanisms of ribosome assembly and the surveillance mechanisms that monitor this process and couple it to growth are poorly understood. Causative mutations and deletions of genes involved in ribosome biogenesis define an emerging group of disorders known as the ribosomopathies. Recent work from my laboratory strongly supports the hypothesis that Shwachman-Diamond syndrome (SDS) is a ribosomopathy caused by defective maturation of the large ribosomal subunit. Elucidation of the specific function of the SBDS protein that is deficient in SDS is revealing unexpected new insights that extend our understanding of the mechanisms underlying the late cytoplasmic steps of ribosome assembly and the quality control surveillance pathways that monitor 60S maturation. Genetic dissection of this pathway may inform novel therapeutic strategies for SDS. 1. Wong C.C., Traynor D., Basse N., Kay R.R., Warren A.J. Defective ribosome assembly in Shwachman-Diamond syndrome. Plenary Paper, Blood. 2011 Oct 20;118(16):4305-12. 2. Finch A.J., Hilcenko C., Basse N., Drynan L.F., Goyenechea B., Menne T.F., González Fernández Á., Simpson P., D’Santos C.S., Arends M.J., Donadieu J., Bellanné-Chantelot C., Costanzo M., Boone C., McKenzie A.N., Freund S.M., Warren A.J. Uncoupling of GTP hydrolysis from eIF6 release on the ribosome causes Shwachman-Diamond syndrome. Genes and Development (2011) 25: 917-929. 3. Menne T.M., Goyenechea B., Sánchez-Puig N., Wong C.C., Tonkin L.M., Ancliff P., Brost R.L., Costanzo M., Boone C. and Warren A.J. The Shwachman-Bodian-Diamond syndrome protein mediates translational activation of ribosomes in yeast. Nature Genetics (2007) 39: 486-95. Disclosures: No relevant conflicts of interest to declare.

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Shwachman-Diamond Syndrome and the Quality Control of Ribosome Assembly

Blood ◽

10.1182/blood.v128.22.sci-42.sci-42 ◽

2016 ◽

Vol 128 (22) ◽

pp. SCI-42-SCI-42

Author(s):

Alan J. Warren

Keyword(s):

Quality Control ◽

Ribosomal Proteins ◽

Active Sites ◽

Structural Integrity ◽

Conflicts Of Interest ◽

Bone Marrow Failure ◽

Ribosomal Subunit ◽

Ribosome Assembly ◽

Final Maturation ◽

Shwachman Diamond Syndrome

Abstract The synthesis of new ribosomes is a fundamental conserved process in all cells. Ribosomes are pre-assembled in the nucleus and subsequently exported to the cytoplasm where they acquire functionality through a series of final maturation steps that include formation of the catalytic center, recruitment of the last remaining ribosomal proteins and the removal of inhibitory assembly factors. Surprisingly, a number of key factors (SBDS, DNAJC21, RPL10 (uL16)) involved in late cytoplasmic maturation of the large (60S) ribosomal subunit are mutated in both inherited and sporadic forms of leukemia. In particular, biallelic mutations in the SBDS gene cause Shwachman-Diamond syndrome (SDS), a recessive bone marrow failure disorder with significant predisposition to acute myeloid leukemia. By using the latest advances in single-particle cryo-electron microscopy to elucidate the function of the SBDS protein, we have uncovered an elegant mechanism that couples final maturation of the 60S subunit to a quality control assessment of the structural integrity of the active sites of the ribosome. Further molecular dissection of this pathway may inform novel therapeutic strategies for SDS and leukemia more generally. References: 1. Weis F, Giudice E, Churcher M,et al. Mechanism of eIF6 release from the nascent 60S ribosomal subunit. Nat Struct Mol Biol, (2015) Nov;22(11):914-9. 2. Wong CC, Traynor D, Basse N, et al. Defective ribosome assembly in Shwachman-Diamond syndrome. Plenary Paper, Blood. 2011 Oct 20;118(16):4305-12. 3. Finch AJ, Hilcenko C, Basse N, et al. Uncoupling of GTP hydrolysis from eIF6 release on the ribosome causes Shwachman-Diamond syndrome. Genes Dev (2011) 25: 917-929. 4. Menne TM, Goyenechea B, Sánchez-Puig N, et al. The Shwachman-Bodian-Diamond syndrome protein mediates translational activation of ribosomes in yeast. Nature Genetics (2007) 39: 486-95. Disclosures No relevant conflicts of interest to declare.

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