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Identification of RPS14 as a 5q- syndrome gene by RNA interference screen

Nature volume 451, pages 335–339 (2008)Cite this article

Abstract

Somatic chromosomal deletions in cancer are thought to indicate the location of tumour suppressor genes, by which a complete loss of gene function occurs through biallelic deletion, point mutation or epigenetic silencing, thus fulfilling Knudson’s two-hit hypothesis1. In many recurrent deletions, however, such biallelic inactivation has not been found. One prominent example is the 5q- syndrome, a subtype of myelodysplastic syndrome characterized by a defect in erythroid differentiation2. Here we describe an RNA-mediated interference (RNAi)-based approach to discovery of the 5q- disease gene. We found that partial loss of function of the ribosomal subunit protein RPS14 phenocopies the disease in normal haematopoietic progenitor cells, and also that forced expression of RPS14 rescues the disease phenotype in patient-derived bone marrow cells. In addition, we identified a block in the processing of pre-ribosomal RNA in RPS14-deficient cells that is functionally equivalent to the defect in Diamond–Blackfan anaemia, linking the molecular pathophysiology of the 5q- syndrome to a congenital syndrome causing bone marrow failure. These results indicate that the 5q- syndrome is caused by a defect in ribosomal protein function and suggest that RNAi screening is an effective strategy for identifying causal haploinsufficiency disease genes.

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Figure 1: Screen of the common deleted region for the 5q - syndrome.
Figure 2: Multiple shRNAs targeting RPS14 recapitulate the 5q- syndrome in vitro.
Figure 3: RPS14 is required for 18S pre-rRNA processing and 40S ribosomal subunit formation.
Figure 4: RPS14 overexpression rescues erythroid differentiation in samples from patients with 5q deletions.

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Acknowledgements

We thank Broad Institute RNAi and Genetic analysis platforms for advice, single-nucleotide polymorphism analysis and reagents. This work was supported by grants from the National Heart Lung and Blood Institute to T.R.G., B.L.E. and S.R.E. T.R.G. is an investigator of the Howard Hughes Medical Institute.

Author Contributions B.L.E., J.P., J.B., C.Y.C., P.T. and S.R.E. performed experiments and analysed data. D.E.R. provided essential reagents. N.G., A.R. and E.A. provided samples from patients. B.L.E. and T.R.G. wrote the manuscript.

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Authors and Affiliations

  1. Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA ,

    Benjamin L. Ebert, Jennifer Pretz, Jocelyn Bosco, Cindy Y. Chang, Pablo Tamayo, David E. Root & Todd R. Golub

  2. Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA ,

    Benjamin L. Ebert & Todd R. Golub

  3. Division of Hematology, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA,

    Benjamin L. Ebert

  4. Division of Hematology Oncology, University of Massachusetts Medical School, Worcester, Massachusetts 01655, USA,

    Naomi Galili & Azra Raza

  5. Division of Hematology/Oncology, Department of Medicine, Massachusetts General Hospital, Boston, Massachusetts 02114, USA,

    Eyal Attar

  6. Department of Biochemistry and Molecular Biology, University of Louisville, Kentucky 40292, USA

    Steven R. Ellis

  7. Howard Hughes Medical Institute, Chevy Chase, Maryland 20815, USA ,

    Todd R. Golub

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  1. Benjamin L. Ebert
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Corresponding author

Correspondence to Todd R. Golub.

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This paper contains Supplementary Figures S1-S11 with Legends and Supplementary Tables S1-S5. (PDF 851 kb)

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Ebert, B., Pretz, J., Bosco, J. et al. Identification of RPS14 as a 5q- syndrome gene by RNA interference screen. Nature 451, 335–339 (2008). https://doi.org/10.1038/nature06494

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