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 Blood, 16 April 2009, Vol. 113, No. 16, pp. 3690-3695.
Prepublished online as a Blood First Edition Paper on February 2, 2009; DOI 10.1182/blood-2008-10-176396.

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HEMATOPOIESIS AND STEM CELLS

Forward RNAi screens in primary human hematopoietic stem/progenitor cells

Nicole Ali1,2, Christine Karlsson3, Marie Aspling1,2, Guang Hu4, Nir Hacohen5,6, David T. Scadden1,2,7, and Jonas Larsson13

1 Center for Regenerative Medicine, Massachusetts General Hospital, Harvard Medical School, Boston; 2 Harvard Stem Cell Institute, Harvard University, Cambridge, MA; 3 Molecular Medicine and Gene Therapy, Lund Stem Cell Center, Lund University, Lund, Sweden; 4 Howard Hughes Medical Institute and Department of Genetics, Center for Genetics and Genomics, Brigham and Women's Hospital, Harvard Medical School, Boston, MA; 5 Broad Institute of Massachusetts Institute of Technology and Harvard University, Cambridge; 6 Center for Immunology and Inflammatory Diseases, Massachusetts General Hospital, Harvard Medical School, Boston; and 7 Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA

The mechanisms regulating key fate decisions such as self-renewal and differentiation in hematopoietic stem and progenitor cells (HSPC) remain poorly understood. We report here a screening strategy developed to assess modulators of human hematopoiesis using a lentiviral short hairpin RNA (shRNA) library transduced into cord blood-derived stem/progenitor cells. To screen for modifiers of self-renewal/differentiation, we used the limited persistence of HSPCs under ex vivo culture conditions as a baseline for functional selection of shRNAs conferring enhanced maintenance or expansion of the stem/progenitor potential. This approach enables complex, pooled screens in large numbers of cells. Functional selection identified novel specific gene targets (exostoses 1) or shRNA constructs capable of altering human hematopoietic progenitor differentiation or stem cell expansion, respectively, thereby demonstrating the potential of this forward screening approach in primary human stem cell populations.


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