Recent advances have highlighted extensive phenotypic and functional similarities between normal stem cells and cancer stem cells. This raises the question of whether disease therapies can be developed that eliminate cancer stem cells without eliminating normal stem cells. Here we address this issue by conditionally deleting the Pten tumour suppressor gene in adult haematopoietic cells. This led to myeloproliferative disease within days and transplantable leukaemias within weeks. Pten deletion also promoted haematopoietic stem cell (HSC) proliferation. However, this led to HSC depletion via a cell-autonomous mechanism, preventing these cells from stably reconstituting irradiated mice. In contrast to leukaemia-initiating cells, HSCs were therefore unable to maintain themselves without Pten. These effects were mostly mediated by mTOR as they were inhibited by rapamycin. Rapamycin not only depleted leukaemia-initiating cells but also restored normal HSC function. Mechanistic differences between normal stem cells and cancer stem cells can thus be targeted to deplete cancer stem cells without damaging normal stem cells.

In mammalian cells, DNA double-strand breaks (DSBs) cause rapid phosphorylation of the H2AX core histone variant (to form ␥-H2AX) in megabase chromatin domains flanking sites of DNA damage. To investigate the role of H2AX in mammalian cells, we generated H2AX-deficient (H2AX ⌬/⌬ ) mouse embryonic stem (ES) cells. H2AX ⌬/⌬ ES cells are viable. However, they are highly sensitive to ionizing radiation (IR) and exhibit elevated levels of spontaneous and IR-induced genomic instability. Notably, H2AX is not required for NHEJ per se because H2AX ⌬/⌬ ES cells support normal levels and fidelity of V(D)J recombination in transient assays and also support lymphocyte development in vivo. However, H2AX ⌬/⌬ ES cells exhibit altered IR-induced BRCA1 focus formation. Our findings indicate that H2AX function is essential for mammalian DNA repair and genomic stability. The DNA in eukaryotic cells is packaged into chromatin, the fundamental unit of which is the nucleosome. The nucleosome consists of DNA wrapped around an octamer of the four core histones-H2A, H2B, H3, and H4 (1). The H2A histones consist of several subfamilies that contain distinct, conserved amino-and carboxyl-terminal amino acid sequences (2). The H2AX subfamily contains a conserved carboxyl-terminal SerGln-Glu (SQE motif) amino acid sequence. This SQE motif represents the consensus in vitro phosphorylation site for members of the phosphoinositide 3-kinase related kinase (PIKK) family that includes the protein kinases DNA-dependent protein kinase catalytic subunit (DNA-PKcs), ataxia telangiectasia mutated (ATM), and ATM and Rad3-related (ATR) (3).The repair of spontaneous and induced DNA double-strand breaks (DSBs) is critical for the maintenance of genomic integrity. In eukaryotic cells, the two major pathways of DSB repair are nonhomologous end-joining (NHEJ) and homologous recombination (HR; refs. 4 and 5). Covalent modifications of core histones via phosphorylation, acetylation, and methylation have been proposed to form a ''histone code'' that is read by cellular proteins to facilitate downstream molecular events (6). In response to DNA damage by agents that induce DNA doublestrand breaks, Mec1, the Saccharomyces cerevisiae homologue of ATR, phosphorylates the SQE motif of H2A (7). This phosphorylation event is required for the efficient repair of chromosomal DSBs by NHEJ but does not appear to be as important for homologous recombination (7). In mammalian cells, H2AX is rapidly phosphorylated on the induction of DSBs by ionizing radiation (IR) and DNA damaging agents (8, 9), resulting in formation of ␥-H2AX foci along megabase chromatin domains flanking DNA damage sites (9).Foci of ␥-H2AX also form at the immunoglobulin heavy chain locus during class switch recombination (CSR) in activated mature B cells (10). CSR occurs between large, highly repetitive S regions and also may be initiated by DSBs (10, 11) and completed by NHEJ factors (12)(13)(14)(15). Notably, CSR is significantly impaired in the absence of H2AX (10). Earlier during lymphocyte development...