Regulation of Hematopoietic Stem Cell Quiescence by Ku70.

Abstract 3613

Poster Board III-549

Genomic integrity is essential for organism development and longevity, and in large part is mediated by DNA repair proficiency. Non-homologous end-joining (NHEJ) is essential for DNA double-strand break repair in all cells and for VDJ processing in B and T cells. NHEJ is also critical for hematopoietic stem cell (HSC) maintenance and function, but the mechanisms by which the NHEJ pathway regulates HSC function are not known. Ku70 is a key component of NHEJ; Ku70-deficient mice are hypersensitive to radiation and show a leaky SCID phenotype. Previously we have shown that HSC from Ku70-deficient mice are defective in repopulation, self-renewal and competitive repopulation. Also, mice defective in other NHEJ components, Ku80 or ligase 4, display impaired HSC function and reduced HSC numbers during aging. Here we used Ku70-deficient mice to investigate the role of the NHEJ pathway in HSC function.

Ku70-deficient mice show comparable HSC (Sca1+,c-Kit+,Lin-) frequency to Wild-type (WT) mice, and HSCs from Ku70-deficient mice display similar apoptosis rates as HSCs from WT mice. SKL cells from both Ku70-deficient and WT mice proliferate at similar rates in vitro in the presence of IL3, SCF, Flt3-L and Tpo. However, a greater percentage of HSCs from Ku70-deficient mice are actively cycling, and fewer are quiescent than those obtained from WT mice. Further, recipients of Ku70-deficient BM are more sensitive to 5-FU treatment than those receiving WT BM. In addition, WT HSCs efficiently replace endogenous Ku70-deficient HSCs when WT BM cells are transplanted into Ku70-deficient mice in the absence of any conditioning.

Together, our data indicate that loss of quiescence in Ku70-deficient HSCs correlates with dramatic defects observed in repopulation, self-renewal and competitive repopulation. These results suggest that the NHEJ pathway, either through the processes of DNA repair and genome maintenance, or through a stress response signaling pathway, is critical for maintaining HSCs in a quiescent state, avoiding age-related physiological stress, and allowing functional stem cell transplantation.