Rev1 hematopoietic stem cell (HSC) display competitive and proliferative defects (see alsosupplemental Figure 1). The involvement of TLS in tolerance of endogenous DNA damage in the hematopoietic system was investigated by analyzing Rev1 blood and bone marrow, by competitive repopulation experiments, and by culture of hematopoietic stem and progenitor cells (HSPCs) in vitro. *P < .05; **P < .01; ***P < .001; ****P < .0001. Data are mean ± standard error of the mean (SEM). (A) Helix-distorting nucleotide lesions (blue spheres) can be repaired by ggNER, dependent on the Xpc gene. In case a lesion escapes timely repair, it arrests processive replication (black rectangle), resulting in replication stress and DNA damage signaling. The lesion can be bypassed postreplicatively by Rev1-dependent DNA TLS (zig-zag line). Thereby, TLS prevents the induction of replication stress and double-stranded DNA (dsDNA) breaks. TLS frequently misincorporates (in red) opposite the damaged nucleotide, which originates nucleotide substitution mutations. (B) Cytopenia in 26- to 30-month-old Rev1 mice (n = 11), compared with age-matched WT mice (n = 6). (C) Relative contribution of myeloid and lymphoid cells in the WT and Rev1 blood at 3 months of age (3m) and when moribund (MB). N = 10. (D) Frequencies of LSK, LSK34-, and LSK-SLAM cells in bone marrow of 5-month-old Rev1 (n = 5) and WT mice (n = 5). Frequencies are depicted as percent of mononuclear cells. (E) Impaired function of Rev1-deficient HSCs as demonstrated by competitive repopulation assays. Scheme of competitive transplantation experiments (top). Competitive transplantation of WT (n = 9) and Rev1 HSCs (n = 8) (bottom). (See also supplemental Figure 1.) (F) Impaired proliferative capacity of HSPCs as demonstrated by reduced CAFC numbers from 5-month-old WT (n = 4) and Rev1 (n = 4) mice. (G) Sizes of colonies after single-cell sorting of LSK-SLAM cells from 5-month-old Rev1 (n = 3) and WT (n = 3) mice.