Abstract Although initially identified about 90 years ago, Fanconi Anemia (FA) remains a fatal genetic disease with few therapeutic options. Patients with the genetic disorder FA exhibit developmental malformations, bone marrow failure (BMF), and increased cancer susceptibility. Twenty-one FA genes cooperate in a genome stability pathway that is essential for repair of DNA interstrand crosslink (ICL) damage and tolerance of replication stress. The cell proliferative and survival defects in FA result in hematopoietic stem cell (HSC) exhaustion that contributes to BMF. However, how FA pathway disruption selectively affects HSC function is not understood. Signaling events that can reduce FA severity remain to be established. Importantly, interventions to mitigate FA associated HSC defects do not exist, aside from allogeneic stem cell transplant. This R01 renewal is based on our discovery that activation of cytokine/JAK signaling ameliorates the HSPC defects associated with FA. Specifically, we showed that loss of LNK (or SH2B3), a critical negative regulator of cytokine/JAK signaling fully restored hematopoietic stem/progenitor cell (HSPC) functions in FA mutant mice and prevented FA associated genome instability. HSCs nullizygous for both Lnk and the central component of the FA pathway, Fancd2, exhibited near normal repopulation and self-renewal capability in serial transplantation assays. Interestingly, LNK did not play an overt role in repair of ICL DNA damage; rather, Lnk deficiency stabilized stalled replication forks and alleviated replication stress that is characteristic of FA cells. These results were strongly associated with increased HSC fitness, ex vivo growth, survival and genomic stability in FA HSPCs that harbored concomitant Lnk deficiency. Here, we propose comprehensive and in-depth analyses on the role of LNK in FA pathogenesis and therapy. In aim 1, we propose to study the mechanisms by which LNK-regulated signaling pathways stabilize replication forks and mitigate replication stress. Moreover, the FA pathway is reported to play a critical role in mitigating two types of endogenous genotoxic stress, oxidative DNA damage caused by reactive oxygen species and aldehyde-induced DNA damage generated by cellular metabolism. Our preliminary data suggested that Lnk deficiency reduces both types of stress. Thus, in aim 2 we will determine mechanism by which Lnk deficiency alleviates endogenous genotoxic stress and preserves HSPC functions. Lastly and importantly, we will determine if targeting LNK could be widely used as a FA suppressor in aim 3. Since mutations in the FA core complex FANCA/C/G account for ~90% FA in humans, we plan to expand our studies to test if Lnk deficiency could restore HSC function in Fanca/c/g deficient mice. This study will culminate in exploring therapeutic avenues of inhibiting LNK to rescue hematopoietic defects in primary HSPCs from FA patients. To our knowledge, this suppression of HSPC dysfunction in vivo w...