2007;449:105C108. or LSD1 to KU70 impacts chromatin structure but does not correlate well with their direct histone modification LY2794193 functions, and SIRT1 helps maintain histone H4K16 acetylation and open chromatin for repair. The competitive KU70 binding by these proteins affects cancer cells’ ability to repair broken DNA and acquire resistant genetic mutations in CML and prostate cancer cells. We identify that the core domain of KU70 binds both LSD1 and SIRT1, forming a molecular basis for the competition. The C-terminal SAP motif of KU70 mediates LSD1/SIRT1 IL-10C competitive interaction by suppressing LSD1 binding to KU70 and ectopic expression of SAP-deleted KU70 to CML cells compromises their ability to acquire BCR-ABL mutations. Our study reveals a novel cellular stress response mechanism in cancer cells and a key role of LSD1/SIRT1/KU70 dynamic interaction in regulating DNA repair and mutation acquisition. strong class=”kwd-title” Keywords: chronic myeloid leukemia, BCR-ABL, NHEJ, lysine deacetylase, lysine demethylase INTRODUCTION Transformation of hematopoietic stem cells by the BCR-ABL fusion oncogene leads to development of chronic myeloid leukemia (CML). Tyrosine kinase inhibitor imatinib mesylate (IM) is an effective treatment for the disease [1], but forfeits its efficacy in some patients, particularly those in advanced phases of the disease, due to acquired resistance through BCR-ABL mutations [2, 3]. To dissect the mechanisms of resistance, we previously developed a culture model with a blast crisis CML cell line that recapitulates clinical CML acquired resistance through BCR-ABL mutations [4]. Using LY2794193 this model, we showed that NAD+ dependent protein lysine deacetylase SIRT1 is critically involved in promoting acquisition of BCR-ABL mutations in response to IM treatment [5]. We also demonstrated that LY2794193 induction of cell differentiation by all-trans retinoid acid (ATRA) increases expression of cellular NAD+ cyclase CD38 that reduces cellular NAD+ concentration, inhibits SIRT1 activity and blocks BCR-ABL mutation acquisition [6]. SIRT1 is a multi-functional enzyme that deacetylates histones including H4K16 to regulate gene expression and many nonhistone proteins for biological functions [7]. A key downstream effector of SIRT1 is KU70, a crucial factor for non-homologous end joining (NHEJ). NHEJ is a major DNA repair mechanism in mammalian cells for double strand breaks (DSBs) that can arise from intrinsic sources such as reactive oxygen species or from external sources such as cancer chemotherapeutic agents and ionizing radiation [8]. NHEJ repair is initiated when KU70/KU80 heterodimer binds to broken DNA ends. Both KU factors are essential for NHEJ as deletion of either one leads to DSB repair impairment and sensitivity to radiation [9, 10]. KU70 is subjected to lysine acetylation modification [11], and deacetylation of KU70 by SIRT1 stimulates KU70-mediated NHEJ repair [5, 12]. Besides its well-known function in NHEJ, KU70 has roles in non-DNA repair events, which are less understood. Among them, SIRT1 deacetylation of KU70 sequesters BAX protein in the cytoplasm to prevent apoptosis initiation and extend cell survival [13]. We have shown that SIRT1 promotes acquisition of resistant BCR-ABL mutations in CML cells in association with its ability to stimulate aberrant NHEJ activity by deacetylating KU70 [5, 6]. Lysine specific demethylase 1 (LSD1) is a monoamine oxidase homolog that demethylates histone H3K4 and H3K9 [14C16], and functions to regulate gene expression [17, 18]. LSD1 also demethylates nonhistone proteins such as for example p53 for regulating cell success [19]. Previously, we showed that p53 deacetylation by SIRT1 has a key function for drug level of resistance of CML stem/progenitor cells [20, 21]. As a result, both SIRT1 and LSD1 can target on a single non-histone protein to modulate its functions. In addition, LSD1 and SIRT1 may co-exist within a repressor organic to modify gene transcription [22]. However, it really is unknown if LSD1 may regulate KU70 and NHEJ features. We hypothesized that SIRT1 and LSD1 might co-regulate KU70 initially.
