The extracellular signal-regulated kinase 1/2 modulates the intracellular localization of DNA methyltransferase 3A to regulate erythrocytic differentiation.

DNA methylation, catalyzed by DNA methyltransferases (DNMTs), is a heritable epigenetic mark, participating in numerous physiological processes. DNMT3A is of particular relevance to hematopoietic differentiation, because DNMT3A mutations are strongly related to hematopoietic malignancies. Additionally, DNMT3A deficiency has been reported to increase the hematopoietic stem cell pool by limiting their differentiation. Our previous study demonstrated that complete loss of DNMT3A resulted in anemia, while DNMT3A haploinsufficiency caused an elevated population of erythrocytes in the content of oncogenic KRAS. Since erythropoiesis is tightly regulated via the erythropoietin (EPO)-mediated RAS-RAF-MEK-ERK1/2 pathway, the question arises whether DNMT3A cooperates with RAS signaling to modulate erythropoiesis. Human leukemia cell lines were used, with differentiation capabilities towards megakaryocyte and erythroid lineages. Overexpression of DNMT3A was found to enhance erythrocytic differentiation of K562 cells, while DNMT3A knockdown suppressed differentiation. Furthermore, higher DNMT3A expression was detected in late-stage mouse erythroblasts along with the DNMT3A translocation to the nucleus. Further studies demonstrated that both ERK1/2-DNMT3A interaction and serine-255 phosphorylation in DNMT3A led to DNMT3A translocation into the nucleus, and modulated erythrocytic differentiation. Our results not only explore the critical role of DNMT3A in erythropoiesis, but also provide a new insight into ERK1/2-DNMT3A interaction in the hematopoietic system.

Dnmt3a haploinsufficiency cooperates with oncogenic Kras to promote an early-onset T-cell acute lymphoblastic leukemia.

Mutations in DNA methyltransferase 3A (DNMT3A) are prevalent in various myeloid and lymphoid malignancies. The most common DNMT3A R882 mutations inhibit methyltransferase activity of the remaining wild-type DNMT3A proteins at a heterozygous state due to their dominant-negative activity. Reports and COSMIC database analysis reveal significantly different frequencies of R882 mutations in myeloid versus T-cell malignancies, inspiring us to investigate whether downregulation of DNMT3A regulates malignancies of different lineages in a dose-dependent manner. In a competitive transplant setting, the survival of recipients with KrasG12D/+ ; Dnmt3a+/- bone marrow (BM) cells was significantly shortened than that of recipients with KrasG12D/+ cells. Moreover, all of the recipients with KrasG12D/+ ; Dnmt3a+/- cells developed a lethal T-cell acute lymphoblastic leukemia (T-ALL) without significant myeloproliferative neoplasm (MPN) phenotypes, while ~20% of recipients with KrasG12D/+ cells developed MPN with or without T-ALL. This is in sharp contrast to the recipients with KrasG12D/+ ; Dnmt3a-/- cells, in which ~60% developed a lethal myeloid malignancy (MPN or acute myeloid leukemia [AML]). Our data suggest that in the context of oncogenic Kras, loss of Dnmt3a promotes myeloid malignancies, while Dnmt3a haploinsufficiency induces T-ALL. This dose-dependent phenotype is highly consistent with the prevalence of DNMT3A R882 mutations in AML versus T-ALL in human.

Camptothecin promotes the production of nitric oxide that triggers subsequent S-nitrosoproteome-mediated signaling cascades in endothelial cells.

Camptothecin (CPT) has been used for colorectal cancer therapy. At low concentration of 10-9M, CPT modulates endothelial nitric oxide production following the phosphorylation of LKB1 Ser431, AMPK-α Thr172, eNOS Ser633 and Ser1177. Elevated nitric oxide (NO) was observed by FA-OMe fluorescent probe. 726 S-nitrosoproteins were identified by iTRAQ quantitative proteomics. IPA analysis indicated that ERK/MAPK was closely linked in the signaling network. Further studies showed that CPT phosphorylated p38 MAPK Thr180/Tyr182 and dephosphorylated Tau Ser199/202. CPT also suppressed the TNF-α-induced expression of the inflammasome and cyclooxygenase 2. All this suggests that in addition to the original character of CPT in attenuating the binding of topoisomerase I and DNA in cancer cells, the role of CPT in triggering NO production and the subsequent S-nitrosylated signaling including anti-inflammatory effects in endothelial cells are proposed here. CPT, therefore, provides a potential application addition in preventing vascular disorders.