Study Of The Epigenetic Mechanism Of Leukemogenesis And The Reprogrammed Hematopoietic Stem Cell

Section ?.Mechanistic study of the mutually exclusive mutations of epigenetic factors DNMT3A and MLL in acute myeloid leukemiaAcute leukemias are a group of hematological tumors that arise from malignant mutations in hematopoietic stem cells,which further advance the clonal evolution.In recent years,high-throughput genome sequencing studies have found that in acute myeloid leukemia(AML),many genes inccluding DNMT3A and MLL associated with epigenetic regulation are frequently mutated,with higher mutation rates compared with other genes.However,DNMT3A and MLL seldom mutate in the same case at the same time,showing the mutually exclusive mutation patternDNMT3A is an important DNA methyltransferase in eukaryotic cells,whereas MLL is an important histone methyltransferase that exerts a trimethylation modification on histone H3 lysine 4(H3K4me3).In AML,the mutation of DNMT3A mainly occurs in arginine at position 882(Arg882),which usually leads to the missense mutation,whereas the MLL gene dominantly undergoes translocation with the generation of various fusion proteins.As two important epigenetic regulators in cells,DNMT3 A and MLL play regulatory roles on DNA and histone proteins,respectively.What is the relationship between DNMT3A and MLL,and why they are mutually exclusive in the pathogenesis of AML,these questions are largely unknown In this work,we've found that in AML,DNMT3A mutant and MLL fusion protein both can cause the overexpression of HOX and MEIS1 genes,which is vitally important for the leukemogenesis of AML.The reason that both of DNMT3A mutant and MLL fusion protein can induce the overexpression of HOX and MEIS1 genes is probably due to the following mechanism:when DNMT3 A or MLL is wild-type,either of them is able to recruit another negative epigenetic factor,BMI1,to the promoter regions of HOX and MEIS1 genes,so that BMI1 can exert its negatively regulatory function on controlling the expression of HOX and MEIS1 genes to the normal level.When DNMT3A or MLL undergoes mutation,either DNMT3A mutant or MLL fusion protein is severely weakened for recruiting BMI1 to the promoter regions of HOX and MEIS1 genes,therefore,HOX and MEIS1 genes can be overexpressed due to the weakened negative regulation from BMI1.Since either DNMT3A mutation or MLL translocation can lead to the decreased recruitment of BMI1,even if they are co-mutated,no more decreased recruitment of BMI1 is likely to be occurred,which means no more proliferative advantage of AML cells due to the more overexpressed HOX and MEIS1 genes would happen.Therefore,seldom could we observe the co-mutations of DNMT3A and MLL in AML patients.The interpretation of this phenomenon would help us further understand the epigenetic regulation mechanism of AML,and provide new ideas for the study of other types of leukemia with such mutually exclusive mutationsSection ?.Respecifying human iPSC-derived blood cells into highly engraftable hematopoietic stem and progenitor cells with a single factorHematopoietic stem cell(HSC)is the only cell type that can differentiate into all the blood cells in human body,and HSC transplantation(HSCT)has so far been the only effective treatment for many diseases including malignant blood diseases such as leukemia.Despite of the enormous demand of HSC,the great shortage of the donor HSC has severely limited the clinical application of HSCT.As the emergence of induced pluripotent stem cell(iPSC)technology,it provides a possibility to revert the somatic cell from the patient into iPSC followed by differentiating iPSC into HSC,which may realize the allogeneic stem cell transplantation,and thereby overcome the hurdles of the shortage of donor HSC and the transplant rejection from allogeneic HSCT.When human iPSC has been obtainable since 2007,to acquire human iPSC-derived transplantable HSC has been a hot area of research for the last decade,however,with unexpected challenges.After a decade,there are only few reports that have achieved iPSC-derived hematopoietic stem and progenitor cell(iPSC-HSPC)with engraftability,however,most of these iPSC-HSPCs lack the ability to re-establish lymphopoiesis or long-term hematopoiesis in vivo,or are generated by highly complicated procedures with very low efficiency.We speculate that by adding transcription factor that can simultaneously improve the self-renewal ability and lymphopoietic differentiation potential into iPSC-HSPC,it may be possible to correct the myeloid differentiation bias of iPSC-HSPC in vivo and realize the long-term engraftment.Our investigation reveals that by adding a single factor,MLL-AF4,into iPSC-derived blood cells can revert these blood cells into highly engraftable HSPC with unbiased differentiation ability in vivo.This method is highly simple and effective,which only requires the transient expression of MLL-AF4 in a non-integrative way.In addition,we also conduct the same MLL-AF4 induction experiment in primary HSPCs in order to parallelly compare the induced iPSC-HSPCs with the primary HSPCs.The results reveal that iPSC-HSPCs are more prone to leukemic transformation during the long-term engraftment period,while primary HSPCs with the same induction can sustain the long-term engraftment without leukemic transformation.These findings demonstrate the feasibility of activating the HSC network in human iPSC-derived blood cells through expression of a single factor and suggest iPSC-HSPCs are more genomically instable than primary HSPCs,which merits further attention.