Transcriptional Regulation Of ?-globin Gene Expression By LYAR And Epigenetic Regulation Of Slug Gene Expression By NATD

Hemoglobins play an important role in oxygen transport in the body.In adults,a hemoglobin molecule is composed of two a-globin subunits and two ?-globin subunits.Mutations in(3-globin gene are associated with the hemoglobinopathies,such as sickle-cell disease and ?-thalassemia,which are two of the most important single gene genetic diseases.Therefore,studies of these diseases and related mechanisms are of great importance.Elevated level of ?-globin to compensate for the loss of ?-globin has been proved to be an effective strategy for the treatment.Thus,it is very important to study the mechanisms involved in ?-globin gene regulation.In the first part of the thesis,we discovered a novel role of a nuclear protein,LYAR,in ?-globin gene transcriptional regulation which may provide a new prospective therapeutic target for sickle-cell disease and ?-thalassemia treatment.Human globin gene expression during development is regulated by transcription factors in a stage-dependent manner.However,the mechanisms controlling the process are still largely unknown.In our previous study,we found that PRMT5 played an important role in ?-globin gene silencing.Here we identified the interaction between PRMT5 and LYAR.Furthermore,we found that the binding of PRMT5 on ?-globin promoter is LYAR-dependent.The results of CASTing(Cyclic Amplification and Selection of Targets)and EMSA assay confirmed that LYAR directly binds a consensus sequence,5'-GGTTAT-3',in the DNA region corresponding to the 5'UTR of ?-globin gene.In addition,?-globin gene can be reactivated in both K562 cells and human primary erythroid progenitor cells when LYAR was knocked down in these cells using the shRNA approach.These findings suggest that LYAR functions as a novel transcriptional repressor of ?-globin gene expression.Gene activity not only can be regulated transcriptionally,but also can be controlled under epigenetic mechanisms.Epigenetics is the study of heritable changes in gene activity which are not caused by changes in the DNA sequences.Over the]ast decade,epigenetics has developed very rapidly.A comprehensive study of epigenetic mechanisms has helped us develop new therapeutic strategies toward various diseases including cancer.Cancer is one of the major threats to human health,and metastasis is the most important contributor to the mortality of cancer patients.Tumor metastasis consists of a series of discrete biological processes.Epithelial-mesenchymal transition(EMT)is one of the most important pathways involved in cancer progression and metastasis.In the second part of my thesis,we focused on the epigenetic role of N-?-acetyltransferase,NATD,in controlling Slug gene expression and related EMT process.N-?-acetylation is catalyzed by N-a-acetyltransferases(NATs).NATD is responsible for N-?-acetylation of histone H4.However,very little is known about the biological function of NATD.Here we found that NATD mRNA and protein levels have significantly increased in lung cancer specimens compared with those in normal tissues.The migration capabilities of lung cancer H1299 cells were significantly decreased when NATD expression was reduced.We also showed that expression of Slug,an important regulator in EMT pathway,was significantly reduced in NATD knockdown H1299 cells.We found that NATD was required for the maintenance of associate histone markers on Slug promoter.Together,our data indicate that NATD is a novel key epigenetic regulator of Slug gene expression,and thus mediates EMT during lung cancer development.