The Roles Of Linker Histone H1.2 And Histone Demethylase KDM6A In Non-small Cell Lung Cancer

Purpose: Non-small cell lung cancer(NSCLC),which accounts for 85% of lung cancer cases,is a serious malignance with high incidence and mortality.A large number of NSCLC patients face the lack of effective drug therapies.Thus,it is of great importance to study the underneath pathogenesis mechanisms.Linker histone H1 and core histone are important components of chromatin;they can influence the structure of chromatin,and regulate gene transcription via epigenetic mechanisms with DNA sequence unchanged.H1.2,a most common H1 variant,is involved in cell cycle,apoptosis and other cell life processes,and affects the proliferation and invasion of various tumors.However,the role of H1.2 in lung cancer is not clear.In addition,a histone demethylase KDM6 A,which regulates the methylation of core histone H3,plays as either tumor suppressor or oncogene in cancers;yet its role in NSCLC remains elusive.Most of the known mechanisms focus on how KDM6 A regulates downstream target gene transcription,and the information about the upstream network of KDM6 A is limited.The aim of this study is to investigate the role of H1.2 and KDM6 A,two molecules with epigenetic regulation functions,in NSCLC and their specific molecular mechanisms.Methods: Immunoblotting were applied to detect the levels of H1.2 and KDM6 A in clinical samples from lung cancer patients.H1.2 or KDM6 A were knocked out,knocked down or overexpressed in multiple lung cancer cell lines to investigate phenotypic changes in cell proliferation and invasion.In the studies on H1.2,transgenic mouse models for primary lung cancer was used for phenotype investion and transcriptome sequencing analysis;antioxidant capacity of cells was assessed by detecting reactive oxygen species and glutathione levels.In the studies on KDM6 A,multiple small molecule inhibitors were applied to explore the association between epidermal growth factor receptor(EGFR)pathways and KDM6 A.In addition,the association of H1.2 and KDM6 A with prognosis in lung cancer was analyzed using online databases.Results: In the first part of the Results,the expression level of H1.2 was significantly higher in tumor tissues than in paratumor tissues,and NSCLC patients with high H1.2 level comparing to those with low H1.2 level showed worse prognosis.H1.2 knock out inhibited malignant proliferation of lung cancer in NSCLC cells and mice.Furthermore,H1.2 could bind with Nrf2,an important nuclear transcription factor mediating antioxidation reaction.Upon H1.2 knock out,the level of Nrf2 droped in nuclei,the transcription level of its downstream target gene GCLC(one of the speed-limiting enzymes of glutathione synthesis)decreased,glutathione synthesis was down-regulated,suggesting impaired antioxidant capacity.In the second port of the Results,KDM6 A levels in clinical samples were positively correlated with the phosphorylation of EGFR.Activation of EGFR phophorylates its downstream transcriptional regulator STAT3 to up-regulate m RNA and protein levels of KDM6 A.Moreover,knockdown of KDM6 A or inhibition of its enzyme activity could inhibit the proliferation of NSCLC cells.Conclusion: In general,the two parts of this paper are based on the detection results of clinical samples from NSCLC patients.On the one hand,based on the high expression of H1.2 in tumors and its association with worse prognosis,we found that H1.2 promotes the malignant proliferation of NSCLC via regulating antioxidant systems.On the other hand,based on the positive correlation between KDM6 A and EGFR phosphorylation level in tumors,we found that KDM6 A was transcriptionally regulated by the activation of EGFR and STAT3,and its tumor-promoting effect was dependent on enzyme activity,suggesting the potential of KDM6 A to be a therapeutic target for certein types of lung cancer.Innovation points: These findings demonstrate the oncogenic roles of H1.2 and KDM6 A in lung cancer,providing new information for the pathogenesis of NSCLC,a new theoretical basis for the further development of epigenetic drugs,and a new perspective for the crosstalk of epigenetic regulation and other pathways in cancer.