Study On Pathogenic Mechanism Of Susceptibility Gene LCE3D And The Lysine Crotonylation In Psoriasis

Background Psoriasis is a common chronic relapsing immune skin disease and a polygenic hereditary skin disease.The exact cause of psoriasis is unclear.According to the pathogenesis,psoriasis susceptibility genes can be mainly divided into two categories.One is immune-related genes such as HLA,IL-12 B,IL-23 R and NFKB.The second is genes associated with skin barrier function such as LCE3,S100 and so on.At present,multiple teams have found that LCE3 is a susceptibility gene for psoriasis in multiple ethnic groups.The genetic loci in this region are associated with psoriasis in a variety of genetic ways,and these loci are in a linkage disequilibrium.These studies suggest that the LCE3 may have an important role in the pathogenesis of psoriasis.LCE3(Late Cornified Envelope)has been discovered as an important susceptibility gene for psoriasis for 10 years.LCE has a complex gene cluster,and it is still unclear which gene cluster has a major role in the pathogenesis of psoriasis.The objective reason is that the gene homology between the LCE3 subfamilies is extremely high.For example,the gene sequence similarity of LCE3 D and LCE3 E is 97%,and these homologous genes may have functional complementarity.After the loss of a certain gene function,other genes may be complementarily executed in terms of protein function because of their high degree of similarity.Therefore,the function of a single gene cannot be expressed after deletion,so it is difficult to perform functional studies on these genes separately.In this study,in combination with the above factors,a method of transfecting LCE3 D into the skin by an adenovirus was used to effectively proliferate the expression of the target gene.By simulating the rapid increase of LCE3 D levels in psoriasis tissue,the related gene functions were studied,and the possible roles and mechanisms of LCE3 D in the pathogenesis of psoriasis were discussed.Method To explore the mechanism of action of LCE in psoriasis.We first performed expression profiling on clinical samples,animal models,and human keratinocyte lines.The gene expression patterns of the LCE gene family in the psoriasis model and the control model were analyzed.Through the alignment analysis of these data,it was found that LCE3 D is the most relevant gene for psoriasis.Therefore,we constructed an overexpressed LCE3 D adenovirus model to perform functional verification and exploration of genes at the animal and cellular levels.Results Gene expression analysis results of clinical samples,mouse models,and cell models,and differential expression of LCE gene family expression in psoriasis samples and psoriasis models.It is indicated that the LCE3 gene cluster has the largest expression difference and significance in the psoriasis samples.Significant differences in gene expression are positively correlated with the psoriasis.In addition,LCE3 D is the most significant gene in the LCE3 gene cluster,indicating that LCE3 D may play an important role in psoriasis.LCE1 protein is distributed in all layers of the epidermis and may belong to the basal expression protein,which is used to maintain the normal growth of keratinocytes.However,when stress is present,LCE1 expression is decreased and function is weakened.LCE2 is expressed in the stratum corneum.The stratum corneum is the last layer of keratinocyte differentiation.This layer of cells has no cell morphology,mainly protective physical effects,and it is difficult to regulate the body's response through the biological activity of cells.LCE3(D)is mainly expressed in the spinous layer and the granular layer,which are the most abundant cellular components in the epidermis,which can secrete many metabolites and substances that regulate self-keratinization.We found that overexpression of LCE3 D promotes increased levels of keratinocyte synthesis,including the synthesis of t RNA and the generation of a series of complexes that promote protein translation,and the addition of subcellular organelles to provide the basic equipment needed for new cell formation.On the other hand,the expression of this gene also promotes the process of keratinization,allowing newly formed(basal layer)cells to enter the differentiation stage as soon as possible,and promote the increase of acanthal and granular cells.In animal models,we found that LCE3 D overexpression promoted skin thickening in mice and made mice more sensitive to the induction of imiquimod.There was a significant difference in the PASI scores of the mice compared to the control empty group at the time of transfection for three days.Conclusion Clinical samples,animal models and cell models have demonstrated that LCE3 subfamily gene expression is positively correlated with psoriasis,and LCE3 D gene expression is most strongly associated with psoriasis.LCE3 D is mainly expressed in the spinous layer and the granular layer,which promotes translation and energy synthesis and accelerates keratinocyte differentiation.LCE3 D is an important stress response gene.The LCE3 D gene rapidly responds to internal and external stress and increases expression,promotes keratinocyte proliferation and rapidly differentiates,causing skin thickening and increased scales.Background Psoriasis is a chronic immunoinflammatory dermatosis characterized by erythema and scaly,which is unclear.It is easy to relapse and is considered to be a multifactorial complex disease.Genome-wide association analysis has identified more than 80 Genegenic loci,but the genetic end of psoriasis is only 28%,only explain some of the pathogenic factors,DNA will eventually be translated into proteins or regulate the expression of other proteins to perform functions,and ultimately function The protein,many protein modifications such as phosphorylation,methylation,acetylation and ubiquitination,undergo functional changes,so protein modification omics research is very meaningful,in recent years,with high-resolution protein mass spectrometry The innovation has greatly promoted the research of post-translational modification of proteins.Researchers have successively identified many new PTMs modifications.Among them,Lysine Crotonylation(Kcr)is a kind of modification that has received much attention.This modification usually occurs in histones in the active chromatin region,and Kcr modification is closely related to reproductive regulation and is involved in the regulation of cell developmental genes.Several biomarkers for the diagnosis of psoriasis have been identified by proteomic analysis.Although their role in disease is unknown,non-histone lysine crotonylation has been shown to be widespread.Objective To analyze the Kcr proteomics of 45 cases of psoriasis lesions and adjacent non-lesional lesions by high-resolution protein mass spectrometry,explore the Kcr map of psoriasis protein,and study the protein Kcr modification,Whether it has a role in the pathogenesis of psoriasis.Methods After processing the skin tissue,extract the protein,isolate the peptide,add the TMT labeled quantitative protein group to the peptide mixture,analyze the peptide by high performance liquid chromatography,and carry out affinity enrichment of lysine crotonylation,and generate by mass spectrometry.MS spectra,the mass spectrometry results were retrieved in the database and subjected to biological analysis to screen for sites with significant differences in modification.In this study,the Kcr locus quantitative ratio(cutaneous lesions/non-lesional lesions)was up-regulated by >1.2,and the quantitative <0.38 was the down-regulated standard.After bioinformatics analysis,the sites with significant differences were screened out and existing.Proteomics conducts correlation analysis,speculating on the role of protein Kcr modification in the pathogenesis of psoriasis,supplementing the interpretation of the pathogenesis of psoriasis.Results We studied a total of 90 samples of 45 psoriasis lesions and their adjacent normal skin tissues,and conducted experiments in three groups.During the three replicate experiments,the entire proteome and lysine were recorded.Dynamic changes in acid crotonylation.Reproducibility analysis showed that the three replicates were consistent.A total of 3686 and 3008 proteins were identified and quantified in the lesions,of which 102 proteins were up-regulated and 124 were down-regulated.The expression of SART 1(P=3.55×10-5)and recombinant human glycol ester transporter(GLTP)(P=1.54×10-3)was the most obvious down-regulation and up-regulation,respectively.Nearly 90% of these differentially regulated proteins exhibit the same expression trends as the differential proteins in the online RNA sequencing data set for psoriasis.19 different regulatory proteins were negatively correlated with DNA methylation data of psoriatic lesions.In summary,1703 of the 934 and 690 proteins and 2080 lysine crotonylation sites were identified and quantified,respectively.33 of the 31 and 29 proteins were up-and down-regulated,respectively.A 88.0% differentially regulated lysine crotonylation site was negatively correlated with protein expression.These differentially expressed proteins are enriched in immune responses,ribosomes,antigen processing and presentation,and the IL-17 signaling pathway.Conclusions This study is the first comprehensive analysis of proteomics and lysine crotonylation of psoriasis.The results indicate that changes in the proteome and lysine crotonylation are involved in the process of psoriasis play an important regulatory role.