Involvement of BCL6 in variable splicing regulation of recurrent abortion in HTR-8 cells
Objective To construct a lentiviral overexpression of B cell lymphoma 6(BCL6)gene in the HTR-8 stable cell model for high-throughput sequencing to analyze potential variable splicing sites and pathways.Methods Lentiviral in-fection of HTR-8 cells for 1-2 weeks to obtain a stable cell line was followed by bipartite sequencing using the Illumina No-vaseq 6000 sequencing platform,and hierarchical indexing for spliced alignment of transcripts(HISAT2)was used to analyze the splicing sites of each sample.The student t-test(t-test)was utilized to filter out differentially expressed genes(DEG)and differentially alternative splicing genes(DASG)by comparing the change in variable splicing level of the same splicing type of each gene in various cell samples.DASG and DEG were improved for Kyoto Encyclopedia of Genes and Genomes(KEGG)and Gene Ontology(GO)study.Results Sequencing results showed that HTR-8 cells overexpressing the BCL6 gene had a higher proportion of variable splicing events than controls in all cases,in which intron retention occurred with significantly increased frequency.Sequencing screened a total of 880 DASG genes,and 670 DEG genes,of which a total of 32 genes were expressed,and GO enrichment showed that the biological functions were mainly protein binding,which existed with the cy-tosol.KEGG pathway was enriched in starch and sucrose metabolism.Conclusion Overexpression of the BCL6 gene acts on herpes simplex virus type Ⅰ infection and starch-sucrose metabolism pathway to increase the number of variable splicing sites to cause the occurrence of recurrent miscarriage further,BCL6 may be a potential biomarker for recurrent miscarriage.
human chorionic trophoblast cellsrecurrent miscarriagehigh-throughput sequencingBCL6
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