The Prediction Of CTCF Binding Sites In Herpes Simplex Virus Type Ⅱ And The Study Of Its Function In Expression Regulation

Herpes Simplex Virus is the main pathogen of herpes symptoms,including two types,Herpes Simplex Virus Ⅰ(HSV-1)and Herpes Simplex Virus Ⅱ(HSV-2),and HSV-2 is the main pathogen of genital herpes.HSV-2 establishes a lifelong latency infection within sensory neurons after initial infection.Latency-assistant transcript(LAT)is the only transcript can be found in latent infection.CTCF(CTCF-binding factor)is a multivalent eukaryotic transcription factor,binding on multiple target genes and expressing different functions.It has demonstrated that HSV-1 genome contain clustered CTCF binding motifs,and the latency or reactivation of HSV-2 is related with CTCF binding or disruption.Because of the high homology between HSV-1 and HSV-2 and the difference in LAT region,whether CTCF is also involved in the HSV-2 regulation of latency-activation need to be researched.The objective of this project is to find the function of CTCF-binding motifs in HSV-2 latency-reactivation regulation.Firstly,bioinformatics analysis was taken to analyze HSV-2 whole genome structure,and a contrast was taken between HSV-1 and HSV-2.Bioinformatics analysis and Jaspar analysis coupled with sequence analysis were used to find the CTCF-binding motifs.Then,the predicted binding motifs were amplified and embedded in p GL3-promoter plasmids,and dual luciferase reporter assays were taken place to identified the functions of these motifs.In this project,we predicted three CTCF-binding motifs located in LAT upstream(CTUL),downstream(CTa’m)and intron(CTRL),respectively.Luciferase reporter assays revealed that LAT intron(p GL3-promoter-CTRL)and LAT downstream(p GL3-promoter-CTa’m)recombinant plasmids expressed lower luciferase activities than p GL3-promoter(P<0.001),While,the p GL3-promoter-CTUL recombinant plasmids group had no statistically significant difference compared with the p GL3-promoter group(P>0.05).Our project predicted three CTCF-binding motifs,located in HSV-2 LAT upstream,downstream and intron region,and LAT downstream and intron predicted binding motifs may a promoter-blocking activity.In this study,we provided a preliminary prediction of CTCF binding motifs,and gave an expression regulation research in predicted motifs.Our study fills the CTCF research vacancy in HSV-2,and finds a new direction to research the mechanism of HSV-2 latency.