| Chicken Marek’s Disease(MD)is an infectious tumour disease caused by Marek’s Disease Virus(MDV),which occurs widely in various places.Although vaccines are available,outbreaks can still occur when MDV virulence changes or preventive measures are not in place.Marek’s disease is also an ideal model for studying the pathogenesis of cancer diseases.In this study,the chicken MD animal model was used to validate the differential expressed new LncRNAs related to p53 expression screened in previous studies in vivo and in vitro,and to detect and analyze their serum levels in order to explore the changes and possible functions of newly discovered LncRNAs in the process of MD infection,and to provide the screening of clinical detection markers for MD serum.The transcription level of p53-related new LncRNAs in chicken spleen was analyzed by q PCR in MDV-Infected Chickens.The results showed that the transcription levels of PIG3 LncRNA,TP53PB2 LncRNA,TP53BP1 LncRNA-1,TP53BP1 LncRNA-2,TP53BP1 LncRNA-3 in spleen tissues of MDV infected chickens were significantly higher than those of the control group(P<0.01),while the transcription levels of TP53TG5 LncRNA were significantly lower(P<0.05).The results of in vivo validation analysis of the above six LncRNAs showed that the expression of p53-related LncRNAs was significantly up-regulated or down-regulated in the spleen tissues of chickens infected with MDV.This change may be related to the development of MD tumors,thus promoting the occurrence of MD tumors.We constructed TP53BP1 LncRNA-3 overexpression vector to transfect DF-1 cells.The positive cells were screened by resistance screening markers.The efficiency of transfection was detected by fluorescence markers.The overexpression level of TP53BP1 LncRNA-3 in DF-1 cells and its effect on p53 expression were detected by q PCR and Western Blot.The results showed that compared with the blank control group,the transcription level of TP53BP1 LncRNA-3 increased significantly(P<0.01);the transcription level and protein expression level of tumor-related factor p53 increased significantly(P< 0.01);the protein expression level increased significantly(P< 0.05).After MDV infection,the transcription level of TP53BP1 LncRNA-3 in serum increased significantly compared with that in normal control chickens(P< 0.05).The transcription level of TP53BP1 LncRNA-3 in spleen increased significantly(P< 0.01).The difference between the two groups was the same,but the difference between groups was different.The analysis of the transcription level of TP53BP1 LncRNA-3 in serum can reflect the changes of this index in chicken spleen to a certain extent,indicating that this index may be used as a marker for clinical detection of MDV infection.The results showed that six LncRNAs related to p53 in chicken spleen tissues were significantly up-regulated or down-regulated after MDV infection.Overexpression of TP53BP1 LncRNA-3 in DF-1 cells in vitro showed that the transcriptional levels of TP53BP1 LncRNA-3 and p53 were significantly increased,and the expression of p53 protein was significantly increased.These results suggest that the transcriptional levels of six newly discovered LncRNAs associated with tumor-related factor p53 are all affected by MDV,and their changes may affect the expression level of p53,thus promoting the occurrence of MD tumors.The expression level of TP53BP1 LncRNA-3 in serum and spleen of chickens infected with MDV showed the same trend of increasing difference between the two groups.The change of the expression level of TP53BP1 LncRNA-3 in serum could provide a reference for the screening of serum detection markers for MD tumors. |
