Identification Of ADAR2 Splicing Variants Expression In Glioma Tissues And The GluA2 Q/R Site RNA Editing Mediated By ADAR2 Isoforms

Objective: To test the A-to-I RNA editing level of Glu A2 subunit Q/R site in glioma tissues. To identify the alternative splicing site of ADAR2 m RNA, and the expression features of ADAR2 splicing variants. To investigate the potential relationship between Q/R site RNA editing, expression of ADAR2 and its splicing variants and glioma recurrence, peritumoral edema and RNA editing rescure.Method: 30 frozen tissues of gliomas and other 3 of white matter were randomly selected in the tissue bank of China-Japan Union Hospital, which the storage time ranged from 2011 to 2015. Isolating Total RNA, RT-PCR, sequencing the PCR product, then test the A-to-I RNA editing level of Glu A2 Q/R site. Identify the alternative splicing site of ADAR2 m RNA by RT-PCR and sequencing. After that, basing on the identified splicing site, we propagate the specific splicing variants of ADAR2, and analysis the expression properties. Analysis the potential correlations between Q/R site RNA editing, ADAR2 splicing variants expression properties with RNA editing rescure and peritumoral edema.Result: RNA editing level of Glu A2 Q/R site is decreased significantly in primary malignant gliomas compared to white matter controls, without noticeable change of ADAR2 m RNA. Three alternative splicing site in ADAR2 m RNA was identified: insertion of exion1 a, 5a and exon2 was skipped. Real time PCR data indicates that the ratio of ASV(1a+)/ASV(1a-)、ASV(2-)/ASV(2+) and ASV(5a+)/ASV(5a-) in glioma tissues was not changing significantly compared to white matter controls. But in 56.67% of glioma tissues, the ratio of ASV(5a+)/ASV(5a-) was up regulated than the average level of white matter controls. Compared with primary gliomas, the editing level of recurrent gliomas was not significantly changed, so as ADAR2 and its splicing variants. The rescure of Q/R site RNA editing coupled with up regulation of ADAR2 expression but the ratios of ASVs is as normal. The editing level of sever peritumoral edema gliomas is lower than the no convincing and mild edema group(P<0.05). Spearman correlation analysis show the PTE grade of gliomas with the Q/R site editing level of Glu A2 subunit and the ADAR2 m RNA was negative correlation, though there was no noticed change of ADAR2 and ASVs ratios between different PTE groups. There were three alternative splicing site of ADAR2 m RNA.Conclusion: The A-to-I RNA editing level of Glu A2 Q/R site was decreased significantly in primary WHO III~IV gliomas. PTE grade of gliomas with the Q/R site editing level of Glu A2 subunit and the ADAR2 m RNA was negative correlation. RNA editing rescure coupled with up regulation of ADAR2 m RNA.