Abstract: Objective To explore the mutual regulation between TRIM21 and TRIM8 in glioma U251 cells. Methods TRIM21 and TRIM8 were overexpressed or knocked down respectively in U251cells, and their protein and mRNA expression levels were detected by Western blotting and RT-PCR respectively. Immunofluorescence experiment was used to analyze the subcellular localization of TRIM21 and TRIM8. The interaction between TRIM21 and TRIM8 was verified by immunoprecipitation experiment. The ubiquitination modification of TRIM21 or TRIM8 was analyzed by intracellular ubiquitination experiment. Protease inhibitor MG-132 was used to inhibit protease activity. U251 cell apoptosis was detected by flow cytometry assay. Results TRIM21 and TRIM8 could negatively regulate each other on theprotein level in U251 cells which resulted in cell apoptosis inhibition. Mechanism study showed that there was an interaction between TRIM21 and TRIM8 in U251 cells, that interaction furtherly activated ubiquitin-proteasome-dependent degradation pathway through ubiquitination. Conclusion Negatively mutualregulation between TRIM21 and TRIM8 is achieved through ubiquitination modification, following ubiquitin-proteasome-dependent degradation. Our results suggested that it is important to maintain a balance between TRIM21 and TRIM8 on protein level, that is, three might exist an E3 ubiquitin ligase protein homeostasis. The homeostasis disorder will be highly related to the tumorigenesis.

Key words: TRIM21, TRIM8, mutual regulation, ubiquitination, E3 homeostasis, glioma