Molecular mechanism of apoptosis-inducing factor regulated by FBXO22 and its role in cancer cell apoptosis

doi:10.3969/j.issn.1674-8115.2021.04.003

Abstract

Abstract: Objective

·To explore the substrate of F-box only protein 22 (FBXO22) and its mechanism, and further study the role of FBXO22 in cancer cell apoptosis.

Methods

·The interaction protein of FBXO22, namely apoptosis-inducing factor (AIF), was searched and identified by immunoprecipitation (IP)-coupled liquid chromatograph mass spectrometer/mass spectrometer (LC-MS/MS) technology. The effect of FBXO22 on ubiquitination of AIF was detected by ubiquitin modification assay under denatured conditions. Short hairpin RNA (shRNA) knockdown FBXO22 and overexpression FBXO22 were used to detect the expression of AIF. The effect of FBXO22 knockdown or overexpression on apoptosis of colon cancer cells was detected by flow cytometry.

Results

·IP-coupled LC-MS/MS technology showed that FBXO22 could interact with AIF, and ubiquitin modification assay under denaturing conditions showed that FBXO22 could mediate ubiquitination of AIF. Knockdown of FBXO22 could up-regulate the level of AIF, while overexpression of FBXO22 could down-regulate the level of AIF. The results of flow cytometry showed that knockdown of FBXO22 could promote the apoptosis of colon cancer cells induced by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), while overexpression of FBXO22 could inhibit the apoptosis of colon cancer cells induced by MNNG.

Conclusion

· FBXO22 regulates AIF-mediated apoptosis of colon cancer cells by ubiquitination and degradation of AIF.

Key words: F-box only protein 22 (FBOX22), apoptosis-inducing factor (AIF), ubiquitination modification, cell apoptosis

CLC Number:

R363.1⁺4

Meng-di LIU, Yi-lian PAN, Xiao-na ZHU, Shuo YANG, Qian YANG, Yun YU. Molecular mechanism of apoptosis-inducing factor regulated by FBXO22 and its role in cancer cell apoptosis[J]. JOURNAL OF SHANGHAI JIAOTONG UNIVERSITY (MEDICAL SCIENCE), 2021, 41(4): 427-433.

Figures/Tables 5

Fig 1 Immunoprecipitation complexes of the two groups detected by SDS-PAGE and Coomassie bright blue staining

Tab 1 Different protein peptide number of the two groups detected by IP/MS technology

Gene	Protein Name	Peptide (FBXO22 group/Control group)
FBXO22	F-box only protein 22	933/1
SKP1	S-phase kinase-associated protein 1	327/0
CUL1	Cullin-1	76/0
RBX1	E3 ubiquitin-protein ligase RBX1	11/0
AIF	Apoptosis-inducing factor 1	9/0

Fig 2 IP detection of the interaction between FBXO22 and AIF and its effect on the ubiquitination level of AIF

Fig 3 Effect of FBXO22 on AIF level by Western blotting

Fig 4 Effect of FBXO22 on cancer cell apoptosis mediated by AIF

TrendMD

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