Heat shock proteins stimulate APOBEC-3-mediated cytidine deamination in the hepatitis B virus
Zhigang Chen1, Thomas L. Eggerman2, Alexander V. Bocharov1, Irina N. Baranova1, Tatyana G. Vishnyakova1, Roger Kurlander1 and Amy P. Patterson3*
1 Clinical Center, National Institutes of Health, United States;
2 National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, United States;
3 National Heart, Lung and Blood Institute, National Institutes of Health, United States
Author contributions: ZC, TLE, and APP designed the study and wrote the paper. AVB, INB, TGV, and RK provided technical assistance, data analyses and consulting. All authors reviewed the results and approved the final version of the manuscript.
Abstract
Apolipoprotein B mRNA-editing enzyme catalytic subunit 3 (APOBEC-3) enzymes are cytidine deaminases that are broadly and constitutively expressed. They are often upregulated during carcinogenesis and candidate genes for causing the major single-base substitution in cancer-associated DNA mutations. Moreover, APOBEC-3s are involved in host innate immunity against many viruses. However, how APOBEC-3 mutational activity is regulated in normal and pathological conditions remains largely unknown. Heat shock protein levels are often elevated in both carcinogenesis and viral infection and are associated with DNA mutations. Here, using mutational analyses of hepatitis B virus (HBV), we found that Hsp90 stimulates deamination activity of APOBEC3G (A3G), A3B, and A3C during co-expression in human liver HepG2 cells. Hsp90 directly stimulated A3G deamination activity when the purified proteins were used in in vitro reactions. Hsp40, 60, and 70 also had variable stimulatory effects in the cellular assay, but not in vitro. Sequencing analyses further demonstrated that Hsp90 increased both A3G cytosine mutation efficiency on HBV DNA and total HBV mutation frequency. In addition, Hsp90 shifted A3G's cytosine region selection in HBV DNA and increased A3G's 5-end nucleoside preference for deoxycytidine (5'CC). Furthermore, the Hsp90 inhibitor 17-AAG dose dependently inhibited A3G and A3B mutational activity on HBV viral DNA. Hsp90 knockdown by siRNA or by Hsp90 active-site mutation also decreased A3G activity. These results indicate that heat shock proteins, in particular Hsp90, stimulate APOBEC3-mediated DNA deamination activity, suggesting a potential physiological role in carcinogenesis and viral innate immunity.
cancer cytidine deaminase heat shock protein 90 (Hsp90) hepatitis B virus (HBV, Hep B) mutagenesis APOBEC3G cofactor
Received September 26, 2016.
Accepted June 21, 2017.