Ca2+/Calmodulin-Dependent Protein Kinase II Inhibits Hepatitis B Virus Replication from cccDNA via AMPK Activation and AKT/mTOR Suppression.

Jumi Kim,Jaesung Jung,Sun Park,Jin Lim,Kyongmin Kim,Fadia Kalsoom,Hyun Lee,Yong-Joon Chwae,Hyeonjoong Kwon,Muhammad Sajjad,Ho-Joon Shin

doi:10.3390/microorganisms10030498

Jumi Kim, Jaesung Jung + Show 9 more

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https://doi.org/10.3390/microorganisms10030498

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Abstract

Ca2+/calmodulin-dependent protein kinase II (CaMKII), which is involved in the calcium signaling pathway, is an important regulator of cancer cell proliferation, motility, growth, and metastasis. The effects of CaMKII on hepatitis B virus (HBV) replication have never been evaluated. Here, we found that phosphorylated, active CaMKII is reduced during HBV replication. Similar to other members of the AMPK/AKT/mTOR signaling pathway associated with HBV replication, CaMKII, which is associated with this pathway, was found to be a novel regulator of HBV replication. Overexpression of CaMKII reduced the expression of covalently closed circular DNA (cccDNA), HBV RNAs, and replicative intermediate (RI) DNAs while activating AMPK and inhibiting the AKT/mTOR signaling pathway. Findings in HBx-deficient mutant-transfected HepG2 cells showed that the CaMKII-mediated AMPK/AKT/mTOR signaling pathway was independent of HBx. Moreover, AMPK overexpression reduced HBV cccDNA, RNAs, and RI DNAs through CaMKII activation. Although AMPK acts downstream of CaMKII, AMPK overexpression altered CaMKII phosphorylation, suggesting that CaMKII and AMPK form a positive feedback loop. These results demonstrate that HBV replication suppresses CaMKII activity, and that CaMKII upregulation suppresses HBV replication from cccDNA via AMPK and the AKT/mTOR signaling pathway. Thus, activation or overexpression of CaMKII may be a new therapeutic target against HBV infection.

Highlights

Transient transfection of 1.3 mer WT hepatitis B virus (HBV) into Huh7 or HepG2 cells resulted in lower levels of activated calmodulin-dependent protein kinase II (CaMKII), as shown by T286 phosphorylation relative to control cells transfected with empty vector (Figure 1A,B, top panel, lane 1 vs. 2)
These results indicate that HBV replication inhibits CaMKII activation
Upon CaMKII α overexpression, phosphorylation of AMPK was increased and phosphorylation of AKT/mTOR was decreased, resulting in the downregulation of HBV DNA synthesis (Figure 4D, lane 3 vs. 4, and Figure S3B). These results demonstrate that CaMKII α overexpression represses HBV replication by activating AMPK and inhibiting the AKT/mTOR signaling pathway, indicating that CaMKII α is an upstream protein in the AMPK/AKT/mTOR

Summary

Introduction

Hepatitis B virus (HBV), a prototype virus of the Hepadnaviridae family, contains a small, partially double-stranded, relaxed 3.2 kb circular (RC) DNA [1]. An effective vaccine is available, HBV infection remains a public health concern. B (CHB) infection, which has been estimated to affect 257 million individuals worldwide (WHO), is closely associated with cirrhosis and hepatocellular carcinoma (HCC) [2]. HBV infection of a cell, its RC DNA is translocated to the nucleus and converted into a covalently closed circular DNA (cccDNA), which serves as the template for transcription 4.0/).

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