BPIFB3 Regulates Endoplasmic Reticulum Morphology To Facilitate Flavivirus Replication

Abstract

Flaviviruses, including dengue virus (DENV) and Zika virus (ZIKV), rely heavily on the availability of endoplasmic reticulum (ER) membranes throughout their life cycle, and degradation of ER membranes restricts flavivirus replication. Accordingly, DENV and ZIKV restrict ER turnover by protease-mediated cleavage of reticulophagy regulator 1 (RETREG1), also known as FAM134B, an autophagy receptor responsible for targeted ER sheet degradation. Given that the induction of autophagy may play an important role in flavivirus replication, the antiviral role of RETREG1 suggests that specialized autophagic pathways may have differential effects on the flavivirus life cycle. We previously identified BPI fold-containing family B member 3 (BPIFB3) as a regulator of autophagy that negatively controls enterovirus replication. Here, we show that in contrast to enteroviruses, BPIFB3 functions as a positive regulator of DENV and ZIKV infection and that its RNA interference-mediated silencing inhibits the formation of viral replication organelles. Mechanistically, we show that depletion of BPIFB3 enhances RETREG1-dependent reticulophagy, leading to enhanced ER turnover and the suppression of viral replication. Consistent with this, the antiviral effects of BPIFB3 depletion can be reversed by RETREG1 silencing, suggesting a specific role for BPIFB3 in regulating ER turnover. These studies define BPIFB3 as a required host factor for both DENV and ZIKV replication and further contribute to our understanding of the requirements for autophagy during flavivirus infection. IMPORTANCE Flaviviruses and other arthropod-transmitted viruses represent a widespread global health problem, with limited treatment options currently available. Thus, a better understanding of the cellular requirements for their infection is needed. Both DENV and ZIKV rely on expansion of the endoplasmic reticulum (ER) and the induction of autophagy to establish productive infections. However, little is known regarding the interplay between the requirements for autophagy initiation during infection and the mechanisms used by these viruses to avoid clearance through the autophagic pathway. Our study highlights the importance of the host factor BPIFB3 in regulating flavivirus replication and further confirms that the RETREG1-dependent reticulophagy pathway is antiviral to both DENV and ZIKV.

BPIFB3 interacts with ARFGAP1 and TMED9 to regulate non-canonical autophagy and RNA virus infection

“…Given that BPIFB3 regulates a non-canonical autophagy pathway, we sought to characterize which components of the autophagic initiation machinery were required for the induction of autophagy during BPIFB3 depletion. To do this, we used two distinct viruses, CVB and DENV, as read outs for the reversal of autophagy induction given that our previous studies showed that BPIFB3-induced autophagy increases the replication of CVB, while it restricts DENV infection (Delorme-Axford et al, 2014;Evans et al, 2020). We tested components associated with three key phases of autophagy induction-autophagosome nucleation (ULK1), isolation membrane formation (the catalytic component of PI3K (PI3KC3), UVRAG1, and BECN1), and processing of LC3 (ATG7).…”

“…This regulation of autophagy has been demonstrated to have important implications in regulating the infection of two distinct families of viruses, enteroviruses and flaviviruses. Depletion of BPIFB3 specifically enhances the replication capacity of the enterovirus CVB, while restricting the replication of the two flaviviruses DENV and ZIKV (Delorme-Axford et al, 2014;Evans et al, 2020). These disparate impacts on replication highlight key replication differences between the two families of viruses and their sensitivity to turnover by BPIFB3-regulated autophagy.…”

“…Previously, we reported that an ER-localized protein, bactericidal/permeability-increasing protein (BPI) fold-containing family B, member 3 (BPIFB3), regulates a non-canonical form of autophagy to control RNA virus infection, including members of the enterovirus and flavivirus families (Delorme-Axford et al, 2014;Evans et al, 2020). BPIFB3 belongs to the BPIFB family of proteins named for their homology to BPI, a secreted antimicrobial protein.…”

“…Despite the high degree of predicted structural homology, BPIFB3 and other ER-localized members of the family, BPIFB2 and BPIFB6, are not secreted and remain associated with the cytoplasmic face of the ER, despite lacking transmembrane domains (Delorme-Axford et al, 2014). Furthermore, our prior studies have determined that both BPIFB3 and BPIFB6 play important roles in vesicle trafficking and regulation of both enteroviruses and flaviviruses, including coxsackievirus B (CVB), dengue virus (DENV), and Zika virus (ZIKV) (Delorme-Axford et al, 2014;Evans et al, 2020;Morosky et al, 2016).…”

“…These disparate impacts on replication highlight key replication differences between the two families of viruses and their sensitivity to turnover by BPIFB3-regulated autophagy. Recently, we demonstrated that depletion of BPIFB3 not only impacts autophagic flux, but also disrupts ER integrity, which can be reversed by inhibition of ER turnover via RETREG1-dependent reticulophagy (Evans et al, 2020), suggesting that the depletion of BPIFB3 broadly affects multiple aspects of autophagy.…”