The Host Protein Reticulon 3.1A Is Utilized by Flaviviruses to Facilitate Membrane Remodelling

Abstract

Flaviviruses are enveloped, positive-sensed single-stranded RNA viruses that remodel host membranes, incorporating both viral and host factors facilitating viral replication. In this study, we identified a key role for the membrane-bending host protein Reticulon 3.1 (RTN3.1A) during the replication cycle of three flaviviruses: West Nile virus (WNV), Dengue virus (DENV), and Zika virus (ZIKV). We observed that, during infection, RTN3.1A is redistributed and recruited to the viral replication complex, a recruitment facilitated via the WNV NS4A protein, however, not DENV or ZIKV NS4A. Critically, small interfering RNA (siRNA)-mediated knockdown of RTN3.1A expression attenuated WNV, DENV, and ZIKV replication and severely affected the stability and abundance of the NS4A protein, coinciding with a significant alternation and reduction of viral membrane structures in the endoplasmic reticulum. These observations identified a crucial role of RTN3.1A for the viral remodelling of host membranes during efficient flavivirus replication and the stabilization of viral proteins within the endoplasmic reticulum.

Shaping the flavivirus replication complex: It is curvaceous!

“…Viral proteins interact with DNAJC14, which acts as a chaperone to modulate VP formation (Yi et al, , Yi et al, ), potentially on cholesterol‐rich microdomains (Mackenzie et al, ). We speculate that the viral protein NS4A (based on its predicted topology, structure, and membrane remodelling capacity [Roosendaal et al, , Miller et al, ]) induces membrane curvature while interacting with the host RTN3.1A protein (Aktepe et al, ). We suggest that the biogenesis and recruitment of the cone‐shaped lipids ceramide (Aktepe et al, ) and lyso‐PChol (Liebscher et al, ) stabilises and provides the required curvature for the formation of the VP.…”

“…CRISPR and RNA interference screens have identified key host proteins in the replication of many flaviviruses; however, very few studies have subsequently investigated the role of identified proteins in the biogenesis and stability of the viral RC. The reticulon (RTN) protein is the most extensively studied host membrane‐shaping protein during (+)RNA viruses, including WNV and DENV (Aktepe, Liebscher, Prier, Simmons, & Mackenzie, ).…”

“…Recent studies have demonstrated the importance of RTN 3.1A during flavivirus replication. During flavivirus (WNV, DENV, and ZIKV) infection, RTN3.1A is required to induce (WNV and ZIKV) or preserve (DENV) RC architecture by directly (WNV) or indirectly (DENV and ZIKV) interacting with the viral NS4A protein (Aktepe et al, ). The abolishment of this interaction by silencing RTN3.1A destabilises and promotes the degradation of NS4A, which further affects the aptitude of these viruses to remodel intracellular membranes.…”

“…The abolishment of this interaction by silencing RTN3.1A destabilises and promotes the degradation of NS4A, which further affects the aptitude of these viruses to remodel intracellular membranes. This translates into a significant attenuation in viral translation and the generation of viral infectious particles (Aktepe et al, ). Together, these studies demonstrate a novel mechanism by which BMV, E71, and flaviviruses remodel cellular membranes by recruiting host proteins to the RCs.…”

Endoplasmic reticulum: a focal point of Zika virus infection

“…Cleaved capsid protein interacts with capsid-dense lipid droplets and viral RNA to generate nucleocapsid core followed by Env and prM proteins encapsulation of the nucleocapsid core Invagination of ZIKV-induced vesicles into the ER lumen is mediated through the host reticulon 3.1A (RTN 3.1A) that facilitates ER membrane curvature ( Fig. 1, blue box) [38]. Silencing of RTN3.1A impairs NS4A protein stability and results in significant reduction of virusinduced vesicles and virus replication [38].…”