Reticulon‐like properties of a plant virus‐encoded movement protein

Abstract

Plant viruses encode movement proteins (MPs) that ensure the transport of viral genomes through plasmodesmata (PD) and use cell endomembranes, mostly the endoplasmic reticulum (ER), for delivery of viral genomes to PD and formation of PD-anchored virus replication compartments. Here, we demonstrate that the Hibiscus green spot virus BMB2 MP, an integral ER protein, induces constrictions of ER tubules, decreases the mobility of ER luminal content, and exhibits an affinity to highly curved membranes. These properties are similar to those described for reticulons, cellular proteins that induce membrane curvature to shape the ER tubules. Similar to reticulons, BMB2 adopts a W-like topology within the ER membrane. BMB2 targets PD and increases their size exclusion limit, and these BMB2 activities correlate with the ability to induce constrictions of ER tubules. We propose that the induction of ER constrictions contributes to the BMB2-dependent increase in PD permeability and formation of the PD-associated replication compartments, therefore facilitating the virus intercellular spread. Furthermore, we show that the ER tubule constrictions also occur in cells expressing TGB2, one of the three MPs of Potato virus X (PVX), and in PVX-infected cells, suggesting that reticulon-like MPs are employed by diverse RNA viruses.

Constriction of endoplasmic reticulum tubules by the viral movement protein BMB2 is associated with local BMB2 anchorage at constriction sites

“…Recently, HGSV BMB2 has been shown to induce localized constrictions of ER tubules, to exhibit an affinity to highly curved membranes, and to adopt a W-like topology within the ER membrane, showing therefore properties similar to those known for reticulons, cellular proteins that induce membrane curvature to shape the ER tubules. 6 Despite these striking similarities to reticulons, a notable difference in the pattern of ER constrictions induced by BMB2 and reticulons has been observed. While mRFP-BMB2 was localized to discrete ER tubule sites lacking the ER-GFP 7 , a soluble marker for the ER lumen 8 , and thus representing the localized sites of ER tubule constrictions 6 , reticulons induce constrictions without being focally localized to discrete sites in the ER tubules.…”

“…6 Despite these striking similarities to reticulons, a notable difference in the pattern of ER constrictions induced by BMB2 and reticulons has been observed. While mRFP-BMB2 was localized to discrete ER tubule sites lacking the ER-GFP 7 , a soluble marker for the ER lumen 8 , and thus representing the localized sites of ER tubule constrictions 6 , reticulons induce constrictions without being focally localized to discrete sites in the ER tubules. 9,10 BMB2-mRFP, which carries mRFP at the C-terminus retains the ability for transport to peripheral membrane bodies typical for non-fused BMB2 and mRFP-BMB2.…”

“…5 Additionally, BMB2 co-localizes with PDassociated callose and increases the PD size exclusion limit. 6 Moreover, BMB2 is capable of directing BMB1, which is supposed to bind viral RNA, to BMB2-containing membrane bodies, PD, and neighboring cells. 5 Therefore, intra-and intercellular transport of BMB1 and, likely, viral RNA depends on BMB2 functions, in particular, the interactions with ER membranes.…”

“…However, unlike mRFP-BMB2 this protein is unable to induce tubule constrictions and to focally bind to discrete sites along the ER tubules. 6 Such a correlated loss of two protein functions in BMB2-mRFP allows two alternative explanations. First, the formation of BMB2-containing structures at discrete ER sites may be required for BMB2-induced constrictions of ER tubules at these sites.…”

“…In a control, BMB2-mRFP expressed in ER-GFP-transgenic Nicotiana benthamiana plants was observed in ER-derived cell wall-associated bodies (Figure 1 (a-c)), as described previously. 6 When MyoXI2F was coexpressed with BMB2-mRFP in ER-GFP-transgenic plants, only a portion of BMB2-mRFP was localized to cell wall-associated bodies, whereas the rest of the fusion protein was found either in thread-like ER-associated structures as visible in some cells (Figure 1 (d-f)), or distinct small aggregates associated with the cortical ER network as found in other cells (Figure 1 (g-i)). The latter structures resembled those found for mRFP-BMB2 (Figure 1 (j-l)) at constriction sites 6 ; however, unlike for mRFP-BMB2, the normal appearance of cortical ER tubules remained unaffected upon BMB2-mRFP expression in these cells, and no visible constrictions were found.…”