Fig 1: Cyclin D3 associates with SARS-CoV-2 proteins E and M A293T cells were contransfected with HA-cyclin D3 and Strep-tag-SARS-CoV-2 E or nsp9, and control plasmid (EV). Immunoprecipitation was performed using anti-HA antibody. The immunoprecipitates were blotted with anti-Strep, anti-HA and cyclin D3 antibodies.B293T cells were contransfected with HA-cyclin D3 and Strep-tag-SARS-CoV-2 E, M, both E and M or nsp9. Immunoprecipitation was performed using anti-HA antibody. The immunoprecipitates were blotted with anti-Strep, anti-HA antibodies. WCL, whole-cell lysate.C–G293T GFP11 cells were transfected with spike, and/or with envelope, membrane and cyclin D3. 24 h post-transfection, cells were seeded at a 1:1 ratio with Vero-GFP10 cells, and the percentage of GFP + ve area (syncytia) was determined 18 h later. S, spike; E, envelope; M, membrane; D3, cyclin D3. (C, D) Representative images of GFP+ syncytia. Scale bars: 40 µm. (E–G) Quantification of cell-to-cell fusion showing the percentage of the GFP + ve area to the acquired total cell area. (E) n = 4 biological replicates. (F, G) n = 4 biological replicates; technical triplicates shown; one-way ANOVA with Dunnett's multiple comparisons test: ns, non-significant; ****P < 0.0001; ***P < 0.001; **P < 0.01. Bars indicate mean with SD.HDiagram of SARS-CoV-2 VLPs construction in the presence or absence of cyclin D3 in 293T cells.IRepresentative example of western blot from purified SARS-CoV-2 virus-like particles (VLP) and whole-cell lysates (WCL). MCM2 was used as a loading control.JQuantification of protein incorporation (into VLPs) or expression (in WCL). n = 4 biological replicates; one-way ANOVA with Dunnett's multiple comparisons test: ns, non-significant; ***P < 0.001. Bars indicate the mean with SD. N, nucleocapsid; M, membrane; E, envelope; S, spike. Source data are available online for this figure.