Abstract 1040 Gut dysbiosis: ecological causes and causative effects on human disease

Distinct signaling cues on host cell cargo proteins determine their intracellular trafficking routes.One such signaling cue is the dibasic motif (consensus KxKxx) that binds host coatomer (COPI) complexes.COPI complexes facilitate retrograde cargo trafficking from Golgi to ER. Variants of this consensus KxKxx Coatomer-Binding Motif (CBM) are present on the cytoplasmic tails of all coronavirus (CoV) spike glycoproteins.CoVs assemble at the ER-Golgi intermediate compartments (ERGIC), following which the virions bud into compartment lumen, with the virion-containing vesicles then egressing to plasma membranes.It is known that COPI complexes retain spike proteins internally , however it is unclear whether spike CBM variations alter CoV assembly, virus particle structure, and infectivity.To address the relevance of CBM variation, we compared infection parameters amongst several recombinant Mouse Hepatitis Viruses (rMHVs) with varying COPI binding affinities.Viruses with high COPI affinities were severely debilitated, low-yielding and with small plaque morphology.These strong COPI binder rMHVs were also unstable and rapidly acquired suppressor mutations that completely deleted the COPI-binding motif (CBM).By contrast, infection with rMHVs lacking COPI binding secreted abundant virion particles that were 10-fold less infectious.To further investigate differences in specific infectivities, we compared virion protein composition across WT and CBM mutant viruses.When compared with WT rMHVs, western blot analysis revealed less S incorporation into rMHVs lacking COPI binding.Furthermore, vast quantities of M protein secreted from CBM mutant rMHV infected cells.These data support the hypothesis that altered CBM-COPI interactions disrupt efficient spike incorporation into progeny virions.Our data also suggests that CBM-COPI interaction retains virions intracellularly, allowing for effective congregation of virion transmembrane proteins and assembly-budding at ERGIC organelles.