Vaccinia virus proteins A36 and F12/E2 show strong preferences for different kinesin light chain isoforms

Abstract

Vaccinia virus (VACV) utilizes microtubule‐mediated trafficking at several stages of its life cycle, of which virus egress is the most intensely studied. During egress VACV proteins A36, F12 and E2 are involved in kinesin‐1 interactions; however, the roles of these proteins remain poorly understood. A36 forms a direct link between virions and kinesin‐1, yet in its absence VACV egress still occurs on microtubules. During a co‐immunoprecipitation screen to seek an alternative link between virions and kinesin, A36 was found to bind isoform KLC1 rather than KLC2. The F12/E2 complex associates preferentially with the C‐terminal tail of KLC2, to a region that overlaps the binding site of cellular 14‐3‐3 proteins. F12/E2 displaces 14‐3‐3 from KLC and, unlike 14‐3‐3, does not require phosphorylation of KLC for its binding. The region determining the KLC1 specificity of A36 was mapped to the KLC N‐terminal heptad repeat region that is responsible for its association with kinesin heavy chain. Despite these differing binding properties F12/E2 can co‐operatively enhance A36 association with KLC, particularly when using a KLC1‐KLC2 chimaera that resembles several KLC1 spliceforms and can bind A36 and F12/E2 efficiently. This is the first example of a pathogen encoding multiple proteins that co‐operatively associate with kinesin‐1.

Tagging of the vaccinia virus protein F13 with mCherry causes aberrant virion morphogenesis

“…Alternatively, one of the other VACV IEV membrane-associated viral proteins, A33, A34, B5, A56 or F13, might also do so. Most of these only have a small cytoplasmic domain, with the majority of the polypeptide being oriented into the membrane lumen (for a review see [ 47 ]), and a yeast-2-hybrid screen [ 21 ] and co-immunoprecipitation screen [ 22 ] failed to identify a direct interaction between any of these proteins and KLCs.…”

“…A36 is an integral membrane protein that is enriched in the outer envelope of IEVs [ 19 ] and associates with the kinesin light chain (KLC) component of kinesin-1 [ 20, 21 ]. Recently, A36 was shown to bind specifically to KLC isoform 1 and not to KLC2 [ 22 ]. Intracellular movement of IEVs at speeds consistent with MT-mediated trafficking occurs in the absence of A36, albeit at reduced efficiency and with reduced run lengths, and surface virions are present at 50 % of wild-type (WT) levels late during infection [ 23 ].…”

The Effects of Combined Exercise Training (Resistance-Aerobic) on Serum Kinesin and Physical Function in Type 2 Diabetes Patients with Diabetic Peripheral Neuropathy (Randomized Controlled Trials)

“…The connection of cell shipments to kinesin-1 is established through two kinesin light chains (KLC) [7]. KLC1 is responsible for connecting intercellular shipments such as cellular mitochondria and proteins [8] and plays a major role in axonal (nerve) transport [9].…”

Loss of Actin-Based Motility Impairs Ectromelia Virus Release In Vitro but Is Not Critical to Spread In Vivo

“…Once WV are formed at a perinuclear site, they are translocated to the cell periphery by the action of A36, F12, and E2, which collectively stabilize the microtubule motor complex, kinesin-1, at the viral surface [ 11 , 12 ]. The cytoplasmic domain of A36 contains two Trp-Asp/Trp-Glu (WD/WE) motifs that associate with kinesin-1 via tetratricopeptide repeats on kinesin light chain (KLC) isoform 1 [ 13 , 14 ]. F12 and E2 associate with KLC isoform 2, stabilizing the virus–kinesin-1 association [ 14 ].…”

“…The cytoplasmic domain of A36 contains two Trp-Asp/Trp-Glu (WD/WE) motifs that associate with kinesin-1 via tetratricopeptide repeats on kinesin light chain (KLC) isoform 1 [ 13 , 14 ]. F12 and E2 associate with KLC isoform 2, stabilizing the virus–kinesin-1 association [ 14 ]. At the cell membrane, the outer WV membrane fuses with the plasma membrane resulting in cell-associated extracellular virus particles that occupy membrane pits at the cell surface [ 8 ].…”