Zeile W L, Condit R C, Lewis J I, Purich D L, Southwick F S
Department of Medicine, Division of Infectious Diseases, University of Florida College of Medicine, Gainesville, FL 32610, USA.
Proc Natl Acad Sci U S A. 1998 Nov 10;95(23):13917-22. doi: 10.1073/pnas.95.23.13917.
Vaccinia uses actin-based motility for virion movement in host cells, but the specific protein components have yet to be defined. A cardinal feature of Listeria and Shigella actin-based motility is the involvement of vasodilator-stimulated phosphoprotein (VASP). This essential adapter recognizes and binds to actin-based motility 1 (ABM-1) consensus sequences [(D/E)FPPPPX(D/E), X = P or T] contained in Listeria ActA and in the p90 host-cell vinculin fragment generated by Shigella infection. VASP, in turn, provides the ABM-2 sequences [XPPPPP, X = G, P, L, S, A] for binding profilin, an actin-regulatory protein that stimulates actin filament assembly. Immunolocalization using rabbit anti-VASP antibody revealed that VASP concentrates behind motile virions in HeLa cells. Profilin was also present in these actin-rich rocket tails, and microinjection of 10 microM (intracellular) ABM-2 peptide (GPPPPP)3 blocked vaccinia actin-based motility. Vinculin did not colocalize with VASP on motile virions and remained in focal adhesion contacts; however, another ABM-1-containing host protein, zyxin, was concentrated at the rear of motile virions. We also examined time-dependent changes in the location of these cytoskeletal proteins during vaccinia infection. VASP and zyxin were redistributed dramatically several hours before the formation of actin rocket tails, concentrating in the viral factories of the perinuclear cytoplasm. Our findings underscore the universal involvement of ABM-1 and ABM-2 docking sites in actin-based motility of Listeria, Shigella, and now vaccinia.
痘苗病毒利用基于肌动蛋白的运动方式在宿主细胞内移动病毒粒子，但其具体的蛋白质成分尚未明确。单核细胞增生李斯特菌和志贺氏菌基于肌动蛋白的运动方式的一个主要特征是血管舒张刺激磷蛋白（VASP）的参与。这种重要的衔接蛋白可识别并结合李斯特菌ActA以及志贺氏菌感染产生的p90宿主细胞纽蛋白片段中所含的基于肌动蛋白的运动1（ABM-1）共有序列[（D/E）FPPPPX（D/E），X = P或T]。反过来，VASP提供ABM-2序列[XPPPPP，X = G、P、L、S、A]用于结合肌动蛋白调节蛋白丝切蛋白，该蛋白可刺激肌动蛋白丝组装。使用兔抗VASP抗体进行免疫定位显示，VASP在HeLa细胞中运动的病毒粒子后方聚集。丝切蛋白也存在于这些富含肌动蛋白的“火箭尾”中，显微注射10微摩尔（细胞内）的ABM-2肽（GPPPPP）3可阻断痘苗病毒基于肌动蛋白的运动。纽蛋白在运动的病毒粒子上不与VASP共定位，而是保留在粘着斑中；然而，另一种含有ABM-1的宿主蛋白斑联蛋白则聚集在运动病毒粒子的尾部。我们还研究了痘苗病毒感染过程中这些细胞骨架蛋白位置随时间的变化。在肌动蛋白“火箭尾”形成前数小时，VASP和斑联蛋白就发生了显著的重新分布，集中在核周细胞质的病毒工厂中。我们的研究结果强调了ABM-1和ABM-2对接位点在李斯特菌、志贺氏菌以及现在的痘苗病毒基于肌动蛋白的运动中的普遍参与。
