Grb2 and Nck act cooperatively to promote actin-based motility of vaccinia virus.
The Wiskott-Aldrich syndrome protein family member N-WASP is a key integrator of the multiple signalling pathways that regulate actin polymerization via the Arp2/3 complex. Our previous studies have shown that N-WASP is required for the actin-based motility of vaccinia virus and is recruited via Nck and WIP. We now show that Grb2 is an additional component of the vaccinia actin tail-forming complex. Recruitment of Nck and Grb2 to viral particles requires phosphorylation of tyrosine residues 112 and 132 of A36R, the vaccinia actin tail nucleator, respectively. The presence of Grb2 on the virus is also dependent on the polyproline-rich region of N-WASP. The Grb2 pathway alone is therefore unable to nucleate actin tails, as its recruitment requires the prior recruitment of N-WASP by Nck. However, Grb2 does play an important role in actin-based motility of vaccinia, as in its absence, the mean number of actin tails per cell is reduced 2.6-fold. Thus, both Nck and Grb2 act in a cooperative manner to stabilize and/or activate the vaccinia actin-nucleating complex. We suggest that such cooperativity between "primary" and "secondary" adaptor proteins is likely to be a general feature of receptor-mediated signalling.
Scaplehorn
Niki
N
European Molecular Biology Laboratory, Meyerhofstrasse 1, D-69117 Heidelberg, Germany.
Holmström
Anna
A
Moreau
Violaine
V
Frischknecht
Freddy
F
Reckmann
Inge
I
Way
Michael
M
D000199:Actins / Q000378:metabolism*; D048868:Adaptor Proteins, Signal Transducing; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D051380:GRB2 Adaptor Protein; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D015513:Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D014616:Vaccinia virus / Q000235:genetics / Q000502:physiology*; D051316:Wiskott-Aldrich Syndrome Protein
D000199:Actins; D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D008164:Luminescent Proteins; C078332:Nck protein; D015513:Oncogene Proteins; D011506:Proteins; D011993:Recombinant Fusion Proteins; C495767:WAS protein, human; D051316:Wiskott-Aldrich Syndrome Protein; D049452:Green Fluorescent Proteins