Rac2 regulates neutrophil chemotaxis, superoxide production, and myeloid colony formation through multiple distinct effector pathways.
Polymorphonuclear neutrophils (PMN) are an important component of the innate immune system. We have shown previously that migration and superoxide (O2*-) production, as well as some kinase signaling pathways are compromised in mice deficient in the Ras-related Rho GTPase Rac2. In this study, we demonstrate that Rac2 controls chemotaxis and superoxide production via distinct pathways and is critical for development of myeloid colonies in vitro. The Rac2 mutants V36A, F37A, and N39A all bind to both Pak1 and p67(phox), yet are unable to rescue superoxide production and chemotaxis when expressed in Rac2-/- PMN. In contrast, the N43A mutant, which binds to Por1 (Arfaptin 2), p67phox, and Pak1, is able to rescue superoxide production but not chemotaxis. The F37A mutant, demonstrated to have reduced binding to Por1, shows reduced rescue of fMLP-induced chemotaxis. Finally, the Rac2Y40C mutant that is defective in binding to all three potential downstream effectors (Pak1, p67phox, and Por1) is unable to rescue chemotaxis, motility, or superoxide production, but is able to rescue defective growth of myeloid colonies in vitro. These findings suggest that binding to any single effector is not sufficient to rescue the distinct cellular phenotypes of Rac2-/- PMN, implicating multiple, distinct, and potentially parallel effector pathways.
Carstanjen
Dirk
D
Division of Experimental Hematology, Children's Hospital Research Foundation, Cincinnati, OH 45229, USA.
Yamauchi
Akira
A
Koornneef
Annemart
A
Zang
Heesuk
H
Filippi
Marie-Dominique
MD
Harris
Chad
C
Towe
Jason
J
Atkinson
Simon
S
Zheng
Yi
Y
Dinauer
Mary C
MC
Williams
David A
DA
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000818:Animals; D002634:Chemotaxis, Leukocyte / Q000502:physiology*; D003114:Colony-Forming Units Assay; D007113:Immunity, Innate; D066298:In Vitro Techniques; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009154:Mutation; D009504:Neutrophils / Q000166:cytology* / Q000276:immunology / Q000502:physiology*; D010750:Phosphoproteins / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D015398:Signal Transduction; D013481:Superoxides / Q000378:metabolism*; D014161:Transduction, Genetic; D054462:p21-Activated Kinases; D020744:rac GTP-Binding Proteins / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D054460:rho-Associated Kinases; D000097766:RAC2 GTP-Binding Protein
D048868:Adaptor Proteins, Signal Transducing; D047908:Intracellular Signaling Peptides and Proteins; D010750:Phosphoproteins; D017346:Protein Serine-Threonine Kinases; D013481:Superoxides; D054462:p21-Activated Kinases; D020744:rac GTP-Binding Proteins; D054460:rho-Associated Kinases; D000097766:RAC2 GTP-Binding Protein; C079789:neutrophil cytosol factor 67K; C515905:Pak1 protein, mouse