Rac GTPases are activated by extracellular stimuli and contribute to cellular responses including cytoskeletal changes and cell migration. Dominant-negative Rac1 has been used to implicate Rac GTPases in these responses, but which of the three mammalian Rac isoforms it inhibits is not known. We show that mouse bone marrow-derived macrophages express Rac1, low levels of Rac2 but not Rac3. As Rac1-null mice die early in development, we have used mice with a loxP-flanked allele of Rac1 and the type I interferon-inducible Mx1-Cre transgene to address for the first time the specific role of Rac1 in cell motility. Bone marrow-derived macrophages isolated from mice treated with polyIC to induce interferon lack detectable Rac1, and there is no compensatory increase in Rac2 or Cdc42 expression. Rac1-deficient macrophages have an altered morphology: they are significantly more elongated than control cells and have a reduced adhesive area. Re-expression of Rac1 reverts the morphology to that of control cells. Loss of Rac1 reduces but does not completely prevent membrane ruffling in response to CSF-1. However, Rac1-deficient macrophages show normal migration and chemotaxis. Thus in macrophages Rac1 is primarily responsible for regulating cell morphology, contributes to membrane ruffling, but is not required for migration.

Rac GTPases are believed to contribute to migration in leukocytes by transducing signals from cell surface receptors to the actin and microtubule cytoskeletons. Mammals have three closely related Rac isoforms, Rac1, Rac2 and Rac3, and it is widely assumed that cell migration requires the activity of these Rac GTPases. We have previously shown that Rac1-null mouse macrophages have altered cell shape and reduced membrane ruffling but normal migration speed. Here we investigate the behaviour of macrophages lacking Rac2 (Rac2–/–) or Rac1 and Rac2 (Rac1/2–/–). Rac2–/– macrophages have reduced F-actin levels and lack podosomes, which are integrin-based adhesion sites, and their migration speed is similar to or slightly slower than wild-type macrophages, depending on the substrate. Unexpectedly, Rac1/2–/– macrophages, which do not express Rac1, Rac2 or Rac3, migrate at a similar speed to wild-type macrophages on a variety of substrates and perform chemotaxis normally, although their morphology and mode of migration is altered. However, Rac1–/– and Rac1/2–/– but not Rac2–/– macrophages are impaired in their ability to invade through Matrigel. Together, these data show that Rac1 and Rac2 have distinct roles in regulating cell morphology, migration and invasion, but are not essential for macrophage migration or chemotaxis.

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