Different isoforms of the neuronal guidance molecule Slit2 directly cause chemoattraction or chemorepulsion of human neutrophils 1

The movement of neutrophils between blood and tissues appears to be regulated by chemoattractants and chemorepellents. Compared to neutrophil chemoattractants, relatively little is known about neutrophil chemorepellents. Slit proteins are endogenously cleaved into a variety of N and C terminal fragments, and these fragments are neuronal chemorepellents and inhibit chemoattraction of many cell types, including neutrophils. In this report, we show that the 140 kDa N-terminal Slit2 fragment (Slit2-N) is a chemoattractant and the 110 kDa N-terminal Slit2 fragment (Slit2-S) is a chemorepellent for human neutrophils. The effects of both Slit2 fragments were blocked by antibodies to the Slit2 receptor Robo1 or the Slit2 co-receptor syndecan-4. Slit2-N did not appear to activate Ras, but increased PIP3 levels. Slit2-N induced chemoattraction was unaffected by Ras inhibitors, reversed by PI3 kinase inhibitors, and blocked by cdc42 and Rac inhibitors. In contrast, Slit2-S activated Ras but did not increase PIP3 levels. Slit2-S induced chemorepulsion was blocked by Ras and Rac inhibitors, not affected by PI3 kinase inhibitors, and was reversed by cdc42 inhibitors. Slit2-N but not Slit2-S increased neutrophil adhesion, myosin II light chain phosphorylation, and polarized actin formation and single pseudopods at the leading edge of cells. Slit2-S induced multiple pseudopods. These data suggest that Slit-2 isoforms use similar receptors but different intracellular signaling pathways, and have different effects on the cytoskeleton and pseudopods, to induce neutrophil chemoattraction or chemorepulsion.