Similar enhancement was also observed in Ag-activated NTAL KOs. These data support the hypothesis that competition between NTAL and LAT as kinase substrates could attenuate the response in WT cells through decreased tyrosine phosphorylation of LAT, followed by decreased binding and activation of phospholipase Cc1 and subsequent events. Fourth, BMMCs with both NTAL KD and NTAL KO exhibited enhanced F-actin depolymerization after stimulation with Ag alone and even more after simultaneous triggering with both Ag + SCF. F-actin depolymerization precedes degranulation and the observed decrease in amount of F-actin could account for the observed higher degranulation in NTAL-deficient cells than in WT cells after simultaneous activation with Ag + SCF. Fifth, cells activated through FceRI or KIT exhibited enhanced spreading on fibronectin. In cells with NTAL KD spreading was significantly decreased after activation with antigen, but was unaffected after SCF triggering. The observed data suggest that positive regulatory role of NTAL on Ag-mediated spreading is not the result of developmental compensatory events. Rather, spreading could be related to transient actin depolymerization which was observed in Ag-activated WT cells and even more in NTAL-deficient cells, but not in SCF-activated cells, WT or NTAL-deficient. Sixth, BMMCs with NTAL KD exhibited migration towards Ag comparable with that seen in NTAL KO cells, and significantly higher than in WT cells. We recently showed that the level of Acetylcorynoline active RhoA in resting NTAL KO BMMCs is at least twice as high as in WT cells. Although active RhoA transiently decreased after FceRI triggering, more in NTAL KO cells than in WT cells, it is likely that differences in regulation of RhoA Moexipril HCl activity in NTALdeficient cells and WT cells are responsible for the enhanced NTAL-regulated chemotaxis. It should be stressed that previous reports have shown that RhoA regulates chemotaxis in other cell types, such as neutrophils, macrophages, dendritic cells and lymphocytes.The data presented in this study, together with those obtained in mice experiencing systemic anaphylaxis indicate that in mouse mast cells NTAL is a negative regulator of FceRI signaling. In contrast to mouse cells, NTAL in human mast cells and rat basophilic leukemia -2H3 cells was described as a positive regulator of mast cells signaling.