Loss of CCR2 leads to enhanced early event of BCR activation
BCR clustering and spread are essential for the initiation of the BCR signal. Based on the boosted F-actin accumulation in CCR2 KO B cells, we investigated the influence CCR2 depletion has on the early activation of B cells. Upon stimulation with mAg for 3, 5, and 7 min, the distribution pattern of F-actin in CCR2 KO B cells appeared at the outer edge of the central BCR cluster, which was similar to the pattern observed in WT B cells (Fig. 5A ). However, the MFI of F-actin in the contact zone of KO B cells increased over time from 3 to 5 min post-stimulation, which was significantly higher than that in WT B cells (Fig. 5B-C ). Consistent with the augmented F-actin accumulation, the recruitment of pWASP in KO B cells was also increased at 3 min after stimulation (Fig. 5A, D ). Moreover, these changes were accompanied by the increased accumulation of pCD19 in KO B cells at 3 and 5 min post-stimulation, compared to that of WT B cells (Fig. 5E-F ). As was the case for pCD19, the recruitment of pY, pBtk, and pSHIP in KO B cells remained higher than that of WT B cells (Fig. 5G-K ). Convincingly, these findings suggest that CCR2 KO promotes the aggregation of BCR microclusters and accumulation of signalosome at an early phase of B cell activation, which subsequently influences the formation of the central clusters at the terminal phase; thus, positively regulating B cell signaling.