CD22 is a negative regulator of B cell signaling, an activity modulated by its interaction with glycan ligands containing α2-6-linked sialic acids. B cells deficient in the enzyme (ST6Gal I) that forms the CD22 ligand show suppressed BCR signaling. Here we report that mice deficient in both CD22 and its ligand (Cd22^−/−St6gal1^−/− mice) showed restored B cell receptor (BCR) signaling, suggesting that the suppressed signaling of St6gal1^−/− cells is mediated through CD22. Coincident with suppressed BCR signaling, B cells lacking ST6Gal I showed a net redistribution of the BCR to clathrin-rich microdomains containing most of the CD22, resulting in a twofold increase in the localization of CD22 together with the BCR. These studies suggest an important function for the CD22-ligand interaction in regulating BCR signaling and microdomain localization.