Class II-mediated signals play potential roles in B cell activation and antigen presentation. The regions of the class II molecule participating in B cell signaling are incompletely defined. Our prior analysis of structural requirements of the cytoplasmic domain of A beta revealed that only the eight membrane-proximal residues are required for signaling. Here, we report that the sequence and position of two of these are critical, and present direct evidence that the A beta transmembrane domain is also involved in signaling, via a pathway distinct from the cytoplasmic domain. These results demonstrate that specific regions in both the cytoplasmic and transmembrane domains of the class II molecule have distinct signaling functions.