Abstract Non-classical major histocompatibility complex class-II (MHCII) protein H2-M edits peptides bound to conventional MHCII (pMHCII) in favor of stable pMHCII complexes. Although the impact of H2-M deficiency on pMHCII expression and T cell activation is well-studied, information on how this protein affects the development and phenotypic profile of APCs is lacking. We found that the absence of H2-M in C57BL/6 (B6) mice, resulted in a loss of surface expression and peptide-cargo of MHCII molecules in B cells across lymphoid organs. Lower pMHCII expression reduced MHCII association with BCR, affecting the integrated BCR signaling. Importantly, in H2-M deficient B6 mice, compared to the wildtype mice, frequency and abundance of B-1 cells, but not conventional B-2 cells, was reduced in the spleen and peritoneum. This H2-M mediated effect on the B-1 cell population was only evident in the B6 background (I-Ab), but not in BALB/c (I-Ad/I-Ed), indicating an MHCII haplotype-dependent phenomenon. A decrease in B-1 cell number also was evident in immature B-1 cells, emphasizing that H2-M deficiency affects B-1 cell development. In H2-M KO mice compared to WT B6, B-1 cells display a significantly lower self-renewal capacity and a higher rate of apoptosis. Furthermore, H2-M deficiency alters the B-1 BCR repertoire, selecting for B-1 cells specific for highly abundant epitopes, but not for low-frequency epitopes. Collectively, these data identify the impact of H2-M/MHCII interaction as a regulator of BCR signaling that influences the selection, maturation, and maintenance of mature B-1 cell clones in the periphery.