Ly49B Is Expressed on Multiple Subpopulations of Myeloid Cells

Abstract

Using a novel mAb specific for mouse Ly49B, we report here that Ly49B, the last remaining member of the C57 Ly49 family to be characterized, is expressed at low levels on ∼1.5% of spleen cells, none which are NK cells or T cells but which instead belong to several distinct subpopulations of myeloid cells defined by expression of CD11b and different levels of Gr1. Much larger proportions of bone marrow and peritoneal cells expressed Ly49B, all being CD11b+ and comprising multiple subpopulations defined by light scatter, F4/80, and Gr1 expression. Costaining for Ly49Q, also expressed on myeloid cells, revealed that Ly49B and Ly49Q were most strongly expressed on nonoverlapping subpopulations, Ly49Qhigh cells being mostly B220+CD4+ and/or CD8+, Ly49B+ cells lacking these markers. Myeloid populations that developed from bone marrow progenitors in vitro frequently coexpressed both Ly49B and Ly49Q, and Ly49B expression could be up-regulated by LPS, α-IFN, and γ-IFN, often independently of Ly49Q. PCR analysis revealed that cultured NK cells and T cells contained Ly49B transcripts, and Ly49B expression could be detected on NK cells cultured in IL-12 plus IL-18, and on an immature NK cell line. Immunohistochemical studies showed that Ly49B expression in tissues overlapped with but was distinct from that of all other myeloid molecules examined, being particularly prominent in the lamina propria and dome of Peyer’s patches, implicating an important role of Ly49B in gut immunobiology. In transfected cells, Ly49B was found to associate with SHP-1, SHP-2, and SHIP in a manner strongly regulated by intracellular phosphorylation events.

SHIP Influences Signals from CD48 and MHC Class I Ligands That Regulate NK Cell Homeostasis, Effector Function, and Repertoire Formation

“…4A) or an H-2 d (B10.D2) background (Fig. 4B) in both the spleen and bone marrow (BM), whereas virtually no Ly49B expression was detected on WT NK cells from both haplotypes, consistent with the findings of Gays et al (21). We performed Ly49B blocking studies using hybridoma supernatants or purified Ab against Ly49B (21) Whether there are functional consequences for NK cells in vivo that arise owing to lineage-inappropriate expression of Ly49B will require the development of Ly49B 2/2 mice and thus remain to be determined.…”

“…These data suggest that 2B4 may be necessary to limit the expression of these activating receptors. In addition, we find that SHIP and 2B4 are required to prevent the lineage-inappropriate expression of Ly49B, the polyspecific MHC-I receptor that is normally restricted to myeloid cells (21). This finding establishes a previously unappreciated role for SHIP and 2B4 in maintaining lineage restricted expression of immune receptors.…”

“…SHIP acting in trans from 2B4 or in cis from the same MHC-I receptors could limit PI3K/Akt survival signals, and thereby prevent inappropriate expansion of such NK subsets. SHIP can also be recruited directly to MHC-I receptors; therefore, SHIP may also limit these subsets in cis (3,21,24). This cis activity may be important when MHC-I receptors are in a genetic background where high-affinity MHC-I ligands are also present, as suggested by the overexpression of Ly49A in SHIP 2/2 H-2 d mice.…”

“…Ly49B and Ly49Q are the only members of the Ly49 gene family not expressed by NK cells, but instead are restricted to myeloid lineage cells (21). Like the Ly49A and C receptors we previously found to be regulated by SHIP, Ly49B is also a promiscuous MHC-I receptor that does not exhibit precise specificity for a given MHC haplotype (24).…”

Optimized Tetramer Analysis Reveals Ly49 Promiscuity for MHC Ligands

“…Additionally, two Ly49, Ly49B and Ly49Q, are expressed on myeloid-derived cells, including macrophages, neutrophils, and DCs (6,7). Initial cDNA screening of inbred mouse strains revealed extreme genetic diversity and polymorphism in different mouse strains (8), which was later revealed to be a result of genomic plasticity within the Ly49 region resulting in cluster sizes ranging from 8 to 22 Ly49 genes (9,10).…”