Identification and Characterization of a Novel Human Myeloid Inhibitory C-type Lectin-like Receptor (MICL) That Is Predominantly Expressed on Granulocytes and Monocytes

Abstract

Inhibitory and activatory C-type lectin-like receptors play an important role in immunity through the regulation of leukocytes. Here, we report the identification and characterization of a novel myeloid inhibitory Ctype lectin-like receptor (MICL) whose expression is primarily restricted to granulocytes and monocytes. This receptor, which contains a single C-type lectin-like domain and a cytoplasmic immunoreceptor tyrosine-based inhibitory motif, is related to LOX-1 (lectin-like receptor for oxidized low density lipoprotein-1) and the ␤-glucan receptor (Dectin-1) and is variably spliced and highly N-glycosylated. We demonstrate that it preferentially associates with the signaling phosphatases SHP-1 and SHP-2, but not with SHIP. Novel chimeric analyses with a construct combining MICL and the ␤-glucan receptor show that MICL can inhibit cellular activation through its cytoplasmic immunoreceptor tyrosine-based inhibitory motif. These data suggest that MICL is a negative regulator of granulocyte and monocyte function.The functional balance of the immune system is regulated, in part, by inhibitory and activatory receptors found on all leukocytes as well as on many non-immune cells (1). Within these activatory and inhibitory receptor families, cytoplasmic consensus motifs have been identified and shown to associate with particular signaling mechanisms. These motifs include the immunoreceptor tyrosine-based activation motifs (ITAMs), 1 which become phosphorylated upon stimulation, sending their activatory signals through a variety of intracellular enzyme cascades and resulting in a wide range of cellular outcomes, and the immunoreceptor tyrosine-based inhibitory motifs (ITIMs), which suppress such activatory signals by recruiting phosphatase enzymes that dephosphorylate ITAMs as well as other components of the activatory pathways.The array of inhibitory and activatory receptors expressed by natural killer cells has recently been the focus of much investigation (2). In humans, these receptors can be divided into two families: the Ig superfamily, including the killer cell Ig-like receptors (3) that bind classical major histocompatibility complex class I or HLA G (non-classical), and the C-type lectin-like family (4), including the NKG2 receptors, which bind the nonclassical HLA E or other distantly related molecules that are up-regulated on stressed, transformed, or infected cells (5, 6).The C-type lectin and lectin-like proteins are classified into 14 groups based on the arrangement of their C-type lectin-like domains (CTLDs).2 In particular, group V receptors include the lectin-like NKG2 family mentioned above, but also a distinct subgroup of these receptors that are expressed predominantly on myeloid and endothelial cells (7). The members of this subgroup appear to bind diverse ligands, as exemplified by LOX-1 (lectin-like receptor for oxidized low density lipoprotein-1) (8) and the ␤-glucan receptor (BGR), which is an activatory phagocytic receptor for ␤-glucans (9).3 Although they share structuralhomologywithclassic...

Human MICL (CLEC12A) is differentially glycosylated and is down‐regulated following cellular activation

“…MICL contains an ITIM in its cytoplasmic tail and our initial studies using chimeras suggested that this molecule could function as an inhibitory receptor. To gain more insight into the role of this receptor, we generated a novel mAb and have used it here to examine MICL on primary cells.Our analysis of MICL, in both transduced cell lines [14] and primary cells, suggests that this molecule is highly N-glycosylated compared to its most closely MICL expression was predominantly restricted to myeloid cells in both peripheral blood and in tissues, and although low levels were observed on CD4 + T cells, this receptor was not detected on any other lymphocyte population. This expression profile of MICL is similar to that of the related receptor, beta-glucan receptor/ Dectin-1, found in the same gene cluster [21], and is largely consistent with previous transcript analyses [14,16,17], although some differences were noted, as discussed above.…”

“…MICL can be alternatively spliced into at least three isoforms and is highly N-glycosylated when expressed in heterologous cell lines. Others and we have shown that MICL can associate with the inhibitory signalling phosphatases, SHP-1 and SHP-2, and that MICL can function as an inhibitory receptor [14][15][16]. The ligand of this receptor is unknown.…”

“…7). To determine whether these discrepancies in MICL molecular mass were due to the expression of different splice variants [14] or to variable glycosylation, cell lysates were de-glycosylated and examined by Western blotting (Fig. 7).…”

“…The genes of most C-type lectin-like molecules are located within the 'natural killer complex' (NKC), but many of those expressed by myeloid and other cells are found in a distinct cluster of genes within the NKC [10]. This cluster includes Dectin-1/ betaglucan receptor [11], the lectin-like receptor for oxidized LDL (LOX-1) [12], CLEC-1 and CLEC-2 [13], two unstudied transcripts [14], and myeloid inhibitory C-type lectin-like receptor (MICL) [14].…”

“…The genes of most C-type lectin-like molecules are located within the 'natural killer complex' (NKC), but many of those expressed by myeloid and other cells are found in a distinct cluster of genes within the NKC [10]. This cluster includes Dectin-1/ betaglucan receptor [11], the lectin-like receptor for oxidized LDL (LOX-1) [12], CLEC-1 and CLEC-2 [13], two unstudied transcripts [14], and myeloid inhibitory C-type lectin-like receptor (MICL) [14].MICL (also called CLL-1, KLRL1 and DCAL-2) has been independently identified by several groups and shown to be a type II transmembrane receptor comprising a single C-type lectin-like domain (which is not predicted to bind either calcium or sugar), a stalk region, a transmembrane domain and a short cytoplasmic tail containing an ITIM motif [14][15][16][17]. MICL orthologs have been identified in the mouse, rat, and dog, indicating that the receptor is conserved across species.…”