C-type lectin domain group 14 proteins in vascular biology, cancer and inflammation

Abstract

The C-type lectin domain (CTLD) group 14 family of transmembrane glycoproteins consist of thrombomodulin, CD93, CLEC14A and CD248 (endosialin or tumour endothelial marker-1). These cell surface proteins exhibit similar ectodomain architecture and yet mediate a diverse range of cellular functions, including but not restricted to angiogenesis, inflammation and cell adhesion. Thrombomodulin, CD93 and CLEC14A can be expressed by endothelial cells, whereas CD248 is expressed by vasculature associated pericytes, activated fibroblasts and tumour cells among other cell types. In this article, we review the current literature of these family members including their expression profiles, interacting partners, as well as established and speculated functions. We focus primarily on their roles in the vasculature and inflammation as well as their contributions to tumour immunology. The CTLD group 14 family shares several characteristic features including their ability to be proteolytically cleaved and engagement of some shared extracellular matrix ligands. Each family member has strong links to tumour development and in particular CD93, CLEC14A and CD248 have been proposed as attractive candidate targets for cancer therapy.

Keywords: C-type lectin; CD248; CD93; CLEC14A; cancer; extracellular matrix; group XIV; immuno-oncology; thrombomodulin; vascular targeting.

Figure 1

CTLD group 14 family proteins. Schematic diagrams of the CTLD group 14 family proteins. Each protein is drawn to relative scale based on primary amino acid sequence length. The CTLD is shown in red, the sushi in blue and the EGF repeats in green.

Figure 2

Alignments of CTLD group 14 family members based on amino acid sequence. Amino acid alignments of the whole primary sequence of each human family member using PRALINE 229. The following protein sequences were used thrombomodulin (http://www.uniprot.org/uniprot/P07204), CD93 (http://www.uniprot.org/uniprot/Q9NPY3), CLEC14A (http://www.uniprot.org/uniprot/Q86T13) and CD248 (http://www.uniprot.org/uniprot/Q9HCU0).

Figure 3

Schematic of thrombomodulin protein structure with ligand binding partners. Thrombomodulin CTLD has been shown to interact with fibronectin, HMGB1, Kringle 1–5, Lewis Y antigen and HSP70‐1. The CTLD may be proteolytically cleaved by an as yet unidentified MMP. Thrombin binds to the 5th and 6th EGF domains, this binding is in competition with Ang1 and/or Ang2. RHBDL2 can cleave the whole ECD of thrombomodulin as can neutrophil elastase, cathepsin G and proteinase 3. The cytoplasmic tail binds to ezrin which in turn links thrombomodulin to the actin cytoskeleton.

Figure 4

Schematic of CD248 structure with ligand binding partners. CD248 CTLD binds to fibronectin, Mac‐2 BP, Collagens I and IV and MMRN2. There are currently no known direct intracellular interaction partners for CD248.

Figure 5

Schematic of CD93 structure with ligand binding partners. CD93 CTLD binds to MMRN2. The whole ECD has been shown to be cleaved by an as yet unidentified metalloproteinase. The intracellular cytoplasmic domain binds to moesin which in turn links CD93 to the actin cytoskeleton. The cytoplasmic domain also binds to Cbl and GIPC1 and src.

Figure 6

Schematic of CLEC14A protein with ligand binding partners. CLEC14A CTLD binds to MMRN2 and to HSP70‐1A. The whole ECD can be cleaved by RHBDL2. There are currently no known direct intracellular partners for CLEC14A.

Figure 7

Expression of CTLD group 14 family members in mouse tissues. The Tabula Muris database was used to determine which mouse cell types expressed each CTLD group 14 family gene from data acquired through fluorescence activated cell sorting and single‐cell gene expression analysis. The t‐SNE plot at the top displays annotations of each cell type and shows a legend of colours corresponding to which organ or tissue type that cell was from. The lower t‐SNE plots display in which cell types each family member was expressed (purple), ln(1 + CPM) is the natural logarithm of counts per million + 1.

Figure 8

Endothelial expression of CTLD group 14 family members in mouse tissues. (A) The Tabula Muris database was used to create t‐SNE plots of all endothelial cells from different organs as well as brain pericytes. The t‐SNE plot at the top left displays a legend of colours corresponding to which organ or tissue type that cell was from. Expression of each CTLD group 14 family member within these cell types are displayed as t‐SNE plots. (B) Single‐cell sequencing data analysed as fragments per kilobase million was used to compare CD93 and CLEC14A expression in different endothelial cells from different organs. Wilcoxon statistical test was used to compare ****P ≤ 0.0001 Aorta ECs n = 262, Brain nonmyeloid EC n = 1250, Diaphragm EC n = 154, Fat EC n = 1180, Heart EC n = 2274, Heart endocardial cell n = 350, Kidney EC n = 80, Limb Muscle EC n = 258, Liver EC n = 392, Lung EC n = 1476, Mammary gland EC n = 98, Pancreas EC n = 98, Trachea EC n = 66. (C) t‐SNE plots of lung endothelium alone were created which revealed the presence of a cluster of cells expressing low levels of CD93 when compared with all other lung endothelial cells but similar levels of CLEC14A (grey ellipse).