Efficient activation of the lymphangiogenic growth factor VEGF-C requires the C-terminal domain of VEGF-C and the N-terminal domain of CCBE1

Abstract

The collagen- and calcium-binding EGF domains 1 (CCBE1) protein is necessary for lymphangiogenesis. Its C-terminal collagen-like domain was shown to be required for the activation of the major lymphangiogenic growth factor VEGF-C (Vascular Endothelial Growth Factor-C) along with the ADAMTS3 (A Disintegrin And Metalloproteinase with Thrombospondin Motifs-3) protease. However, it remained unclear how the N-terminal domain of CCBE1 contributed to lymphangiogenic signaling. Here, we show that efficient activation of VEGF-C requires its C-terminal domain both in vitro and in a transgenic mouse model. The N-terminal EGF-like domain of CCBE1 increased VEGFR-3 signaling by colocalizing pro-VEGF-C with its activating protease to the lymphatic endothelial cell surface. When the ADAMTS3 amounts were limited, proteolytic activation of pro-VEGF-C was supported by the N-terminal domain of CCBE1, but not by its C-terminal domain. A single amino acid substitution in ADAMTS3, identified from a lymphedema patient, was associated with abnormal CCBE1 localization. These results show that CCBE1 promotes VEGFR-3 signaling and lymphangiogenesis by different mechanisms, which are mediated independently by the two domains of CCBE1: by enhancing the cleavage activity of ADAMTS3 and by facilitating the colocalization of VEGF-C and ADAMTS3. These new insights should be valuable in developing new strategies to therapeutically target VEGF-C/VEGFR-3-induced lymphangiogenesis.

CCBE1 is required for coronary vessel development and proper coronary artery stem formation in the mouse heart

“…Although our expression analysis does not show Ccbe1 expression in the vicinity of the endocardium, we think that epicardial-derived cells (EPDC), which migrate into the myocardium, may be an additional source of CCBE1. In agreement with this, there is evidence that Ccbe1 is expressed in the wall of blood vessels, fibroblasts and ECs (Tian et al, 2016;Jha et al, 2017) and promotes tube formation in vitro (Tian et al, 2016). We think the methods used in this study are not sensitive enough to detect subtle changes in Ccbe1 expression levels.…”

“…In agreement with its expression in the epicardium, VEGF‐C has been recently shown to play an important role during coronary vasculature development (Chen et al, ; Chen et al, ). Despite the requirement of CCBE1 for the maturation of VEGF‐C during lymphangiogenesis (Jeltsch et al, ; Le Guen et al, ; Jha et al, ), whether Ccbe1 is expressed in the vicinity of the developing coronary vessels and its requirement for VEGF‐C signaling during coronary vasculature development are still unknown. We first sought to know if Ccbe1 is expressed in the heart at the stages coronary vessel formation.…”

Loss of ADAMTS3 activity causes Hennekam lymphangiectasia–lymphedema syndrome 3

“…ADAMTS3 seems a requisite for this. Indeed, we reported elsewhere that an ADAMTS3 change (c.1694 G > A; p.Arg595Gln), which does not change VEGFC processing but reduces ADAMTS3 interaction with CCBE1, renders ADAMTS3 unable to retain CCBE1 at the cell surface, resulting in disruption of VEGFR3 signaling (24).…”

CHAPLE syndrome uncovers the primary role of complement in a familial form of Waldmann's disease

“…The mechanism how FAT4 mutations lead to lymphatic dysplasia remains to be identified. Recently, a third genetic cause of this syndrome has been identified (termed Hennekam lymphangiectasia‐lymphedema syndrome 3); A disintegrin and metallopeptidase with thrombospondin type 1 motif 3 (ADAMTS3) . The symptoms of the patients were reminiscent of those of Hennekam syndrome 1, caused by CCBE1 mutation .…”