Expression analysis of endoglin missense and truncation mutations: insights into protein structure and disease mechanisms

Hereditary hemorrhagic telangiectasia (HHT) is an inherited autosomal domin ant vascular dysplasia caused by mutations in either endoglin (HHT1) or act ivin-like kinase receptor-1 (ALK-1) (HHT2). The majority of the mutations i n endoglin cause frameshifts and premature stop codons. Although initial re ports suggested a dominant-negative model for HHT1, more recent reports hav e suggested that mutations in endoglin lead to haploinsufficiency. In this study, we investigated six different missense mutations and two truncation mutations in the endoglin gene to examine whether mechanisms other than hap loinsufficiency might be involved in HHT1. Expression of the missense mutan ts alone revealed that they are misfolded and that most show no cell surfac e expression. When co-expressed with wildtype endoglin, the missense mutant s are able to dimerize with the normal endoglin protein and are trafficked to the cell surface, We also show that although one truncation mutation act s through haploinsufficiency, the other acts in a dominant-negative way. Th is implies that either dominantnegative protein interactions or haploinsuff iciency can cause HHT1. The biochemical analyses for the different mutation s suggest that the endoglin N-terminus is important for correct protein fol ding and that cysteine residues in the first 350 amino acids are involved i n intramolecular disulfide bonds, whereas cysteines located closer to the C -terminus of the extracellular domain are responsible for intermolecular di sulfide bond dimerization.