Functional domains of VLDLR with positions of the mutations relative to the exons (A), domains (B), and the analysis of VLDLR transcript (C and D). (A) The gene consists of 19 exons. Arrows indicate the locations of the mutations. (B) VLDLR consists of ligand-binding type repeat (LBTR), epidermal growth factor repeat (EGFR) I–III, YWTD β-propeller (YWTD), O-linked sugar domain (OLSD), transmembrane domain (TD), and cytoplasmic domain (CD) () (www.expasy.org/uniprot/P98155). (C) Restriction-based analysis with HphI revealed the presence of only the mutant (347 bp) and both the mutant and wild type (396 and 347 bp; please note that the 49-bp fragment is not visible) VLDLR transcripts in patient VI:20 and carrier V:18 (both from family A), respectively. M is a DNA size marker. (D) Quantitative RT-PCR analysis of VLDLR transcript from peripheral blood samples of all probands in families A and D and controls was performed. Relative expression ratios were normalized according to the housekeeping gene GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and the endothelial marker KDR (kinase insert domain receptor). ΔCt values were calculated from duplicate samples and were converted to linear scale (). Control denotes “VLDLR expression in controls,” VLDLR-GAPDH denotes “VLDLR expression in patients normalized to GAPDH,” and finally VLDLR-KDR denotes “VLDLR expression in patients normalized to KDR.”