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Figure 2 | BMC Medical Genetics

Figure 2

From: Clinical and genetic analyses of three Korean families with hereditary hemorrhagic telangiectasia

Figure 2

Functionality of the aberrant translation start codon. (A) Schematic representation of the pENG-luc constructs. Wild-type (WT) ENG promoter produces ENG mRNA including the 413-nt-5'-UTR. pENG-luc(WT) was designed to code for luciferase mRNA containing the same ENG 5'-UTR under the control of the immediate early promoter of CMV (CMV-IE). In pENG-luc(M), the 'C' at position -127 in pENG-luc(WT) was replaced with 'T' (underlined), which generated the aberrant translation start codon. The substitution was found in the mutant allele of HHT patients in Family 2. The aberrant translation start codon putatively produces a 72-amino acid-protein out of frame with the luciferase coding sequence. In pENG-luc(M+1), a 'C' nucleotide (bold) was inserted between -41 and -40 of pENG-luc(M), thus producing an aberrant translation start codon that was in frame with the luciferase coding sequence. Nucleotides are numbered with c.1 corresponding to the 'A' of the ATG translation start codon in the reference sequence [GenBank:NM_000118.2]. (B) Bar graph illustrating the results of luciferase assays. HepG2 cells were transfected with pENG-luc plasmids [pENG-luc(WT), pENG-luc(M), or pENG-luc(M+1)] plus the β-galactosidase plasmid. Luciferase activity was normalized to β-galactosidase activity. The luciferase activity of pENG-luc(M) was significantly reduced compared to that of pENG-luc(WT). The activity was restored to levels even higher than WT levels in pENG-luc(M+1), which produced an in-frame luciferase fusion protein (44 aa-luciferase). Separate transfections were performed for each of three separate experiments (n = 9). Data represent mean ± SD. *p < .0001, as determined by Student's t test.

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