Fig. 2

Multiple truncated and non functional CISD2 isoforms. a The 5′-RACE extended an expected 500 nt PCR product in the healthy control and two products of approximately 400 and 800–900 bases in the probands. b A schematic representation of the exon structure of wild-type (a) and mutant (a1-a3) CISD2 variants, and putative pre-mRNAs (b1 and b2). The homozygous mutation in the donor splice of intron 1 generated multiple transcripts characterized by the whole or partial absence of exon 1 (a1-a3) and inefficiently spliced pre-mRNAs retaining a large segment of intron 1 (b1 and b2). c The sequencing resulting from the 400 nt 5′-RACE products. The cDNA and amino acids sequences are shown. The exons are indicated in uppercase letters and alternate colours. The 5′-UTR is indicated in black lowercase letters. a1 and a2: Short stretches of genomic DNA are joined to exon 2 perfectly spliced with exon 3. Genomic coordinates: Chr4:102,828,100–102,828,185 and Chr4:102,861,131–102,861,212, for a1 and a2 respectively. CpG island coordinates: Chr4:102,826,475–102,828,235. Pugo gene: Chr4:102,827,193–102,829,052. The ATG, predicted in the beginning of exon 2 (Chr4:102,885,223; NG_008636.2:21,246), shifted the open reading frame and introduced a downstream amino acid changes (underscored) in addition to premature stop. a3: The partial absence of exon 1 (Chr4:102,869,118–102,869,187; NG_008636.2:5141–5210) caused a frame shift, amino acids changes (underscored) and a premature stop. The transcriptional start sites are in bold letters and underscored, and they are located at 111 and 43 nt upstream the ATG for patient 1 and 2, respectively (position at 4997 and 5064 referred to NG_008636.2; position at 102868974 and 102,869,041 referred to Chr4). The SNP rs223332 (NG_008636.2: g.5052G > T) is indicated in red and lowercase letter. d The frequency of mutant isoforms in WFS2 patients