Abstract
Molecular investigations on a young patient and her family were undertaken to identify the molecular defect responsible for a mild form of osteogenesis imperfecta (OI) with blue sclerae, dentinogenesis imperfecta and joint laxity. Analysis of collagenous proteins from the proband's fibroblasts showed the presence of two populations of α2(I) chains, one normal and one migrating faster on SDS gels, thereby suggesting deletion of amino acid sequences. The faster migrating chains were retained mainly in the cell layer and not found in the extracellular matrix deposited by cultured fibroblasts. Chemical cleavage of mismatch (CCM) analysis on the patient's proα2(I) mRNA:normal cDNA heteroduplexes localized the molecular defect. cDNA sequencing revealed a deletion of exon 20 (54 bp) in about half of the molecules. Genomic DNA sequencing revealed heterozygosity for a G-to-C transversion of the last nucleotide of intron 19, which changed the 3′ consensus splicing site. As a consequence proα2(I)mRNA was abnormally spliced from the last codon of exon 19 to the first codon of exon 21. To our knowledge, this is the first acceptor site mutation so far described in an OI patient. Restriction analysis indicated that the mutation was present also in three other affected family members. The full sequence of COL1A2 introns 19 and 20 are reported.
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References
Bateman JF, Golub S (1990) Assessment of procollagen processing defects by fibroblasts cultured in the presence of dextran sulfate. Biochem J 267:573–577.
Bateman JF, Lamande SR, Hannagan M, Moeller I, Dahl HHM, Cole WG (1993) Chemical cleavage method for the detection of RNA base changes: experience in the application to collagen mutations in osteogenesis imperfecta. Am J Med Genet 45:233–240.
Bonadio J, Ramirez F, Barr M (1990) An intron mutation in the human α1(I) collagen gene alters the efficiency of pre-mRNA splicing and is associated with osteogenesis imperfecta type II. J Biol Chem 265:2262–2268.
Byers PH, Wallis GA, Willing MC (1991) Ostenogenesis imperfecta: translation of mutation to phenotype. J Med Genet 28:433–442.
Chiodo AA, Hockey A, Cole WG (1992) A base substitution at the splice acceptor site on intron 5 of the COL1A2 gene activates a cryptic splice site within exon 6 and generates abnormal type I procollagen in a patient with Ehlers-Danlos syndrome type VII. J Biol Chem 267:6361–6369.
Cotton RGH, Rodrigues NR, Campbell RD (1988) Reactivity of cytosine and thymine in single base pair mismatches with hydroxylamine and osmium tetroxide and its application to the study of mutations. Proc Natl Acad Sci USA 85:4397–4401.
D'Alessio M, Ramirez F, Blumberg BD, Wirtz MK, Rao VH, Godfrey MD, Hollister DW (1991) Characterization of a COL1A1 splicing defect in a case of Ehlers-Danlos syndrome type VII: further evidence of molecular homogeneity. Am J Hum Genet 49:400–406.
Filie JD, Orrison BM, Wang Q, Lewis B, Marini JC (1993) A de novo G+1-A mutation at the α2(I) exon 16 splice donor site causes skipping of exon 16 in the cDNA of one allele of an OI type IV proband. Hum Mutat 2:380–388.
Krawczak M, Reiss J, Cooper DN (1992) The mutational spectrum of single base-pair substitutions in mRNA splice junctions of human genes: causes and consequences. Hum Genet 90:41–54.
Kuivaniemi H, Tromp G, Chu ML, Prockop DJ (1988a) Structure of a full-length cDNA clone for the preproα2(I) chain of human type I procollagen. Biochem J 252:633–640.
Kuivaniemi H, Sabol C, Tromp G, Sippola-Thiele M, Prockop DJ (1988b) A 19-base pair deletion in the pro-α2(I) gene of type I procollagen that causes in-frame RNA splicing from exon 10 to exon 12 in a proband with atypical osteogenesis imperfecta and in his asymptomatic mother. J Biol Chem 263:11407–11413.
Kuivaniemi H, Kontusaari S, Tromp G, Zhao M, Sabol C, Prockop DJ (1990) Identical G+1 to A mutations in three different introns of the type III procollagen gene (COL3A1) produce different patterns of RNA splicing in three variants of Ehlers-Danlos syndrome IV. J Biol Chem 265:12067–12074.
Kuivaniemi H, Tromp G, Prockop DJ (1991) Mutations in collagen genes: causes of rare and some common diseases in humans. FASEB J 5:2052–2060.
Lee B, Vitale E, Superti-Furga A, Steinmann B, Ramirez F (1991) G to T transversion at position +5 of a splice donor site causes skipping at the preceding exon in the type III procollagen transcripts of a patient with Ehlers-Danlos syndrome type IV. J Biol Chem 266:5256–5259.
Mottes M, Sangalli A, Valli M, Gomez Lira M, Tenni R, Buttitta P, Pignatti PF, Cetta G (1992) Mild dominant osteogenesis imperfecta with intrafamiliar variability: the cause is a serine for glycine α1(I) 901 substitution in a type I collagen gene. Hum Genet 89:480–484.
Nicholls AC, Oliver J, Renouf DV, Heath DA, Pope FM (1992) The molecular defect in a family with mild atypical osteogenesis imperfecta and extreme joint hypermobility: exon skipping caused by an 11-bp deletion from an intron in one COL1A2 allele. Hum Genet 88:627–633.
Sillence DO, Senn A, Danks DM (1979) Genetic heterogeneity in osteogenesis imperfecta. J Med Genet 16:101–116.
Sippola M, Kaffe S, Prockop DJ (1984) A heterozygous defect for structurally altered proα2(I) chains of type I procollagen in a mild variant of osteogenesis imperfecta. J Biol Chem 259:14094–14100.
Superti-Furga A, Raghunath M, Pistone FM, Romano C, Steinmann B (1993) An intronic deletion leading to skipping of exon 21 of COL1A2 in a boy with mild osteogenesis imperfecta. Connect Tissue Res 29:31–40.
Tenni R, Rossi A, Valli M, Mottes M, Pignatti PF, Cetta G (1990) Anomalous cysteine in type I collagen: localisation by chemical cleavage of the protein using 2-nitro-5-thio-cyanobenzoic acid and by mismatch analysis of cDNA heteroduplexes. Matrix 10:20–26.
Tiller GE, Weis MA, Eyre DR, Rimoin DL, Cohn DH (1992) An RNA splicing mutation in the gene for type II collagen (COL2A1) produces spondyloepiphyseal dysplasia congenita (SEDC) (abstract). Am J Hum Genet 51 [Suppl]: A37 (Abstract 136).
Tromp G, Prockop DJ (1988) Single base mutation in the proα2(I) collagen gene that causes efficient splicing of RNA from exon 27 to exon 29 synthesis of shortened but in-frame proα2(I) chain. Proc Natl Acad Sci USA 85:5254–5258.
Valli M, Mottes M, Tenni R, Sangalli A, Gomez Lira M, Rossi A, Antoniazzi F, Cetta G, Pignatti PF (1991) A de novo G-to-T transversion in a proα1(I) collagen gene for a moderate case of osteogenesis imperfecta. J Biol Chem 266:1872–1878.
Valli M, Rossi A, Forlino A, Tenni R, Cetta G (1993) Extracellular matrix deposition in cultured dermal fibroblasts from probands affected by osteogenesis imperfecta. Matrix 13:275–280.
Vasan NS, Kuivaniemi H, Vogel B, Minor RR, Wootton JAM, Tromp G, Weksberg R, Prockop DJ (1991) A mutation of the proα2(I) gene (COL1A2) for type I procollagen in Ehlers-Danlos syndrome type VII: evidence suggesting that skipping of exon 6 in RNA splicing may be a common cause of the phenotype. Am J Hum Genet 48:305–317.
Wenstrup R, Shrago A, Phillips C, Byers P, Cohn D (1990) Analysis for mutations in α2(I) chains of type I collagen by α2(I)-specific cDNA synthesis and polymerase chain reaction. Ann NY Acad Sci 580:546–548.
Winterpacht A, Hubert M, Schwarze U, Mundlos S, Spranger J, Zabel BU (1993) Kniest and Stickler dysplasia phenotypes caused by collagen type II gene (COL2A1) defect. Nature Genet 3:323–326.
Wu Y, Kuivaniemi H, Tromp G, Strobel D, Romanic AM, Prockop DJ (1993) Temperature sensitivity of aberrant RNA splicing with a mutation in the G+5 position of intron 37 of the gene for type III procollagen from a patient with Ehlers-Danlos syndrome type IV. Hum Mutat 2:28–36.
Zhuang J, Tromp G, Kuivaniemi H, Nakayasu K, Prockop DJ (1993) Deletion of 19 base pairs in intron 13 of the gene for the proα2(I) chain of type I procollagen (COL1A2) causes exon skipping in a proband with type I osteogenesis imperfecta. 91:210–216.
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Mottes, M., Sangalli, A., Valli, M. et al. A base substitution at IVS-19 3′-end splice junction causes exon 20 skipping in proα2(I) collagen mRNA and produces mild osteogenesis imperfecta. Hum Genet 93, 681–687 (1994). https://doi.org/10.1007/BF00201570
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DOI: https://doi.org/10.1007/BF00201570