A Fourth Example Suggests That Premature Termination Codons in the COL2A1 Gene Are a Common Cause of the Stickler Syndrome: Analysis of the COL2A1 Gene by Denaturing Gradient Gel Electrophoresis

doi:10.1006/geno.1993.1306

Genomics

Volume 17, Issue 1, July 1993, Pages 218-221

https://doi.org/10.1006/geno.1993.1306 Get rights and content

Abstract

A series of oligonucleotide primers was designed to generate polymerase chain reaction products that contained exons 6 to 49 of the human gene for type II procollagen (COL2A1) and that could be used to detect sequence variations by denaturing gradient gel electrophoresis. To improve the sensitivity of the analysis, GC clamps were introduced into one primer of each pair. The procedure successfully detected 10 neutral single-base variations in the gene. In addition, the procedure detected a single-base deletion in exon 43 that introduced a premature termination codon in exon 44 and caused the Stickler syndrome (arthro-ophthalmopathy) in one family. The mutation is the fourth mutation in the COL2A1 gene shown to cause the Stickler syndrome. The mutation is similar to the first three mutations causing the disease in that they also introduced premature termination signals. Since only one mutation introducing a premature termination codon was found in the course of defining 120 or more mutations in type I and III procollagens, the results suggest that such mutations may have a special relationship to the Stickler syndrome.

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Cited by (58)

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In our previous study, potential pathological mutations of RetNet genes were detected in 23.8% (71/298) of probands with early-onset high myopia (eoHM), based on whole exome sequencing (WES). The current study aimed to confirm this finding in an additional 325 probands with eoHM and to clarify its specificity by comparison of 195 probands with late-onset high myopia (loHM). Variants in the 234 RetNet genes were selected from whole-exome sequencing data and were filtered using multistep bioinformatics analyses. Potential pathological variants in 33 genes were detected in 76 of 325 (23.4%) probands with eoHM and 14 of 195 (7.2%) probands with loHM. Thirty-five of the 76 (46.1%) probands with eoHM had mutations in COL2A1, COL11A1, RPGR, and CACNAIF, while only 2/14 (14.3%) probands with eoHM were detected. The mutation frequency and spectrum of RetNet genes in the 325 probands with eoHM were similar to our previous study but were significantly different in 195 probands with loHM (P = 2 × 10⁻⁶ and 0.04). Data from eoHM and loHM strongly suggest that a significant proportion of eoHM is caused by mutations in RetNet genes. These results also provide initial genetic evidence that eoHM is different from loHM. The presence of mutations in 7.2% probands with loHM raises questions about pathogenicity and the variable manifestation of some mutations. The functional studies of the mutations in question and more extensive investigations of related phenotypes in the mutation carriers and their family members may provide valuable information to address these questions.
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Alternative splicing of exon 2 of the collagen type II pre-mRNA results in two forms of expressed collagen chains that affects different functional properties of type II collagen. These two forms may be associated with vitreous pathologies such as those observed in Stickler’s syndrome (STL), Wagner syndrome and Kniest dysplasia (Maumenee and Traboulsi, 1985; Ritvaniemi et al., 1993; Snead and Yates, 1999; Kaarniranta et al., 2006). Thus, vitreous anomaly is one of the clinical signs of these syndromes and dysplasia.
Floaters and flashes are most commonly symptoms of age-related degenerative changes in the vitreous body and posterior vitreous detachment. The etiology and pathogenesis of floaters’ formation is still not well understood. Patients with acute-onset floaters, flashes and defects in their visual field, represent a medical emergency with the need for same day referral to an ophthalmologist. Indirect ophthalmoscopy with scleral indentation is needed in order to find possible retinal break(s), on-time treatment and prevention of retinal detachment. The molecular and genetic pathogenesis, as well as the epidemiology of the ageing changes of the vitreous is summarized here, with view on the several treatment modalities in relation to their success rate and side-effects.
Osteoarthritis in temporomandibular joint of Col2a1 mutant mice
2013, Archives of Oral Biology
Citation Excerpt :
In addition, the ratio of phosphorylated to unphosphorylated eIF2α was increased 4.3-fold in Dmm/+ mice (n = 6) compared to wildtype (n = 5) (p < 0.05) (see Fig. 4). A large number of human chondrodysplasias are caused by mutations in COL2A1.28–33 Most of these mutations, however, are in the triple helical region of the gene.
Col2a1 gene mutations cause premature degeneration of knee articular cartilage in disproportionate micromelia (Dmm) and spondyloepiphesial dysplasia congenita (sedc) mice. The present study analyses the temporomandibular joint (TMJ) in Col2a1 mutant mice in order to provide an animal model of TMJ osteoarthritis (OA) that may offer better understanding of the progression of this disease in humans.
Dmm/+ mice and controls were compared at two, six, nine and 12 months. Craniums were fixed, processed to paraffin sections, stained with Safranin-O/Fast Green, and analysed with light microscopy. OA was quantified using a Mankin scoring procedure. Unfolded protein response (UPR) assay was performed and immunohistochemistry (IHC) was used to assay for known OA biomarkers.
Dmm/+ TMJs showed fissuring of condylar cartilage as early as 6 months of age. Chondrocytes were clustered, leaving acellular regions in the matrix. Significant staining of HtrA1, Ddr2 and Mmp-13 was observed in Dmm/+ mice (p < 0.01). We detected upregulation of the UPR in knee but not TMJ.
Dmm/+ mice are subject to early-onset OA in the TMJ. We observed upregulation of biomarkers and condylar cartilage degradation concomitant with OA. An upregulated UPR may exacerbate the onset of OA. The Dmm/+ mouse TMJ is a viable model for the study of the progression of OA in humans.
Chondrodysplasias
2013, Emery and Rimoin's Principles and Practice of Medical Genetics
A mouse model for Stickler's syndrome: Ocular phenotype of mice carrying a targeted heterozygous inactivation of type II (pro)collagen gene (Col2a1)
2006, Experimental Eye Research
Citation Excerpt :
Different mutations lead to heterogeneous phenotypes of the disease including ocular as well as extraocular signs, such as auditory, skeletal and orofacial symptoms. The most typical ocular manifestations of Stickler syndrome are myopia, vitreous degeneration, and pigmentary degeneration of the retina, retinal ruptures and rhegmatogenous retinal detachment (Ahmad et al., 1991; Brown et al., 1992; Ritvaniemi et al., 1993; Williams et al., 1996; Freddi et al., 2000; Stickler et al., 2001). Interestingly, Stickler syndrome has been found to be the most common inherited cause of retinal detachment (Richards et al., 1996).
The influences of targeted heterozygous inactivation of type II (pro)collagen gene (Col2a1) on eye structures in the 15-month-old C57BL/6JOlaHsd mouse was studied. The eyes were collected from C57BL mice heterozygous for a targeted inactivation of one allele of the Col2a1 gene (Col2a1^+/− mice). The eyes of C57BL mice with normal gene alleles were used as controls (Col2a1^+/+ mice). Ocular histology was analyzed from tissue sections, stained with hematoxylin and eosin, toluidine blue and alcian blue. Type II collagen was localized by immunohistochemistry. Hyaluronan (HA) was stained utilizing the biotinylated complex of the hyaluronan-binding region of aggrecan and link protein (bHABC). The anterior segment of the eye was well-formed in both genotypes, but typical folding of ciliary processes was decreased, while increased stromal extracellular matrix vacuolization was seen in the Col2a1^+/− mice. In the lens of these mice, subcapsular extracellular matrix changes were observed. Differences in retinal structures or the number of the eyes with retinal detachment were not detected between the genotypes. In Col2a1^+/− mice, staining for type II collagen was weaker in cornea, ciliary body, iris, lens, vitreous, retina, choroid and sclera than in the control mice. HA staining was detected in the extraocular tissues, ciliary body, iris and the choroid of both genotypes. HA staining was observed only in the vitreous body of the control animals. Heterozygous inactivation of Col2a1 gene causes structural defects in the murine eye. The observed structural changes in the ciliary body, lens and vitreous of the Col2a1^+/− mice may represent ocular features found in the human Stickler syndrome, where the abnormalities result from COL2A1 gene mutations which lead to functional haploinsufficiency.
Collagens and collagen-related matrix components in the human and mouse eye
2004, Progress in Retinal and Eye Research
The three-dimensional structure of the eye plays an important role in providing a correct optical environment for vision. Much of this function is dependent on the unique structural features of ocular connective tissue, especially of the collagen types and their supramolecular structures. For example, the organization of collagen fibrils is largely responsible for transparency and refraction of cornea, lens and vitreous body, and collagens present in the sclera are largely responsible for the structural strength of the eye. Phylogenetically, most of the collagens are highly conserved between different species, which suggests that collagens also share similar functions in mice and men. Despite considerable differences between the mouse and the human eye, particularly in the proportion of the different tissue components, the difficulty of performing systematic histologic and molecular studies on the human eye has made mouse an appealing alternative to studies addressing the role of individual genes and their mutations in ocular diseases. From a genetic standpoint, the mouse has major advantages over other experimental animals as its genome is better known than that of other species and it can be manipulated by the modern techniques of genetic engineering. Furthermore, it is easy, quick and relatively cheap to produce large quantities of mice for systematic studies. Thus, transgenic techniques have made it possible to study consequences of specific mutations in genes coding for structural components of ocular connective tissues in mice. As these changes in mice have been shown to resemble those in human diseases, mouse models are likely to provide efficient tools for pathogenetic studies on human disorders affecting the extracellular matrix. This review is aimed to clarify the role of collagenous components in the mouse and human eye with a closer look at the new findings of the collagens in the cartilage and the eye, the so-called “cartilage collagens”.

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Short CommunicationA Fourth Example Suggests That Premature Termination Codons in the COL2A1 Gene Are a Common Cause of the Stickler Syndrome: Analysis of the COL2A1 Gene by Denaturing Gradient Gel Electrophoresis

Abstract

Short Communication
A Fourth Example Suggests That Premature Termination Codons in the COL2A1 Gene Are a Common Cause of the Stickler Syndrome: Analysis of the COL2A1 Gene by Denaturing Gradient Gel Electrophoresis