Genotype-phenotype analysis of <i>COL2A1</i> and <i>COL11A1</i> de novo mutations leading to Stickler syndrome types 1 and 2_Chinese Journal of Ocular Fundus Diseases

Authors：

Li Jiayu ¹ , Li Chunhua ¹ , Sun Caihong ¹ , Fang Wei ¹ , Qi Xiaolong ¹ , Li Wenjin ^1,2 , Zhang Shaochi ¹ , Zhang Wen ¹ , Li Rui ² ,  Zhuang Wenjuan ^1,2

1. Department of Ophthalmology, People's Hospital of Ningxia Hui Autonomous Region (Affiliated Hospital of Ningxia Medical University), Ningxia Medical University, Yinchuan 750011, China;
2. Third Clinical College, Ningxia Medical University, Yinchuan 750041, China;

Corresponding author：

Zhuang Wenjuan, Email: zh_wenj@163.com

Keywords：

Stickler syndrome; COL2A1 gene; COL11A1 gene; De novo mutation; Genotype-phenotype study

DOI：

10.3760/cma.j.cn511434-20241112-00422

Video：

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Objective To observe and analyze the clinical phenotype and genetic characteristics of COL2A1 and COL11A1 de novo mutation (DNM) related Stickler syndrome type I and II patients. Methods A family-based cohort study. From December 2023 to November 2024, 4 patients (all probands) with Stickler syndrome diagnosed by clinical and genetic testing in Department of Ophthalmology of People's Hospital of Ningxia Hui Autonomous Region and their parents (8 cases) were included in the study. The patients came from 4 unrelated families. A detailed medical history was taken, and the patients underwent best-corrected visual acuity (BCVA), refraction, and fundus color photography examinations. Systemic examinations included the oral and facial regions, skeletal, joints, and hearing. Peripheral venous blood samples were collected from the patients and their parents, and genomic DNA was extracted. Whole-exome sequencing was used to screen for pathogenic genes and their loci, which were then validated by Sanger sequencing and combined with segregation analysis in the families to identify candidate gene mutation sites. The candidate variants were assessed for pathogenicity according to the American College of Medical Genetics and Genomics (ACMG) criteria and guidelines for the classification of genetic variants. Additionally, cross-species conservation analysis was performed to determine the evolutionary conservation of wild-type amino acids, and protein three-dimensional modeling techniques were used to characterize the spatial conformational changes of the variant proteins and the alterations in their local hydrogen bond networks. Results Among the 4 patients, there were 2 males and 2 females; their ages ranged from 3 to 12 years. There were 2 cases of Stickler syndrome type I (proband of families 1 and 2) and 2 cases of type II (proband of families 3 and 4). The diopters ranged from －8.00 to－11.0 D. BCVA ranged from no light perception to 0.6^-. There were 2 cases each of vitreous membrane-like and “bead-like” opacity. Three cases showed peripapillary atrophy arcs and leopard pattern changes in the retina; one case had bilateral retinal detachment with a large macular hole in the left eye, which had previously been treated with vitrectomy surgery. One case had bilateral sensorineural hearing loss. There were 3 cases of simple micrognathia; one case had a flat nasal bridge, short nose, midface depression, and micrognathia. Two cases had excessive elbow joint extension. The phenotypes of the parents of the 4 patients were normal. Genetic testing results revealed that the probands of families 1 and 2 carried COL2A1 gene c.85+1G>C (M1) splice site variant and c.3950_3951insA (p.M1317Ifs*48) (M2) frameshift variant, respectively; the probands of families 3 and 4 carried COL11A1 gene (NM_001854.4) c.2549 G>T (p.G850V) (M3) missense variant and c.3816+6T>C (M4) splice site variant, respectively. The parents did not carry the related gene variants. Among them, M2, M3, and M4 are newly reported DNM. According to the ACMG guidelines, they were all considered likely pathogenic. The cross-species conservation analysis results showed that the wild-type amino acid of the COL11A1 gene M3 missense variant was highly conserved across multiple different species. Protein local structure modeling analysis revealed that the COL2A1 gene M2 frameshift variant and the COL11A1 gene M3 missense variant significantly altered the tertiary structure conformation of the protein, leading to abnormal spatial arrangement and hydrogen bond network in the key functional domains Conclusion The COL2A1 gene M1 splice site variant, M2 frameshift variant, and the COL11A1 gene M3 missense variant, M4 splice site variant are respectively the potential pathogenic genes for families 1, 2, and families 3, 4; leading to the onset of Stickler syndrome type I in families 1 and 2, and type II in families 3 and 4.

1.	Stickler GB, Belau PG, Farrell FJ, et al. Hereditary progressive artho-ophthalmopathy[J]. Mayo Clin Proc, 1965, 40: 433-455.
2.	Soh Z, Richards AJ, McNinch A, et al. Dominant Stickler syndrome[J/OL]. Genes (Basel), 2022, 13(6): 1089[2022-06-18]. https://pubmed.ncbi.nlm.nih.gov/35741851/. DOI: 10.3390/genes13061089.
3.	Chen X, Ju Y, Gao F, et al. Rhegmatogenous retinal detachment secondary to type I Stickler syndrome: diagnosis, treatment and long-term outcomes[J/OL]. Genes (Basel), 2024, 15(11): 1455[2024-11-11]. https://pubmed.ncbi.nlm.nih.gov/39596655/. DOI: 10.3390/genes15111455.
4.	Robin NH, Moran RT, Ala-Kokko L. Stickler syndrome[M]. GeneReviews® [Internet]. Seattle (WA): University of Washington, Seattle, 2020: 1993-2025.
5.	Acke FR, Malfait F, Vanakker OM, et al. Novel pathogenic COL11A1/COL11A2 variants in Stickler syndrome detected by targeted NGS and exome sequencing[J]. Mol Genet Metab, 2014, 113(3): 230-235. DOI: 10.1016/j.ymgme.2014.09.001.
6.	Nixon TRW, Richards AJ, Martin H, et al. Autosomal recessive Stickler syndrome[J/OL]. Genes (Basel), 2022, 13(7): 1135[2022-06-24]. https://pubmed.ncbi.nlm.nih.gov/35885918/. DOI: 10.3390/genes13071135.
7.	Aylward B, daCruz L, Ezra E, et al. Stickler syndrome[J]. Ophthalmology, 2008, 115(9): 1636-1638. DOI: 10.1016/j.ophtha.2008.04.018.
8.	Richards AJ, McNinch A, Martin H, et al. Stickler syndrome and the vitreous phenotype: mutations in COL2A1 and COL11A1[J/OL]. Hum Mutat, 2010, 31(6): E1461-1471[2010-06-01]. https://pubmed.ncbi.nlm.nih.gov/20513134/. DOI: 10.1002/humu.21257.
9.	Spickett C, Hysi P, Hammond CJ, et al. Deep intronic sequence variants in COL2A1 affect the alternative splicing efficiency of exon 2, and may confer a risk for rhegmatogenous retinal detachment[J]. Hum Mutat, 2016, 37(10): 1085-1096. DOI: 10.1002/humu.23050.
10.	Snead MP, Richards AJ, McNinch AM, et al. Stickler syndrome - lessons from a national cohort[J]. Eye (Lond), 2022, 36(10): 1966-1972. DOI: 10.1038/s41433-021-01776-8.
11.	Richards AJ, Martin S, Yates JR, et al. COL2A1 exon 2 mutations: relevance to the Stickler and Wagner syndromes[J]. Br J Ophthalmol, 2000, 84(4): 364-371. DOI: 10.1136/bjo.84.4.364.
12.	Richards AJ, Baguley DM, Yates JR, et al. Variation in the vitreous phenotype of Stickler syndrome can be caused by different amino acid substitutions in the X position of the type Ⅱ collagen Gly-X-Y triple helix[J]. Am J Hum Genet, 2000, 67(5): 1083-1094. DOI: 10.1016/S0002-9297(07)62938-3.
13.	Richards S, Aziz N, Bale S, et al. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology[J]. Genet Med, 2015, 17(5): 405-424. DOI: 10.1038/gim.2015.30.
14.	Li W, He XD, Yang ZT, et al. De novo mutations contributes approximately 7% of pathogenicity in inherited eye diseases[J]. Invest Ophthalmol Vis Sci, 2023, 64(2): 1-5. DOI: 10.1167/iovs.64.2.5.
15.	Brodsky B, Persikov AV. Molecular structure of the collagen triple helix[J]. Adv Protein Chem, 2005, 70: 301-339. DOI: 10.1016/S0065-3233(05)70009-7.
16.	Bella J, Hulmes DJ. Fibrillar collagens[J]. Subcell Biochem, 2017, 82: 457-490. DOI: 10.1007/978-3-319-49674-0_14.
17.	Chung HJ, Jensen DA, Gawron K, et al. R992C (p. R1192C) substitution in collagen Ⅱ alters the structure of mutant molecules and induces the unfolded protein response[J]. J Mol Biol, 2009, 390(2): 306-318. DOI: 10.1016/j.jmb.2009.05.004.
18.	Liang G, Lian C, Huang D, et al. Endoplasmic reticulum stress-unfolding protein response-apoptosis cascade causes chondrodysplasia in a col2a1 p. Gly1170Ser mutated mouse model[J/OL]. PLoS One, 2014, 9(1): e86894[2014-01-17]. https://pubmed.ncbi.nlm.nih.gov/24475193/. DOI: 10.1371/journal.pone.0086894.
19.	Pace CN, Fu H, Lee Fryar K, et al. Contribution of hydrogen bonds to protein stability[J]. Protein Sci, 2014, 23(5): 652-661. DOI: 10.1002/pro.2449.
20.	Richards AJ, Laidlaw M, Whittaker J, et al. High efficiency of mutation detection in type 1 stickler syndrome using a two-stage approach: vitreoretinal assessment coupled with exon sequencing for screening COL2A1[J]. Hum Mutat, 2006, 27(7): 696-704. DOI: 10.1002/humu.20347.
21.	Alexander P, Snead MP. Prevention of blindness in Stickler syndrome[J/OL]. Genes (Basel), 2022, 13(7): 1150[2022-06-26]. https://pubmed.ncbi.nlm.nih.gov/35885933/. DOI: 10.3390/genes13071150.
22.	Richards AJ, Fincham GS, McNinch A, et al. Alternative splicing modifies the effect of mutations in COL11A1 and results in recessive type 2 Stickler syndrome with profound hearing loss[J]. J Med Genet, 2013, 50(11): 765-771. DOI: 10.1136/jmedgenet-2012-101499.
23.	Schlessinger A, Rost B. Protein flexibility and rigidity predicted from sequence[J]. Proteins, 2005, 61(1): 115-126. DOI: 10.1002/prot.20587.
24.	Jin ZB, Li Z, Liu Z, et al. Identification of de novo germline mutations and causal genes for sporadic diseases using trio-based whole-exome/genome sequencing[J]. Biol Rev Camb Philos Soc, 2018, 93(2): 1014-1031. DOI: 10.1111/brv.12383.
25.	Veltman JA, Brunner HG. De novo mutations in human genetic disease[J]. Nat Rev Genet, 2012, 13(8): 565-575. DOI: 10.1038/nrg3241.
26.	Takata A, Miyake N, Tsurusaki Y, et al. Integrative analyses of de novo mutations provide deeper biological insights into autism spectrum disorder[J]. Cell Rep, 2018, 22(3): 734-747. DOI: 10.1016/j.celrep.2017.12.074.
27.	Jin SC, Homsy J, Zaidi S, et al. Contribution of rare inherited and de novo variants in 2, 871 congenital heart disease probands[J]. Nat Genet, 2017, 49(11): 1593-1601. DOI: 10.1038/ng.3970.
28.	Bishop MR, Diaz Perez KK, Sun M, et al. Genome-wide enrichment of de novo coding mutations in orofacial cleft trios[J]. Am J Hum Genet, 2020, 107(1): 124-136. DOI: 10.1016/j.ajhg.2020.05.018.
29.	Huang L, Chen C, Wang Z, et al. Mutation spectrum and de novo mutation analysis in Stickler syndrome patients with high myopia or retinal detachment[J/OL]. Genes (Basel), 2020, 11(8): 882[2020-08-03]. https://pubmed.ncbi.nlm.nih.gov/32756486/. DOI: 10.3390/genes11080882.

1. Stickler GB, Belau PG, Farrell FJ, et al. Hereditary progressive artho-ophthalmopathy[J]. Mayo Clin Proc, 1965, 40: 433-455.
2. Soh Z, Richards AJ, McNinch A, et al. Dominant Stickler syndrome[J/OL]. Genes (Basel), 2022, 13(6): 1089[2022-06-18]. https://pubmed.ncbi.nlm.nih.gov/35741851/. DOI: 10.3390/genes13061089.
3. Chen X, Ju Y, Gao F, et al. Rhegmatogenous retinal detachment secondary to type I Stickler syndrome: diagnosis, treatment and long-term outcomes[J/OL]. Genes (Basel), 2024, 15(11): 1455[2024-11-11]. https://pubmed.ncbi.nlm.nih.gov/39596655/. DOI: 10.3390/genes15111455.
4. Robin NH, Moran RT, Ala-Kokko L. Stickler syndrome[M]. GeneReviews® [Internet]. Seattle (WA): University of Washington, Seattle, 2020: 1993-2025.
5. Acke FR, Malfait F, Vanakker OM, et al. Novel pathogenic COL11A1/COL11A2 variants in Stickler syndrome detected by targeted NGS and exome sequencing[J]. Mol Genet Metab, 2014, 113(3): 230-235. DOI: 10.1016/j.ymgme.2014.09.001.
6. Nixon TRW, Richards AJ, Martin H, et al. Autosomal recessive Stickler syndrome[J/OL]. Genes (Basel), 2022, 13(7): 1135[2022-06-24]. https://pubmed.ncbi.nlm.nih.gov/35885918/. DOI: 10.3390/genes13071135.
7. Aylward B, daCruz L, Ezra E, et al. Stickler syndrome[J]. Ophthalmology, 2008, 115(9): 1636-1638. DOI: 10.1016/j.ophtha.2008.04.018.
8. Richards AJ, McNinch A, Martin H, et al. Stickler syndrome and the vitreous phenotype: mutations in COL2A1 and COL11A1[J/OL]. Hum Mutat, 2010, 31(6): E1461-1471[2010-06-01]. https://pubmed.ncbi.nlm.nih.gov/20513134/. DOI: 10.1002/humu.21257.
9. Spickett C, Hysi P, Hammond CJ, et al. Deep intronic sequence variants in COL2A1 affect the alternative splicing efficiency of exon 2, and may confer a risk for rhegmatogenous retinal detachment[J]. Hum Mutat, 2016, 37(10): 1085-1096. DOI: 10.1002/humu.23050.
10. Snead MP, Richards AJ, McNinch AM, et al. Stickler syndrome - lessons from a national cohort[J]. Eye (Lond), 2022, 36(10): 1966-1972. DOI: 10.1038/s41433-021-01776-8.
11. Richards AJ, Martin S, Yates JR, et al. COL2A1 exon 2 mutations: relevance to the Stickler and Wagner syndromes[J]. Br J Ophthalmol, 2000, 84(4): 364-371. DOI: 10.1136/bjo.84.4.364.
12. Richards AJ, Baguley DM, Yates JR, et al. Variation in the vitreous phenotype of Stickler syndrome can be caused by different amino acid substitutions in the X position of the type Ⅱ collagen Gly-X-Y triple helix[J]. Am J Hum Genet, 2000, 67(5): 1083-1094. DOI: 10.1016/S0002-9297(07)62938-3.
13. Richards S, Aziz N, Bale S, et al. Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology[J]. Genet Med, 2015, 17(5): 405-424. DOI: 10.1038/gim.2015.30.
14. Li W, He XD, Yang ZT, et al. De novo mutations contributes approximately 7% of pathogenicity in inherited eye diseases[J]. Invest Ophthalmol Vis Sci, 2023, 64(2): 1-5. DOI: 10.1167/iovs.64.2.5.
15. Brodsky B, Persikov AV. Molecular structure of the collagen triple helix[J]. Adv Protein Chem, 2005, 70: 301-339. DOI: 10.1016/S0065-3233(05)70009-7.
16. Bella J, Hulmes DJ. Fibrillar collagens[J]. Subcell Biochem, 2017, 82: 457-490. DOI: 10.1007/978-3-319-49674-0_14.
17. Chung HJ, Jensen DA, Gawron K, et al. R992C (p. R1192C) substitution in collagen Ⅱ alters the structure of mutant molecules and induces the unfolded protein response[J]. J Mol Biol, 2009, 390(2): 306-318. DOI: 10.1016/j.jmb.2009.05.004.
18. Liang G, Lian C, Huang D, et al. Endoplasmic reticulum stress-unfolding protein response-apoptosis cascade causes chondrodysplasia in a col2a1 p. Gly1170Ser mutated mouse model[J/OL]. PLoS One, 2014, 9(1): e86894[2014-01-17]. https://pubmed.ncbi.nlm.nih.gov/24475193/. DOI: 10.1371/journal.pone.0086894.
19. Pace CN, Fu H, Lee Fryar K, et al. Contribution of hydrogen bonds to protein stability[J]. Protein Sci, 2014, 23(5): 652-661. DOI: 10.1002/pro.2449.
20. Richards AJ, Laidlaw M, Whittaker J, et al. High efficiency of mutation detection in type 1 stickler syndrome using a two-stage approach: vitreoretinal assessment coupled with exon sequencing for screening COL2A1[J]. Hum Mutat, 2006, 27(7): 696-704. DOI: 10.1002/humu.20347.
21. Alexander P, Snead MP. Prevention of blindness in Stickler syndrome[J/OL]. Genes (Basel), 2022, 13(7): 1150[2022-06-26]. https://pubmed.ncbi.nlm.nih.gov/35885933/. DOI: 10.3390/genes13071150.
22. Richards AJ, Fincham GS, McNinch A, et al. Alternative splicing modifies the effect of mutations in COL11A1 and results in recessive type 2 Stickler syndrome with profound hearing loss[J]. J Med Genet, 2013, 50(11): 765-771. DOI: 10.1136/jmedgenet-2012-101499.
23. Schlessinger A, Rost B. Protein flexibility and rigidity predicted from sequence[J]. Proteins, 2005, 61(1): 115-126. DOI: 10.1002/prot.20587.
24. Jin ZB, Li Z, Liu Z, et al. Identification of de novo germline mutations and causal genes for sporadic diseases using trio-based whole-exome/genome sequencing[J]. Biol Rev Camb Philos Soc, 2018, 93(2): 1014-1031. DOI: 10.1111/brv.12383.
25. Veltman JA, Brunner HG. De novo mutations in human genetic disease[J]. Nat Rev Genet, 2012, 13(8): 565-575. DOI: 10.1038/nrg3241.
26. Takata A, Miyake N, Tsurusaki Y, et al. Integrative analyses of de novo mutations provide deeper biological insights into autism spectrum disorder[J]. Cell Rep, 2018, 22(3): 734-747. DOI: 10.1016/j.celrep.2017.12.074.
27. Jin SC, Homsy J, Zaidi S, et al. Contribution of rare inherited and de novo variants in 2, 871 congenital heart disease probands[J]. Nat Genet, 2017, 49(11): 1593-1601. DOI: 10.1038/ng.3970.
28. Bishop MR, Diaz Perez KK, Sun M, et al. Genome-wide enrichment of de novo coding mutations in orofacial cleft trios[J]. Am J Hum Genet, 2020, 107(1): 124-136. DOI: 10.1016/j.ajhg.2020.05.018.
29. Huang L, Chen C, Wang Z, et al. Mutation spectrum and de novo mutation analysis in Stickler syndrome patients with high myopia or retinal detachment[J/OL]. Genes (Basel), 2020, 11(8): 882[2020-08-03]. https://pubmed.ncbi.nlm.nih.gov/32756486/. DOI: 10.3390/genes11080882.

Chinese Journal of Ocular Fundus Diseases

Latest ArticlesGenotype-phenotype analysis of COL2A1 and COL11A1 de novo mutations leading to Stickler syndrome types 1 and 2

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