In an integrated program of laboratory and clinical investigation, we study the molecular biology of the heritable connective tissue disorders osteogenesis imperfecta (OI) and Ehlers-Danlos syndrome (EDS). Our objective is to elucidate the mechanisms by which the primary gene defect causes skeletal fragility and other connective tissue symptoms and then apply the knowledge gained from our studies to the treatment of children with these conditions. Structural defects of the heterotrimeric type I collagen molecule are well known to cause the dominant bone disorder osteogenesis imperfecta. A severe recessive form of OI was first postulated in 1979. We hypothesized that the cause of recessive OI with abnormal collagen biochemistry and normal collagen gene sequence would involve a gene(s) whose products interacted with type I collagen. Several years ago the BEMB identified defects in two components of the collagen prolyl 3-hydroxylation complex, CRTAP and P3H1 (encoded by LEPRE1) as the cause of recessive OI. Our work has generated a new paradigm for collagen-related disorders of matrix, in which structural defects in collagen cause dominant OI, while defects in the components of a complex in the endoplasmic reticulum that modifies collagen cause recessive OI. In the expanded nosology for OI, defects in CRTAP and LEPRE1 are designated as types VII (OMIM #610682) and VIII (OMIM #610915) OI, respectively. Recessive OI is now a major area of investigation for the BEMB. The phenotypes of types VII and VIII OI are distinct from classical dominant OI, but difficult to distinguish from each other. Both groups of children have severe/lethal OI with white sclerae, normal or small head circumference, rhizomelia, metacarpal shortening and severe undertubulation of long bones. Biochemically, both groups have normal collagen sequences with absence of 3-hydroxylation of the Pro986 residue, but full overmodification of the helical prolines and lysines by prolyl 4-hydroxylase and lysly hydroxylase. This overmodification of the helix was unexpected and indicates that absence of the components of the 3-hydroxylation complex leads to delayed folding of the collagen helix. We have now shown that the basis of the phenotypic and collagen biochemical similarity of types VII and VIII OI is that CRTAP and P3H1 are mutually protectively in the complex. We observed that CRTAP is severely reduced and P3H1 is absent from cell lysates which have a defect in either component of the complex, although the transcript level of the normal component is not reduced. The interpretation of mutual protection of CRTAP and P3H1 is supported by immunofluorescence microscopy;reduced levels of both proteins are detected in cells with a mutation in either gene. P3H1 is partially rescued in CRTAP-null cells treated with inhibitors. Also, in LEPRE1-null cells, the secretion of CRTAP into conditioned media is increased compared with control cells and accounts for 15-20% of the decreased CRTP detected in cells. Recently, we have collaborated with an Italian team to study the CRTAP mutation found in a non-lethal proband with severe OI caused by homozygosity for a null insertion/deletion mutation. The levels of CRTAP transcripts and protein do not correlate with survival, which may be related to functions of the secreted CRTAP in matrix. Importantly, this study provided the first demonstration that absence of CRTAP results in a severe deficiency of collagen deposited into matrix (10-15% of control), with disorganization of the minimal fibrillar network. This year the BEMB identified a two children with a mutation in the 3rd component of the collagen 3-hydroxylation complex, CyPB which is incoded by PPIB. These siblings have recessivvee OI of moderate severity with white sclerae but without rhizomelia. They have a homozygous mutation in the start of codon of the peptidly prolyl isomerase gene, which results in a total absence of CyPB protein as measured on Western blots with 3 different antibodies and by immunfluoresence microscopy. Surprisingly, the 3-hydroxylation of collagen Pro986 and the hydroxylation of helical lysine and proline residues were both normal. First of all, this means that the two component of the 3-hydroxylation complex, CRTAP and P3H1, can complete collagen modification in the absence of the 3rd component. Second, normal helical modification indicates that the folding rate of the collagen helix is normal. Since CyPB had been previously thought to be the unique collagen cis-trans prolyl isomerase, normal collagen folding in the absence of CyPB means that there must be redundancy for this important function in human cells. Among our LEPRE1-deficient patients, the BEMB identified a common mutant allele, IVS5+1G to T, which occurred in both African-Americans and West Africans. This so-called """"""""West-African allele"""""""" has been found only in individuals of African descent. We determined a carrier frequency in Mid Atlantic USA of 1 in 200-300 African-Americans. In a collaboration which Charles Rotimi of NHGRI, we determined contemporary Ghanians and Nigerians had a carrier frequency for this lethal recessive mutations of 1.5%! This high carrier frequency makes the inheritance of severe OI in African distinct from the dominant form prevalent in North America and Europe, where recessive OI occurs in 5-7% of OI cases. The age of the mutation is calculated to be about 600 years old, consistent with a founder mutation that originated in West African and was introduced into North America by the Atlantic slave trade. Our studies have shown that the mutation is not found in a number of countries in Central and West Africa and hence is not pan-African SNP;the reasons for the limitation of this founder mutation to Ghana/Nigeria may reside in the use of languages with common roots spoken in this region. Finally, we have proposed that pathways common to dominant and recessive OI are likely to provide key insights into disease mechanism. These commonalities include alterations in collagen post-translational modification and folding, abnormalities in both cartilage and bone (osteochondrodystrophy), ER stress, collagen-protein binding, cell-matrix effects, increased bone turnover and hypermineralization of bone tissue.

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Webb, Emma A; Balasubramanian, Meena; Fratzl-Zelman, Nadja et al. (2017) Phenotypic Spectrum in Osteogenesis Imperfecta Due to Mutations in TMEM38B: Unraveling a Complex Cellular Defect. J Clin Endocrinol Metab 102:2019-2028
Zou, Yaqun; Donkervoort, Sandra; Salo, Antti M et al. (2017) P4HA1 mutations cause a unique congenital disorder of connective tissue involving tendon, bone, muscle and the eye. Hum Mol Genet 26:2207-2217
Kang, Heeseog; Aryal A C, Smriti; Marini, Joan C (2017) Osteogenesis imperfecta: new genes reveal novel mechanisms in bone dysplasia. Transl Res 181:27-48
Blouin, St├ęphane; Fratzl-Zelman, Nadja; Glorieux, Francis H et al. (2017) Hypermineralization and High Osteocyte Lacunar Density in Osteogenesis Imperfecta Type V Bone Indicate Exuberant Primary Bone Formation. J Bone Miner Res 32:1884-1892
Fratzl-Zelman, Nadja; Barnes, Aileen M; Weis, MaryAnn et al. (2016) Non-Lethal Type VIII Osteogenesis Imperfecta Has Elevated Bone Matrix Mineralization. J Clin Endocrinol Metab 101:3516-25
Dubnikov, Tatyana; Ben-Gedalya, Tziona; Reiner, Robert et al. (2016) PrP-containing aggresomes are cytosolic components of an endoplasmic reticulum quality control mechanism. J Cell Sci :
Terajima, Masahiko; Taga, Yuki; Chen, Yulong et al. (2016) Cyclophilin-B Modulates Collagen Cross-linking by Differentially Affecting Lysine Hydroxylation in the Helical and Telopeptidyl Domains of Tendon Type I Collagen. J Biol Chem 291:9501-12
Lindert, Uschi; Cabral, Wayne A; Ausavarat, Surasawadee et al. (2016) MBTPS2 mutations cause defective regulated intramembrane proteolysis in X-linked osteogenesis imperfecta. Nat Commun 7:11920
Forlino, Antonella; Marini, Joan C (2016) Osteogenesis imperfecta. Lancet 387:1657-71
Ben-Gedalya, Tziona; Moll, Lorna; Bejerano-Sagie, Michal et al. (2015) Alzheimer's disease-causing proline substitutions lead to presenilin 1 aggregation and malfunction. EMBO J 34:2820-39

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