We are pursuing several lines of research: the characterization of mutations in type I collagen genes that give rise to forms of osteogenesis imperfecta and other disorders, the identification and characterization of mutations in type III collagen genes which give rise to Ehlers-Danlos syndrome type IV, identification of proteins in the intracellular and extracellular processing pathways that identify abnormal collagen proteins, the mechanisms of mRNA processing in collagen genes,the dispersion of repetitive elements within the COL3Al gene of type III collagen, and mutations in the type V collagen genes which give rise to milder forms of EDS.
The majority of mutations in the COL1Al and COLlA2 genes that cause OI result in substitution for glycines within the triple helix. Most of the remainder alter splice sites. Our studies of the mutations suggest that in some instances the order of exon splicing may determine the effects of splice mutations; as a consequence we are studying the order of intron removal in such cell strains. One of the most puzzling aspects of OI has been the failure to identify mutations in all affected individuals. Using long amplification regions, we have noted low level splice defects in some such patients that result in the production of only a small amount of abnormal molecules due to the presence of 5-10% abnormal mRNA species as a consequence of mutations outside the canonical splice site sequences.
We have now characterized almost mutations in our families with EDS type IV. These are more heavily weighted to point mutations that result in substitutions for glycine residues within the triple helix of the molecule than mutations that alter splice site integrity. Some of these mutations prohibit mRNA transport from the nucleus when introns that contain termination codons are included. These findings suggest that there is a link between splicing and nuclear recognition of premature termination codons that may be different from the recognition process that leads to cytoplasmicnonsense-codon mediated mRNA decay. The mechanisms of recognition of these structures is being pursued.
Similar approaches are being taken to disorders which result from several other genes involved in connective tissue biogenesis.
Pepin M, Schwarze U, Superti-Furga A, Byers PH. Classical Ehlers-Danlos syndrome type IV: complications and early death due to mutations in the COL3A1 gene of type III collagen. N Engl J Med 342:673-80, 2000
Byers PH. Disorders of collagen biosynthesis and structure. In: The Metabolic and Molecular Basis of Inherited Disease, 8th edition, CR. Scriver, AL. Beaudet, WS. Sly, D Valle, B Childs, B Vogelstein, editors. McGraw-Hill, New York, pp. 5241-5286, 2000.
Schwarze U, Atkinson M, Hoffman GG, Greenspan DS, Byers PH. Null alleles of the COL5A1 gene of type V collagen are a cause of the classical forms of Ehlers-Danlos syndrome (types I and II). Am J Hum Genet 66:1757-1766, 2000
Pace JM, Kuslich CD, Willing MC, Byers PH. Disruption of one intra-chain disulfide bond in the carboxyl-terminal propeptide of the proa1(I) chain of type I procollagen permits slow assembly and secretion of overmodified, but stable procollagen trimers and results in mild osteogenesis imperfecta. J Med Genet 38:443-9, 2001
Pace JM, Atkinson M, Wallis G, Willing M, Byers PH. Small in-frame deletions and duplications in the triple helical domains encoded by the COL1A1 and COL1A2 genes of type I procollagen result in osteogenesis imperfecta. Human Mutation 18:319-326, 2001
Schwarze U, Schievink WI, Petty E, Jaff MR, Babovic-Vuksanovic D, Pepin M, Byers PH. Haploinsufficiency for type III procollagen results in the vascular type of the Ehlers-Danlos syndrome (EDS type IV). Am J Hum Genet 69:989-1001, 2001
Marlowe A, Pepin MG, Byers PH. Testing for osteogenesis imperfecta in cases of suspected non-accidental injury. J Med Genet 39:382-386, 2002
Takahara K, Schwarze U, Imamura Y, Hoffman GG, Toriello H, Smith LT, Byers PH, Greenspan DS. Order of intron removal influences multiple splice outcomes, including a 2-exon Skip, in a COL5A1 acceptor site mutation that results in abnormal pro-a1(V) N-propeptides and Ehlers-Danlos syndrome type I. Am J Hum Genet 71:451-465, 2002
Pace JM, Corrado M, Missero C, Byers PH. Identification, characterization and expression analysis of a new fibrillar collagen gene, COL27A1. Matrix Biology 22:3-14, 2003
Chamberlain, JR, Schwarze U, Wang P-R, Hirata RK, Hankenson KD, Pace JM, Underwood RA, Song KM, Sussman M, Byers PH, Russell DW. Gene targeting in stem cells from individuals with osteogenesis imperfecta. Science 303:1198-1201, 2004
Schwarze U, Hata RI, McKusick VA, Shinkai H, Hoyme HE, Pyeritz RE, Byers PH. Rare autosomal recessive cardiac valvular form of Ehlers-Danlos syndrome results from mutations in the COL1A2 gene that activate the nonsense-mediated RNA decay pathway. Am J Hum Genet 2004 Apr 9 [Epub ahead of print]