STRUCTURE-FUNCTION RELATIONSHIPS IN TYPE I COLLAGEN 
Jeffrey Bonadio, M.D., Assistant Investigator 
Dr. Bonadio' s laboratory is interested in the rela- 
tionship between structure and fijnction for type I 
collagen, a fibrous protein that resides in the extra- 
cellular matrix of most tissues. Type I collagen was 
chosen partly because of a larger interest in the 
contribution of the matrix to tissue assembly dur- 
ing development. The goal is to define this contri- 
bution in molecular terms, by focusing on a major 
component of the matrix. The heteropolymeric na- 
ture of collagen, its abundance, its supramolecular 
assembly into fibrils, its structural role, and its orga- 
nization into a connective tissue matrix distinguish 
collagen from many other proteins and imply that 
unique structure-function relationships exist. 
L Characterization of an Osteogenesis Imperfecta 
Mutation. 
The opportunity to characterize naturally occur- 
ring mutations provides a powerful set of sequence 
alterations that, given their association with human 
disease, are assumed to occur within important do- 
mains in the molecule. The laboratory recently de- 
scribed a homozygous, G-*A transition at the mod- 
erately conserved +3 position within the splice 
donor site of intron 14 in the human a 1(1) collagen 
gene. The mutation reduced the efficiency of nor- 
mal splice-site selection, since the exon upstream 
of the mutation was alternatively spliced. Moreover, 
the extent of alternative temperatures at which the 
mutant cells were grown suggested that the muta- 
tion had a direct effect on spliceosome assembly. 
The homozygous mutation was associated with a 
lethal phenotype. However, inefficient exon skip- 
ping suggests that splicing mutations in collagen 
genes may be more widespread in the general pop- 
ulation than previously recognized. In other words, 
low-level expression of alternative splicing (e.g., 
with heterozygous mutations) may be associated 
with mild dysfunction of connective tissue. 
II. Transgenic Mouse Model of the Mild Dominant 
Form of Osteogenesis Imperfecta. 
Osteogenesis imperfecta type I (OI-I) is a nonle- 
thal disorder characterized by bone fracture with- 
out deformity, blue sclerae (and other defects in 
nonmineralized connective tissue), normal or near- 
normal stature, and autosomal dominant inheri- 
tance. The incidence of OI-I is estimated to be 
1/20,000 live births; males and females are affected 
equally. Osteopenia is associated with an increased 
rate of long-bone fracture upon ambulation. Prese- 
nile hearing loss is a feature of this disease in about 
half the families. All cases to date have been associ- 
ated with mutations in the extracellular matrix mol- 
ecule type I collagen. Most affected individuals are 
thought to have heterozygous, null collagen muta- 
tions, although no naturally occurring mutation of 
this type has been characterized to date at the mo- 
lecular level. 
The Movl3 strain was generated by the labora- 
tory of Dr. Rudolf Jaenisch by exposing mouse em- 
bryos to Moloney murine leukemia virus. Genetic 
and molecular evidence indicated that a single copy 
of the provirus integrated into the first intron of the 
a 1(1) collagen gene. The proviral insert is associ- 
ated with a change in chromatin conformation and 
de novo methylation of the gene, and it prevents 
initiation of transcription. Mice homozygous for the 
null mutation die in utero due to failure of the vas- 
cular system. However, heterozygous Movl3 mice 
do not display an obvious mutant phenotype. Dr. 
Bonadio's group undertook a detailed biochemical 
and functional analysis to determine whether 
Movl3 mice could serve as a model of OI-I. 
A defect in type I collagen production was associ- 
ated with dominant, morphological and functional 
defects in mineralized and nonmineralized connec- 
tive tissue and with progressive hearing loss. There- 
fore the mutant mice represent a faithful model of 
OI-I. The model provides an opportunity to investi- 
gate the effect of a reduced amount of type I colla- 
gen on the structure and integrity of extracellular 
matrix. It also may represent a system in which 
therapeutic strategies to strengthen connective tis- 
sue can be developed. 
III. Biomineralization. 
The perinatal lethal form of osteogenesis im- 
perfecta (OI-II) is characterized by short stature, a 
soft calvarium, blue sclerae, fragile skin, a small 
thoracic cage, and floppy-appearing lower extremi- 
ties (due to external rotation and abduction of the 
femurs). Radiographic signs of bone weakness in- 
clude compression of femurs, bowing of tibiae, 
broad and beaded-appearing ribs, and poor miner- 
alization (of both membranous and endochondral 
bone). The reasons for bone deformation and poor 
Continued 
31 
