Molecular Biology of the Extracellular Matrix 
Jeffrey F. Bonadio, M.D. — Assistant Investigator 
Dr. Bonadio is also Assistant Professor of Pathology at the University of Michigan Medical School. He re- 
ceived his bachelor 's degree in biology from Marquette University and his M.D. degree from the Medical 
College of Wisconsin, Milwaukee. He studied anatomical pathology with Bruce Beckwith and medical 
molecular genetics with Peter Byers at the University of Washington. 
THE long-term goal of our research is to under- 
stand how proteins of the extracellular ma- 
trix contribute to tissue structure and function. 
Quantitative and qualitative changes in these 
proteins occur during morphogenesis and as part 
of the wound healing process. These observations 
suggest that both the organization and protein 
composition of the matrix are precisely regu- 
lated. It is clear that this regulation occurs in part 
at the level of gene expression and in part at the 
level of the assembly of proteins into a matrix- 
like configuration. 
I have chosen to focus for the most part on the 
matrix molecule type I collagen. This is a poly- 
mer of two related proteins whose sequence has 
been determined. Moreover, the multidomain 
structure of the molecule and a general outline of 
collagen biosynthesis are known, and the mole- 
cule is recognized to be distributed widely 
within tissues such as bone, tendon, ligament, 
tooth, dermis, and sclera. Previous studies have 
implied that type I collagen makes an important 
contribution to the structure, integrity, and nor- 
mal homeostasis of these tissues. Over the last 
year we have continued our work to establish 
model systems that would allow us to study this 
contribution at the molecular level. 
One system is designed to investigate the intra- 
cellular assembly of the collagen molecule. In 
general, this work involves site-specific mutagen- 
esis and assays that quantify the effects of muta- 
tion on the assembly process. These effects are 
studied at two levels. First, we have established 
conditions that allow synthetic peptides to fold 
into a collagen-like triple helix. Peptide folding 
is slow enough that the process can be character- 
ized by methods such as circular dichroism, and 
the triple helix formed in vitro is stable enough 
that its structure can be characterized by nuclear 
magnetic resonance (NMR) . Therefore the effect 
of a given mutation can be quantified by directly 
comparing the behavior of a normal peptide with 
that of a mutant one. Also, cellular transfection 
methods have been developed to express and as- 
semble collagen molecules in vitro; and again, 
the effect of mutation on the assembly process 
can be quantified by directly comparing the be- 
havior of normal and mutant molecules. 
In our initial mutagenesis experiments, we 
characterized a highly conserved region of the 
triple helical domain and demonstrated that it 
made an important contribution to the assembly 
of collagen molecules into a thermodynamically 
stable conformation. We speculate that this re- 
gion was conserved during evolution because it 
plays an important role in collagen biosynthesis 
— i.e., in folding the collagen molecule into its 
correct conformation. In the future, we hope to 
use this model system to define further the nor- 
mal contribution made by other collagen do- 
mains to the assembly process. In addition, we 
are interested in characterizing those regions of 
the molecule that mediate interactions between 
collagen and other matrix molecules such as fi- 
bronectin, heparin sulfate proteoglycan, and in- 
tegrins. These interactions are important because 
they represent a molecular basis for the assembly 
of collagen within the matrix. 
A second system is designed to investigate the 
function of type I collagen at the level of connec- 
tive tissue. Our initial set of experiments utilized 
a transgenic mouse strain that expressed only half 
the normal amount of type I collagen. We demon- 
strated that the mutation adversely affected the 
connective tissue of bone and skin dermis. In ad- 
dition, the mutant mice were profoundly deaf. 
We utilized biomechanical tests to quantify the 
effect of the collagen deficiency at the tissue 
level, and these studies demonstrated that the 
major role of type I collagen is to provide con- 
nective tissue with a high degree of resiliency. 
More recently, we also demonstrated that the 
skeleton of these transgenic mice is able to adapt 
to the inherited collagen deficiency. This adapta- 
tion involves a thickening of cortical bone and 
results from the synthesis of new bone matrix. 
This suggests a signal transduction pathway for 
bone in which the mechanical environment 
(e.g., strain) influences the pattern of osteoblast 
gene expression. Particularly intriguing was our 
observation that the adaptation was associated 
with a significant improvement in bone strength. 
Experiments utilizing other transgenic mouse 
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