Expression of the (V+C)- fibronectin isoform is tightly linked to the presence of a cartilaginous matrix.
Fibronectin is encoded by a single gene, but heterogeneity is introduced by alternative splicing of the pre-mRNA. An unique splice variant, designated (V+C)-, which deletes nucleotides encoding the V, III-15 and I-10 segments, has been identified in articular cartilage. In this study, a ribonuclease protection assay was used to quantitate expression of the (V+C)- isoform in eight canine cartilaginous tissues and in chondrocytes cultured as monolayers or in alginate beads. The (V+C)- fibronectin isoform was detected in all cartilaginous tissues examined, ranging from a low of 11% of steady-state fibronectin mRNA in the nucleus pulposus to 71% in the rib. An age dependent increase, from 18% in the epiphyseal cartilage of a newborn to 54% in the articular cartilage of dogs over 10 months of age, was observed. The ubiquitous presence of this isoform in cartilaginous tissues and its absence in all non-cartilaginous tissues examined to date is consistent with a very strong association of the (V+C)- fibronectin isoform with the cartilaginous phenotype. Results from a ribonuclease protection assay using a probe extending into the V region from III-14 were combined with the quantitative information about (V+C)- fibronection expression to develop an over-all profile of splicing within the V region in cartilage. Monolayer culture of articular chondrocytes altered fibronectin splicing patterns. The (V+C)- isoform was rapidly lost and ED-A(+) fibronectin was induced. Three-dimensional culture in alginate beads prevented induction of ED-A(+) fibronection, but failed to sustain expression of the (V+C)- isoform. Thus, some matrix component or structure, lost in cell culture, may be essential to maintain expression of the (V+C)- isoform. The possible relationship of changing patterns of fibronectin isoforms in cultured chondrocytes to maintenance of the differentiated phenotype is discussed.
James A. Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, Ithaca, New York 14853, USA.
N
N
Burton-Wurster
C
C
Borden
G
G
Lust
J N
JN
Macleod
D017398:Alternative Splicing; D000818:Animals; D002356:Cartilage / Q000166:cytology / Q000378:metabolism*; D002454:Cell Differentiation; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism; D004285:Dogs; D005109:Extracellular Matrix / Q000378:metabolism*; D005353:Fibronectins / Q000096:biosynthesis* / Q000235:genetics; D020033:Protein Isoforms / Q000096:biosynthesis* / Q000235:genetics
D005353:Fibronectins; D020033:Protein Isoforms