Abstract
We have cloned and characterized several overlapping cDNAs that specify a large portion of a Paracentrotus lividus fibrillar collagen molecule. Our conclusions are based on sequencing data, which showed that the clones code for a 786-amino acid collagenous domain composed of an uninterrupted series of Gly-X-Y repeats and for a 265-amino acid carboxyl-terminal globular extension. The latter domain exhibits features highly reminiscent of those of the vertebrate counterparts, notably a putative carboxyl-peptidase cleavage site, a series of similarly arranged cysteinyl residues, and an N-linked glycosylation attachment site. In situ and Northern blot hybridizations have established the size, time of appearance, and tissue localization of the collagen mRNA during sea urchin development. The collagen transcript, 9 kilobases in length, is first detected in the primary and, more predominantly, in the secondary mesenchyme cells of late gastrulae where it progressively accumulates thereafter. This and other work (D'Alessio, M., Ramirez, F., Suzuki, H.R., Solursh, M., and Gambino, R. (1989) Proc. Natl. Acad. Sci. U. S. A. 86, 9303-9307) provide evidence of a genetic heterogeneity of fibrillar collagens in the sea urchin embryo and suggest that the two genes are activated in the same cell lineages at distinct developmental stages.
Highlights
We present similar studies on the P. lividus 2a collagen
Features of the Sea Urchin 2a Collagen Chain-In a previous paper, along with cloning experiments, we have described the features of the P. lividus la! collagen protein and the pattern of developmental expression of the gene [14]
To illustrate the specificity of the hybridizing probes and to compare the relative time of appearance of the two collagen mRNAs, the noncoding regions of both la and 2a cDNAs were used in the Northern analysis
Summary
Attempts have been made to clone sea urchin collagen genes to elucidate the primary structure of the proteins and the developmental pattern of expression of the genes (10,ll) These studies, have relied solely on the isolation by crosshybridization of genomic fragments containing putative collagen-coding exons. A genomic fragment, previously shown to contain five putative collagen-coding exons and to hybridize to a 6-kb developmentally regulated transcript (ll), was used to screen a prism-stage embryonic cDNA library [14] This led to the isolation of several collagen-coding cDNAs, none of which, was found to correspond to the sequences of the original genomic clone.
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