DEVELOPMENTAL GENE-EXPRESSION OF PROCOLLAGEN-III IN BOVINE EXTRAEMBRYONIC MEMBRANES DURING EARLY-PREGNANCY

A major secretory protein produced by bovine chorioallantoic membranes , in vitro, was previously identified as the carboxyl-propeptide of al pha-1 type III collagen. In the present study, the protein and gene ex pression of procollagen III by bovine chorioallantois between days 17 and 45 of pregnancy was investigated. In addition, differential usage of multiple transcription termination sites by chorioallantois was exa mined. Two-dimensional PAGE of proteins synthesized and released by wh ole conceptuses or isolated chorioallantoic membranes into culture med ium demonstrated that the C-terminal of procollagen III was not detect able before day 21 of pregnancy and concentrations increased thereafte r. Developmental gene expression was determined by Northern blot analy sis using a probe (A) that preceded all five polyadenylation sites of the previously sequenced clone 9.22. Procollagen III mRNA expression w as undetectable at day 17, low on day 20, and increased through day 36 . Two major transcripts of 5.9 and 4.9 kb were identified, the latter of which was expressed more prominently. A second probe (B), which ter minated between poly-A sites 2 and 3, was designed to identify transcr ipts that terminated at poly-A site 1 or 2. This probe bound to the 5. 9-kb mRNA only. Two additional procollagen III cDNA clones were isolat ed from our bovine conceptus cDNA library and sequenced. One, designat ed 9.29, terminated at poly-A site 5. The other, designated 11.7, term inated at poly-A site 2, indicating that the bovine conceptus uses the se stop sites in procollagen III transcription. Results from this stud y demonstrate that procollagen III gene and protein expression coincid e with the development of the allantois, which progressively fuses wit h the chorion forming the chorioallantois placenta. In addition, multi ple termination sites are used in procollagen III transcription. (C) 1 997 Wiley-Liss, Inc.