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 alpha-1 type III collagen. In the present study, the protein and gene expression 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 examined. Two-dimensional PAGE of proteins synthesized and released by whole conceptuses or isolated chorioallantoic membranes into culture medium demonstrated that the C-terminal of procollagen III was not detectable before day 21 of pregnancy and concentrations increased thereafter. Developmental gene expression was determined by Northern blot analysis using a probe (A) that preceded all five polyadenylation sites of the previously sequenced clone 9.22. Procollagen III mRNA expression was 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 terminated between poly-A sites 2 and 3, was designed to identify transcripts 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 isolated from our bovine conceptus cDNA library and sequenced. One, designated 9.29, terminated at poly-A site 5. The other, designated 11.7, terminated at poly-A site 2, indicating that the bovine conceptus uses these stop sites in procollagen III transcription. Results from this study demonstrate that procollagen III gene and protein expression coincide with the development of the allantois, which progressively fuses with the chorion forming the chorioallantois placenta. In addition, multiple termination sites are used in procollagen III transcription.