Date of Award


Degree Type


Degree Name

Doctor of Philosophy


cDNA libraries were constructed using the Okayama/Berg cloning strategy, and full length cDNA clones for the growth factor inducible genes TIMP, MRP, MEP and calcyclin were isolated, and used to study the temporal and spatial patterns of expression of these genes. The TIMP and MRP cDNAs were used to express the murine proteins in E. coli. The native proteins were found to be expressed only at low levels, although fusion proteins with MRP and TIMP sequences at the carboxyl terminus of {dollar}\beta{dollar}-galactosidase were expressed at high levels in E. coli. The purified fusion proteins were used to raise MRP and TIMP specific antisera.;To determine the effect of an upstream overlapping ORF, TIMP transcripts either containing or lacking the AUG of the 5{dollar}\sp\prime{dollar} ORF were generated by SP6 polymerase, and translated in reticulocyte lysates. The in vitro translated TIMP was immunoprecipitable with TIMP antiserum. TIMP was translated up to 10-fold more efficiently from the transcript lacking the upstream AUG.;RNA was isolated from the reproductive tissues dissected from female mice on each day of pregnancy and analysed on Northern and slot blots. TIMP mRNA levels were highest on day 7, and then decreased with a specific time course in each of placenta, decidua and uterus during pregnancy. A coinduction of TIMP and Transin-2 mRNA levels was seen in the ovary on day 18 of gestation.;I found that spp, calcyclin, MEP, MRP and transin exhibit unique spatial and temporal patterns of expression in the female reproductive tissues. Calcyclin mRNA increased 5-10 fold over the level seen in virgin uterus, peaking on day 10. spp mRNA was undetectable in virgin uterus and increased to a maximal level, 2{dollar}1\over2{dollar} times higher than that of calcyclin, on day 11. In situ hybridization revealed that spp was expressed in the decidua capsularis as well as in granulated metrial gland cells. Calcyclin expression in adjacent sections was a 'diffuse band' with the highest intensity in the decidua capsularis and decidua lateralis.



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