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AN INTERFERON-STIMULATED GENE (ISG) ENCODING A 15-kDa PROTEIN (ISG15) IS UP-REGULATED DURING IMPLANTATION IN THE MOUSE UTERUS.
Hansen, Thomas1, Austin, Kathy1, Carr, Amy1, Belden, E2, Pru, James1, 1 2
ABSTRACT- Murine ISG15, also called ubiquitin cross reactive protein, was the first ubiquitin homolog identified that was induced in several cell lines following treatment with interferon (IFN). The induction of this ubiquitin homolog in the uterus by pregnancy was first documented in cows and later described in humans. The experiments described herein examine the hypothesis that ISG15 is up-regulated in murine endometrium during implantation of the conceptus. Uterine tissues from pregnant and pseudopregnant (mated to sterile male) ICR (Harlan) mice were collected and examined for the presence of ISG15 mRNA and protein on the following days: 3.5 (pre-implantation), 4.5 (adhesion), 7.5 (invasion) and 9.5 (completion of the exchange organ). The ISG15 cDNA was amplified from murine L-929 cellular RNA following 12 h treatment with recombinant IFN-tau using RT-PCR and was sequenced to confirm identity. This muISG15 cDNA was random-prime labeled and used in Northern blot analysis to detect ISG15 mRNA in total uterine RNA. ISG15 mRNA was more abundant (P < 0.05) on day 7.5 of pregnancy and continued to accumulate (P < 0.05) through day 9.5 of pregnancy when compared to days 3.5 or 4.5 of pregnancy and days 3.5, 4.5, 7.5 or 9.5 of pseudopregnancy. The amount of 18S ribosomal RNA did not change in all tissues examined. Mouse uterine cross sections were immuno-stained for the presence of ISG15 protein using a polyclonal rabbit antibody against recombinant bovine ISG17. There was a significant up-regulation of staining for ISG15 protein in the decidualized stroma and surrounding glandular epithelium from day 7.5 pregnant when compared with pseudopregnant mice. Detection of ISG15, in the mouse uterus during early pregnancy provides further evidence that this interferon-induced protein might be a universal uterine response to the developing and implanting conceptus. The regulator of murine ISG15 is not known but probably is an activator of the Jak/STAT signal transduction pathway. Interferon might assume this role in the murine uterus, but it is clearly recognized that other cytokines or growth factors activating the Jak/STAT pathway could also be involved. NIH HD32475-07.
KEY WORDS: mouse, uterus, interferon, ubiquitin
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