Uterine secretion of ISP1 & 2 tryptases is regulated by progesterone and estrogen during pregnancy and the endometrial cycle.

We have described two novel implantation serine proteinase (ISP) genes that are expressed during the implantation period. The ISP1 gene may encode the embryo-derived enzyme strypsin, which is necessary for blastocyst hatching in vitro and the initiation of invasion. The ISP2 gene, which encodes a related tryptase, is expressed in endometrial glands and is regulated by progesterone during the peri-implantation period. Based on similarities between ISP2 gene expression and that of a progesterone-regulated lumenal serine proteinase activity associated with lysis of the zona pellucida, we have suggested that the strypsin related protein, ISP2, may encode a zona lysin proteinase. Recently strypsin has also been found within uterine fluid, suggesting a second potential role in hatching. Consistently, we have discovered that ISP1 is also expressed in the uterine secretory gland at the time of hatching. In this study we demonstrate that both ISP1 and ISP2 are secreted together into the uterine lumen at peri-implantation, and that the appearance of ISP protein is regulated positively at the transcriptional level by progesterone and negatively at the posttranscriptional level by estrogen. This negative regulation by estrogen may be overridden in pregnancy as ISP protein expression is restored during oil-induced decidualization. ISP1 and ISP2 proteins are also expressed in proestrous suggesting additional roles in the endometrial cycle.

Origin of the murine implantation serine proteinase subfamily.

The S1 serine protease family is one of the largest gene families known. Within this family there are several subfamilies that have been grouped together as a result of sequence comparisons and substrate identification. The grouping of related genes allows for the speculation of function for newly found members by comparison and for novel subfamilies by contrast. Analysis of the evolutionary patterns of genes indicates whether or not orthologs are likely to be identified in other species as well as potentially indicating that hypothesized orthologs are in fact not. Looking at subtle differences between subfamily members can reveal intricacies about function and expression. Previously, we have described genes encoding two novel serine proteinases, ISP1 and ISP2, which are most closely related to tryptases. The ISP1 gene encodes the embryo-derived enzyme strypsin, which is necessary for blastocyst hatching and invasion in vitro. Additionally both ISP1 and ISP2 are co-expressed in the endometrial gland during the time of hatching, suggesting that they may also both participate in zona lysis from within the uterine lumen. Here, we demonstrate that the ISPs are tandemly linked within the tryptase cluster on 17A3.3. We suggest that remarkable similarities within the 5'-untranslated and first intron regions of ISP1 and ISP2 may explain their intimate co-regulation in uterus. We also suggest that ISP genes have evolved through gene duplication and that the ISP1 gene has also begun to adopt an additional new function in the murine preimplantation embryo.

Embryonic hatching enzyme strypsin/ISP1 is expressed with ISP2 in endometrial glands during implantation.

Embryo hatching and outgrowth are the first critical steps on the way to a successful pregnancy. It is generally held that serine proteases are responsible for this process, although the exact mechanisms of action are not clearly understood. Recently, we described two novel implantation serine proteinase (ISP) genes that are expressed during the implantation period. The ISP1 gene encodes the embryo-derived enzyme strypsin, which is necessary for blastocyst hatching in vitro and the initiation of invasion. The ISP2 gene, which encodes a related tryptase, is expressed in endometrial glands and is regulated by progesterone during the peri-implantation period. Based on similarities between ISP2 gene expression and that of a progesterone-regulated lumenal serine proteinase activity associated with lysis of the zona pellucida, we have suggested that the strypsin related protein, ISP2, may encode a zona lysin proteinase. As tryptases naturally assemble to form tetrameric structures, we have hypothesized that ISP1 and ISP2 tetramerize to form strypsin and lysin, respectively. In this study, we demonstrate that like ISP2, the ISP1 gene is also expressed in endometrial glands and is positively regulated by progesterone during implantation. Using in situ hybridization of adjacent tissue sections, we show that the ISP1 and ISP2 genes are co-expressed within the endometrial gland. Following evidence that ISP1 and 2 can efficiently form homotetramers and heterotetramers in silico, we suggest that ISP heterotetramers may be also be secreted into the uterine lumen during the implantation period. That the embryonic hatching enzyme, may also be secreted into the uterine lumen from uterus, may provide insight into the mechanisms of hatching and implantation initiation.