The epidermal growth factor (EGF) family consists of EGF, HB-EGF, TGFα, amphiregulin, epiregulin, betacellulin, and the neuregulins. The neuregulins include neu differentiation factor (NDF), glial growth factors (GGFs), acetylcholine receptor inducing activity (ARIA), sensory and motor neuron-derived factor (SMDF), and the heregulins (human homologues), and are splice products of a single gene (NRG1). Neuregulin-2 (NRG2) and neuregulin-3 (NRG3) are growth factors which contain an EGF-like domain and activate erbB RTKs but are encoded by loci distinct from NRG1.

EGF-like ligands and their erbB receptors have unique spatial and temporal expression patterns in the mouse uterus during the peri-implantation period, suggesting an important role in the implantation process. We previously described a highly restricted expression pattern of NDF around the implanting embryo. Here, we examined the expression of NRG2 and NRG3 in the peri-implantation uterus. In addition, 3 subtypes of NRG1 isoforms exist, based on their sequence and predicted protein structure. Therefore, we also examined SMDF expression as a means to survey isoform-specific expression in the mouse uterus.

RT-PCR was performed on uterine tissues from CD-1 mice on day 5 of pregnancy (d1=vaginal plug). NRG2-specific primers were derived from rat and mouse cDNA sequence and included Ig, spacer, and EGF domains. The anticipated PCR products were detected only in brain (control) tissue, with results confirmed by Southern hybridization and nested RT-PCR. cDNAs representing both alpha and beta NRG2 subtypes were recovered and used as probes for Northern and in-situ hybridization. NRG3-specific primers also failed to detect a NRG3 transcript in peri-implantation uterine samples by similar methods. However, nested RT-PCR specific for mouse SMDF amplified a cDNA of 436 bp from both uterus and brain tissues, suggesting that type III NRG1 isoforms are expressed during this period. Taken together, we propose that selective members of the neuregulin family are utilized during implantation for cell-cell signaling between the embryo and uterus.