POGLUT2/3 mediated EGF O-glucosylation promotes separation of digits 2 and 3 by influencing fibrillin network reorganization, signaling, and cell dynamics

The separation of individual digits is dependent on establishment of digit-interdigit periodicity, remodeling of the interdigital mesenchyme, and invagination of interdigital epithelial tongues. In Protein O-glucosyltransferase 2 and 3 double knockout (Poglut2/3 DKO) mice, digits 2 and 3 are fused, suggesting a defect in one or more processes. POGLUT2/3 add O-linked glucose to epidermal growth factor-like (EGF) repeats. Syndactyly is also observed when genes encoding the POGLUT2/3 substrates fibrillin 2 (FBN2) or both Nidogen 1 and 2 (NID1/2) are knocked out, suggesting that O-glucosylation is important for their function or localization. In this study, we evaluated the distribution of these substrates during digit separation and the effects of the Poglut2/3 DKO on their localization and cell behavior. During digit separation, the FBNs underwent a dramatic reorganization. Aberrant levels and distribution of the FBNs were observed in the Poglut2/3 DKO and microfibrils isolated from Poglut2/3 DKO skin showed altered periodicity in Fibrillin microfibrils. In contrast, the Poglut2/3 DKO had no effect on the levels or localization of NID1. In Poglut2/3 DKOs, bone morphogenetic protein (BMP) signaling was reduced during digit development, especially in the anterior autopod. Early anterior reduction of BMP signaling could potentially affect spacing of digits 2 & 3. While later reduction of BMP signaling in the Poglut2/3 DKO in the digit 2-3 region was likely responsible for defects in clearance of interdigital mesenchyme and interdigital tongue morphogenesis. These results highlight the importance of POGLUT2/3 mediated O-glucosylation for FBN microfibril organization and raise the possibility that O-glucose modulates the biological or physical properties of the FBN microfibril network.