McKnite, A. | bioRxiv Science

bioRxiv2019

BMP7 functions predominantly as a heterodimer with BMP2 or BMP4 during mammalian embryogenesis

BMP7/BMP2 or BMP7/BMP4 heterodimers are more active than homodimers in vitro, but it is not known whether these heterodimers signal in vivo. To test this, we generated knock in mice carrying a mutation (Bmp7R-GFlag) that prevents proteolytic activation of the dimerized BMP7 precursor protein. This mutation eliminates the function of BMP7 homodimers and all other BMPs that normally heterodimerize with BMP7. While Bmp7 null homozygotes are live born, Bmp7R-GFlag homozygotes are embryonic lethal and have broadly reduced BMP activity. Furthermore, compound heterozygotes carrying the Bmp7R-G allele together with a null allele of Bmp2 or Bmp4 die during embryogenesis with defects in ventral body wall closure and/or the heart. Co-immunoprecipitation assays confirm that endogenous BMP4/7 heterodimers exist. Thus, BMP7 functions predominantly as a heterodimer with BMP2 or BMP4 during mammalian development, which may explain why mutations in either Bmp4 or Bmp7 lead to a similar spectrum of congenital defects in humans.

developmental biology↗

bioRxiv2017

Fibronectin type III and intracellular domains of Toll-like receptor 4 interactor with leucine-rich repeats (Tril) are required for developmental signaling

Toll-like receptor 4 interactor with leucine-rich repeats (Tril) is a transmembrane protein that functions as a coreceptor for Toll-like receptors (Tlrs) to mediate innate immune responses in the adult brain. Tril also triggers degradation of the Bmp inhibitor, Smad7, during early embryonic development to allow for normal blood formation. Tril most likely plays additional, yet to be discovered, roles during embryogenesis. In the current studies, we performed a structure-function analysis, which indicated that the extracellular domain, including the fibronectin type III (FN) domain, and the intracellular domain of Tril are required to trigger Smad7 degradation in the early Xenopus embryo. Furthermore, we found that a Tril deletion mutant lacking the FN domain (Tril{Delta}FN) can dominantly inhibit signaling by endogenous Tril when overexpressed in vivo. This finding raises the intriguing possibility that the FN domain functions to bind endogenous Tril/Tlr4 ligands, perhaps including extracellular matrix molecules. We also show that Tril normally cycles between the cell surface and endosomes, and that the Tril extracellular domain is required to retain Tril at the cell surface, while the intracellular domain is required for Tril internalization in Xenopus ectodermal explants. Using a CHO cell aggregation assay, we further show that, unlike other transmembrane proteins that contain leucine rich repeats in the extracellular domain, Tril is not sufficient to mediate homophilic adhesion. Our findings identify Tril{Delta}FN as a valuable tool that can be used to block the function of endogenous Tril in vivo in order to discover additional roles during embryonic development.

cell biology↗