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Proprotein convertases (PCs) are a family of serine endoproteases that proteolytically activate many precursor proteins within various secretory pathway compartments. Loss-of-function studies have demonstrated a critical role for these proteases in embryonic patterning and adult homeostasis, yet little is known about how substrate selectivity is achieved. We have identified Xenopus orthologs of three PCs: furin, PC6, and PC4. In addition to previously described isoforms of PC6 and furin, four novel splice isoforms of PC6, which are predicted to encode constitutively secreted proteases, and a putative transmembrane isoform of PC4 were identified. Furin and PC6 are expressed in dynamic, tissue-specific patterns throughout embryogenesis, whereas PC4 transcripts are restricted primarily to germ cells and brain in adult frogs.
Figure 1. Analysis of temporal and spatial patterns of expression of xFurin in developing Xenopus embryos. A: Schematic illustration of domain structure and amino acid identity shared by Xenopus and mouse furin. B: Sequence alignment of the cytoplasmic tail domain of Xenopus and mouse furin showing conservation of internalization signals (green shading), acidic clusters (yellow shading), and phosphorylated serines (black dots) that direct subcellular trafficking. C-F: Analysis of xFurin expression by Northern blot analysis (C) or whole-mount in situ hybridization (D-F). D: Dorsal (dor), lateral (lat), anterior (ant), and midsagittal (lower right) views of neurula stage (st. 18) embryos are shown. Sm, somitic mesoderm; lp, lateral plate mesoderm; vm, ventralmesoderm. E,F: Lateral views of tail bud stage (st. 28-32) embryos. The embryo shown in F has been cleared. pn, pronephric duct; ov, otic vesicle; m, finmesenchyme; pa, pharyngeal arches; e, eye.
Figure 4. Spatial distribution of PC6 transcripts during Xenopus embryogenesis. A-H: In situ hybridization of probes specific for all xPC6 isoforms (A-F) or for xPC6B-D (G,H) to whole Xenopus embryos. Dorsal (A) and dorsoanterior (B) views of neurula (st. 18) stage embryos or lateral views of tail bud (st. 28-30; C-E,G) and tadpole (st. 36; F,H) stage embryos are shown. D: The embryo was cleared to visualize internal structures. nf, neural fold; e, eye; pd, pronephric duct; ov, otic vesicle; pa, pharyngeal arches; nc, notochord; n, nasal placode; ll, lateral line; gs, gill slits; k, kidney; PC, proprotein convertases. Arrowheads denote staining of specific stripes of cells in the brain, open arrowheads indicate staining of mesenchymal cells posterior to the pharyngeal arches.
