Biosynthesis, Post-translation Modification, and Functional Characterization of Drm/Gremlin

Abstract

Down-regulated by mos (Drm)/Gremlin is a highly conserved protein whose properties and expression pattern suggest a role in early development, tissue-specific differentiation, and cell transformation. We have investigated the biosynthesis and processing of Drm expressed endogenously in rat fibroblasts or overexpressed following transient or stable transfection. Analysis of metabolically labeled cells revealed that Drm exists in secreted and cell-associated forms that exhibit similar mobilities in SDS-polyacrylamide gel electrophoresis. Protein analysis indicated that Drm is present in two major species: a slow migrating glycosylated form and a nonglycosylated form. Both forms of Drm are able to undergo phosphorylation. Drm is released into the media within 30 min of synthesis and is detectable for up to 4 -5 h, whereas the cell-associated form has a half-life of about 1 h. Confocal immunofluorescent microscopy indicates that Drm is present both on the external surface of expressing cells, as well as within the endoplasmic reticulum and the Golgi. Both glycosylated and nonglycosylated forms of Drm exhibit identical distributions and are able to antagonize bone morphogenetic protein signaling. Like the soluble form, the cell-associated forms are capable of binding 125

The bone morphogenic protein antagonist gremlin regulates proximal‐distal patterning of the lung

“…BMP-4 with relatively high affinity, which gives further credence to our hypothesis that gremlin inhibits BMP-4 signaling in SP-C/gremlin mice (Topol et al, 2000). Interestingly, BMP-4 has been shown to regulate proximal-distal patterning during tooth development, suggesting that this regulatory role for BMP-4 may be generalized to tissues other than lung (Tucker et al, 1998;Cheifetz, 1999).…”

“…One important difference between the studies presented here and those reported by Weaver et al is that endogenous gremlin is expressed in the proximal airway epithelium, whereas mouse noggin is normally expressed in the distal mesenchyme of the embryonic lung (Weaver et al, 1999). These data, along with previous reports that gremlin inhibits signaling by BMP-4, strongly implicate gremlin as an important regulator of proximal-distal patterning during lung development and suggests that gremlin may program proximal lung epithelial cell fate through inhibition of BMP signaling (Hsu et al, 1998;Topol et al, 2000).…”

“…Gremlin protein expression during lung development. Immunohistochemistry was performed on embryonic day (E) 12.5 and 18.5 wild-type mouse embryos using a previously characterized rabbit anti-gremlin polyclonal antibody (Topol et al, 2000). At E12.5, gremlin protein expression was observed in the epithelium of the developing esophagus (A, arrow) and the limb bud (D, bracket).…”

“…The rabbit polyclonal anti-rat gremlin and anti-human gremlin antibodies have been previously described (Topol et al, 2000). The mouse monoclonal anti-smooth muscle ␣-actin was obtained from Roche Biochemicals.…”

“…We used previously characterized rabbit polyclonal antisera to analyze gremlin protein expression by immunohistochemistry (Topol et al, 2000). Histologic sections from fixed and embedded embryonic day (E) 12.5 and E18.5 wild-type mouse embryos were stained with a rabbit anti-rat gremlin antisera, and the resulting signal was detected by the use of a secondary anti-rabbit FITC conjugated antibody.…”