Neogenin Interacts with Hemojuvelin through Its Two Membrane-Proximal Fibronectin Type III Domains
Abstract
Hemojuvelin is a recently identified iron-regulatory protein that plays an important role in affecting the expression of hepcidin, a key iron regulatory hormone. Although the underlying mechanism of this process is not clear, several hemojuvelin-binding proteins, including the cell surface receptor neogenin and bone morphogenetic protein (BMP) cytokines, have been identified. The ectodomain of neogenin is composed of four immunoglobulin-like (Ig) domains followed by six fibronectin type III-like (FNIII) domains. Here we report expression of soluble versions of hemojuvelin and neogenin for biochemical characterization of their interaction and the interaction of HJV with BMP-2. Hemojuvelin normally undergoes an autocatalytic cleavage, and as in vivo, recombinant hemojuvelin exists as a mixture of cleaved and uncleaved forms. Neogenin binds to cleaved and noncleaved hemojuvelin, as verified by its binding to an uncleaved mutant hemojuvelin. We localized the hemojuvelin binding site on neogenin to the membrane-proximal fifth and sixth FNIII domains and the juxtamembrane linker and showed that a fragment containing only this region binds 2–3 orders of magnitude more tightly than the entire neogenin ectodomain. Binding to the most membrane-proximal region of neogenin may play a role in regulating the levels of soluble and membrane-bound forms of hemojuvelin, which in turn would influence the amount of free BMP-2 available for binding to its receptors and triggering transcription of the hepcidin gene. Our finding that BMP-2 and neogenin bind simultaneously to hemojuvelin raises the possibility that neogenin is part of a multiprotein complex at the hepatocyte membrane involving BMP, its receptors, and hemojuvelin.
Additional Information
© 2008 American Chemical Society. ACS AuthorChoice. Received 8 January 2008. Date accepted 20 February 2008. Published online 13 March 2008. Published in print 1 April 2008. This work was supported by the National Institutes of Health (1R01 DK60770 to P.J.B.). We thank Caroline Enns and An-Sheng Zhang (Oregon Health and Sciences University) for cDNAs, sharing unpublished data, and critical reading of the manuscript, Inderjit Nangiana at the Caltech Protein Expression Center for assistance with protein expression, G. Hathaway and the Caltech PPMAL for N-terminal sequencing, Richard Olson for help with analytical ultracentrifugation, and Adrian Rice for assistance with light scattering experiments.Attached Files
Published - bi800036h.pdf
Accepted Version - nihms-173385.pdf
Supplemental Material - bi800036h-file003.pdf
Supplemental Material - bi800036h-file004.pdf
Files
Additional details
- PMCID
- PMC2819367
- Eprint ID
- 73907
- Resolver ID
- CaltechAUTHORS:20170201-074623072
- NIH
- 1R01 DK60770
- Created
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2017-02-01Created from EPrint's datestamp field
- Updated
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2021-11-11Created from EPrint's last_modified field