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Article
Identification of Putative Receptors for the Novel Adipokine CTRP3 Using Ligand-Receptor Capture Technology
PLoS ONE
  • Ying Li, East Tennessee State University
  • Tammy Ozment, East Tennessee State University
  • Gary L. Wright, East Tennessee State University
  • Jonathan M. Peterson, East Tennessee State University
Document Type
Article
Publication Date
10-11-2016
Description

C1q TNF Related Protein 3 (CTRP3) is a member of a family of secreted proteins that exert a multitude of biological effects. Our initial work identified CTRP3’s promise as an effective treatment for Nonalcoholic fatty liver disease (NAFLD). Specifically, we demonstrated that mice fed a high fat diet failed to develop NAFLD when treated with CTRP3. The purpose of this current project is to identify putative receptors which mediate the hepatic actions of CTRP3.

Methods

We used Ligand-receptor glycocapture technology with TriCEPS™-based ligand-receptor capture (LRC-TriCEPS; Dualsystems Biotech AG). The LRC-TriCEPS experiment with CTRP3-FLAG protein as ligand and insulin as a control ligand was performed on the H4IIE rat hepatoma cell line.

Results

Initial analysis demonstrated efficient coupling of TriCEPS to CTRP3. Further, flow cytometry analysis (FACS) demonstrated successful oxidation and crosslinking of CTRP3-TriCEPS and Insulin-TriCEPS complexes to cell surface glycans. Demonstrating the utility of TriCEPS under these conditions, the insulin receptor was identified in the control dataset. In the CTRP3 treated cells a total enrichment of 261 peptides was observed. From these experiments 5 putative receptors for CTRP3 were identified with two reaching statistically significance: Lysosomal-associated membrane protein 1 (LAMP-1) and Lysosome membrane protein 2 (LIMP II). Follow-up Co-immunoprecipitation analysis confirmed the association between LAMP1 and CTRP3 and further testing using a polyclonal antibody to block potential binding sites of LAMP1 prevented CTRP3 binding to the cells

Conclusion

The LRC-TriCEPS methodology was successful in identifying potential novel receptors for CTRP3.

Relevance

The identification of the receptors for CTRP3 are important prerequisites for the development of small molecule drug candidates, of which none currently exist, for the treatment NAFLD.

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Copyright Statement

© 2016 Li et al. This document was originally published in PLOS ONE.

Creative Commons License
Creative Commons Attribution 4.0
Citation Information
Ying Li, Tammy Ozment, Gary L. Wright and Jonathan M. Peterson. "Identification of Putative Receptors for the Novel Adipokine CTRP3 Using Ligand-Receptor Capture Technology" PLoS ONE Vol. 11 Iss. 10 (2016) p. e0164593 ISSN: 1932-6203
Available at: http://works.bepress.com/jonathan-peterson/25/