UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) binds to alpha-actinin 1: novel pathways in skeletal muscle?

Abstract

BACKGROUND:

Hereditary inclusion body myopathy (HIBM) is a rare neuromuscular disorder caused by mutations in GNE, the key enzyme in the biosynthetic pathway of sialic acid. While the mechanism leading from GNE mutations to the HIBM phenotype is not yet understood, we searched for proteins potentially interacting with GNE, which could give some insights about novel putative biological functions of GNE in muscle.

METHODOLOGY/PRINCIPAL FINDINGS:

We used a Surface Plasmon Resonance (SPR)-Biosensor based assay to search for potential GNE interactors in anion exchanged fractions of human skeletal muscle primary culture cell lysate. Analysis of the positive fractions by in vitro binding assay revealed alpha-actinin 1 as a potential interactor of GNE. The direct interaction of the two proteins was assessed in vitro by SPR-Biosensor based kinetics analysis and in a cellular environment by a co-immunoprecipitation assay in GNE overexpressing 293T cells. Furthermore, immunohistochemistry on stretched mouse muscle suggest that both GNE and alpha-actinin 1 localize to an overlapping but not identical region of the myofibrillar apparatus centered on the Z line.

CONCLUSIONS/SIGNIFICANCE:

The interaction of GNE with alpha-actinin 1 might point to its involvement in alpha-actinin mediated processes. In addition these studies illustrate for the first time the expression of the non-muscle form of alpha-actinin, alpha-actinin 1, in mature skeletal muscle tissue, opening novel avenues for its specific function in the sarcomere. Although no significant difference could be detected in the binding kinetics of alpha-actinin 1 with either wild type or mutant GNE in our SPR biosensor based analysis, further investigation is needed to determine whether and how the interaction of GNE with alpha-actinin 1 in skeletal muscle is relevant to the putative muscle-specific function of alpha-actinin 1, and to the muscle-restricted pathology of HIBM.

• Authors: Shira Amsili, Hagit Zer, Stephan Hinderlich, Sabine Krause, Michal Becker-Cohen, Daniel G MacArthur, Kathryn N North, Stella Mitrani-Rosenbaum
• Journal: PloS one (PLoS One) Vol. 3 Issue 6 Pg. e2477 (Jun 18 2008) ISSN: 1932-6203 [Electronic] United States
• PMID: 18560563 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• ACTN1 protein, human
• Multienzyme Complexes
• UDP-N-acetylglucosamine 2-epimerase - N-acetylmannosamine kinase
• Actinin

Topics

• Actinin (metabolism)
• Animals
• Biosensing Techniques
• Blotting, Western
• Cell Line
• Humans
• Immunohistochemistry
• Immunoprecipitation
• Mass Spectrometry
• Mice
• Multienzyme Complexes (metabolism)
• Muscle, Skeletal (enzymology, metabolism)
• Protein Binding
• Surface Plasmon Resonance