Agrin

Agrin is a large proteoglycan whose best characterised role is in the development of the neuromuscular junction during embryogenesis. Agrin is named based on its involvement in the aggregation of acetylcholine receptors during synaptogenesis.

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Agrin's mechanism of action

During development, the growing end of motor neuron axons secrete a protein called agrin. This protein binds to several receptors on the surface of skeletal muscle. The receptor which seems to be required for formation of the neuromuscular junction (NMJ) is called the MuSK receptor (Muscle specific kinase). MuSK is a receptor tyrosine kinase - meaning that it induces cellular signaling by causing the addition of phosphate molecules to particular tyrosines on itself, and on proteins which bind the cytoplasmic domain of the receptor.

In addition to MuSK, agrin binds several other proteins on the surface of muscle, including dystroglycan and laminin. Apparently these additional binding steps are required to stabilize the NMJ.

The requirement for Agrin & MuSK in the formation of the NMJ was primarily demonstrated by "knockout" mouse studies. In mice which are deficient for either protein, the neuromuscular junction does not form. Many other proteins also comprise the NMJ, and are required to maintain its integrity. For example, MuSK also binds a protein called "dishevelled" (Dvl), which is in the Wnt signalling pathway. Dvl is additionally required for MuSK-mediated clustering of AChRs, since inhibition of Dvl blocks clustering.

Agrin signaling

The nerve secretes agrin, resulting in phosphorylation of the MuSK receptor.

The MuSK receptor apparently recruits casein kinase 2, which is required for clustering.

A protein called rapsyn is then recruited to the primary MuSK scaffold, to induce the additional clustering of acetylcholine receptors (AChR). This is thought of as the secondary scaffold. A protein called Dok-7 has shown to be additionally required for the formation of the secondary scaffold; it is apparently recruited after MuSK phosphorylation, and before AChR are clustered.

Agrin, a proteoglycan

There are three potential heparan sulfate (HS) attachment sites within the primary structure of agrin but it is thought that only two of these actually carry HS chains when the protein is expressed.

Heparan sulphate glycosaminoglycans covalently linked to the agrin protein have been shown to play a role in the clustering of AChR. Interference in the correct formation of heparan sulfate through the addition of chlorate to skeletal muscle cell culture, results in a decrease in the frequency of spontaneous acetylcholine receptor (AChR) clustering. It may be that rather than solely binding directly to the agrin protein core a number of components of the secondary scaffold may also interact with its heparan sulfate side chains.

A role in the retention of anionic macromolecules within the vasculature has also been suggested for agrin linked HS at the glomerular or alveolar basement membrane.

References

[1] Induction, assembly, maturation and maintenance of a postsynaptic apparatus

[2] Agrin receptors at the skeletal neuromuscular junction

[3] Defective neuromuscular synaptogenesis in agrin-deficient mutant mice

[4] Agrin acts via a MuSK receptor complex. Cell. 1996 May 17;85(4):513-23.

[5] Casein kinase 2-dependent serine phosphorylation of MuSK regulates acetylcholine receptor aggregation at the neuromuscular junction. Genes Dev. 2006 Jul 1;20(13):1800-16

[6]Reduced glycosaminoglycan sulfation diminishes the agrin signal transduction pathway. Dev. Neurosci. 2004 26(1):1-10

[7] Primary structure and high expression of human agrin in basement membranes of adult lung and kidney. Eur. J. Biochem. 1998 254:123-128