AGRIN CAN MEDIATE ACETYLCHOLINE-RECEPTOR GENE-EXPRESSION IN MUSCLE BYAGGREGATION OF MUSCLE-DERIVED NEUREGULINS

The neural isoforms of agrin can stimulate transcription of the acetyl choline receptor (AChR) epsilon subunit gene in electrically active mu scle fibers, as does the motor neuron upon the formation of a neuromus cular junction. It is not clear, however, whether this induction invol ves neuregulins (NRGs), which stimulate AChR subunit gene transcriptio n in vitro by activating ErbB receptors. In this study, we show that a grin-induced induction of AChR epsilon subunit gene transcription is i nhibited in cultured myotubes overexpressing an inactive mutant of the ErbB2 receptor, demonstrating involvement of the NRG/ErbB pathway in agrin-induced AChR expression. Furthermore, salt extracts from the sur face of cultured myotubes induce tyrosine phosphorylation of ErbB2 rec eptors, indicating that muscle cells express biological NRG-like activ ity on their surface. We further demonstrate by RT-PCR analysis that m uscle NRGs have Ig-like domains required for their immobilization at h eparan sulfate proteoglycans (HSPGs) of the extracellular matrix. In e xtrasynaptic regions of innervated muscle fibers in vivo, ectopically expressed neural agrin induces the colocalized accumulation of AChRs, muscle-derived NRGs, and HSPGs. By using overlay and radioligand-bindi ng assays we show that the Ig domain of NRGs bind to the HSPGs agrin a nd perlecan. These findings show that neural agrin can induce AChR sub unit gene transcription by aggregating muscle HSPGs on the muscle fibe r surface that then serve as a local sink for focal binding of muscle- derived NRGs to regulate AChR gene expression at the neuromuscular jun ction.