AGRIN MESSENGER-RNA VARIANTS ARE DIFFERENTIALLY REGULATED IN DEVELOPING CHICK-EMBRYO SPINAL-CORD AND SENSORY GANGLIA

Agrin is a synapse-organizing protein synthesized and externalized by motor neurons in the spinal cord, which organizes the postsynaptic apparatus of the developing neuromuscular junction. Agrin mRNA in the nervous system consists of several alternatively spliced variants. Splicing of agrin gene transcripts at the major site of variability results in four variants: encoding 8 (B-8) or 11 (B-11) amino acid inserts, both (B-19), or a predominant variant (B-0) without inserts, The insert-containing, variants are neuron specific and encode agrin proteins with greater synapse-organizing activity than the B-0 variant. Here, we report the localization and developmental regulation of agrin mRNA variants in chick embryo spinal cord and dorsal root ganglia, In situ hybridization using antisense oligodeoxynucleotide (ODN) probes specific for the B-8 and B-11 sequences shows that the neuron-specific variants are concentrated in ventrolateral cells of the chick embryo spinal cord, presumably motor neurons, beginning at embryonic day 4 (E4). By E14, the insert-containing mRNAs are found almost exclusively in presumptive motor neurons. These variants are also found in dorsal root ganglia and sympathetic ganglia, but not in non-neural tissues. Analysis by polymerase chain reaction showed that the B-11 and B-19 mRNA variants appeared in spinal cord at E4, whereas the B-8 variant was first seen at E14. During development, B-11 decreased and disappeared by E20, whereas B-8 increased from E14 to E20, A similar time course a as seen in dorsal root ganglia. The greatest acetylcholine receptor-aggregating activity in the spinal cord was seen from E6 to E10, coincident with the highest proportion of B-11-containing transcripts and with the peak of synaptogenesis in limb muscles. These data provide the first evidence linking appearance of the neuron-specific agrin mRNA variants with expression of the functional protein, The B-11 and B-19 variants appeared in E2 (stage 15) neural tubes cultured for 2 days with or without notochord and trunk tissues, indicating that there is no peripheral signal required to induce these agrin mRNA variants in developing motor neurons. (C) 1995 John Wiley and Sons, Inc.