No abstract

The amino acid, either a glutamine (Q) or an arginine (R), at the Q/R site of the pore-lining segment (M2) of a vertebrate AMPA receptor subunit critically influences the properties of the receptor. The R codon of the mammalian AMPA receptor subunit 2 (GRIA2) transcript is not coded by the chromosomal sequence, but is created by posttranscriptional RNA editing activities. On the other hand, the R codons of some teleost GRIA2 homologs are coded by chromosomal sequences. To elucidate the evolution of the utilization of Q/R RNA editing in modifying vertebrate GRIA2 transcripts, the GRIA2 genes of five fish species and an amphibian were studied. The putative hagfish GRIA2 homolog (hfGRIA2) encodes an R codon, whereas shark and bullfrog GRIA2 genes specify a Q codon at the genomic Q/R site. All gnathostoma GRIA2 genes possess an intron splitting the coding regions of M2 and the third hydrophobic region (M3). The intronic components required for Q/R RNA editing are preserved in all the Q-coding vertebrate GRIA2 genes but are absent from the R-coding GRIA2 genes. Interestingly, the hfGRIA2 is intronless, suggesting that hfGRIA2 is unlikely evolved from a Q/R editing-competent gene. Results of this study suggest that modification of GRIA2 transcripts by Q/R editing is most likely acquired after the separation of the Agnatha and Gnathostome. ß

The AMPA receptor (AMPAR), a pharmacologically defined ionotropic glutamate receptor, mediates fast excitatory synaptic transmission in the vertebrate central nervous system. Mammalian and avian AMPARs are assembled from the products of four genes (GRIA1-GRIA4) conserved in their translated sequences and gene organizations. Teleost fish also express AMPAR subunits; however, the AMPAR genes have not been extensively investigated in lower vertebrates. To elucidate the evolution of vertebrate AMPAR genes, reverse-transcriptase PCR-based surveys of subunits expressed in the brains of eight nonmammalian vertebrates were performed. The newly cloned vertebrate AMPAR subunits were classified by their sequence identities to the mammalian AMPAR subunits. The results of molecular and phylogenetic analyses indicated that the members of the AMPAR gene family increased from two in the jawless hagfish to four in the tetrapods and the shark and to more than four in the teleost fish. The sizes of AMPAR gene families correlate well with those of many multigene families observed in various vertebrates. Moreover, all vertebrates expressed at least one AMPAR subunit bearing an arginine (R) at the Q/R site, at which no invertebrate glutamate receptor subunit has been found to have an R residue, suggesting that the low calcium-permeable AMPARs appeared at early evolutionary stages of vertebrate central nervous systems. Uniquely, the loop 1 (L1) regions between hydrophobic domain 1 and hydrophobic domain 2 of the hagfish putative GRIA2 and all the teleost GRIA1 subunits were much longer than those of the remaining known ionotropic glutamate receptor subunits. The length and sequence of the L1 of teleost GRIA1 subunits were heterogeneous, suggesting that the amino acid residues in L1 were not highly selected.

An insertion element, ISAE1, was discovered during the molecular analysis of mutants defective in the autotrophic growth (Aut-) of Alcaligenes eutrophus H14, a mitomycin C-generated derivative of strain HI.ISAEI is 1,313 bp long, has 12-bp nearly perfect inverted terminal repeats, and contains an open reading frame that has a coding capacity of 408 amino acids. Direct repeats of 8 bp were generated by insertion of ISAEI into chromosomes or plasmids. Most insertions were found in the AT-rich target sites. The distribution of ISAEI is limited to A. eutrophus HI (ATCC 17698) and H16 (ATCC 17699). Variants with newly transposed copies of ISAEI could be isolated at an elevated frequency by changing the growth conditions.