Sequence Analysis of a tRNA Gene Cluster: Drosophila Leucine-tRNA Genes contain Intervening Sequences

A recombinant DNA phage containing a cluster of Drosophila melanogaster tRNA genes has been isolated and analyzed. In situ hybridization localizes this phage to chromosomal region 50AB, a known tRNA site. Nucleotide sequencing of the entire Drosophila tRNA coding region reveals seven tRNA genes spanning 2.5 kb of chromosomal DNA. This cluster is separated from other tRNA regions on the chromosome by at least 2.7 kb on one side, and 9.6 kb on the other. Two tRNA genes are nearly identical and contain intervening sequences in the anticodon loop. These two genes are assigned to be tRNALeu genes because of significant sequence homology with yeast tRNA3Leu, and secondary structure homology with yeast tRNA3Leu intervening sequences. In addition, an 8 base sequence (AAAAUCUU) is conserved in the same location in the inter- vening sequences of Drosophi1a tRNALeu genes and a yeast tRNA3Leu gene. Similar sequences occur in all other tRNAs containing intervening sequences. The remaining five genes are identica1 tRNAIle genes, which are identical to a tRNAIle gene from chromosomal region 42A. The 5' flanking regions are only weakly homologous, but each set of isoacceptors contains short regions of strong homology approximately 20 nucleotides preceding the tRNA coding sequences: GCNTTTTG preceding tRNAIles; and GANTTTGG preceding tRNALeus. The genes are irregularly organized on both DNA strands; spacing regions are divergent in sequence and length. Preliminary in vitro transcription experiments with isolated restriction fragments demonstrate that both tRNALeu genes and at least two of the five tRNAIle genes are selectively transcribed in vitro by cytoplasmic extracts from HeLa cells.