Antimicrobial peptides (AMPs) are major innate immune mechanisms in insects. Among seven AMPs known to be in Drosophila melanogaster, drosomycin is recognized as a major AMP encoded by seven genes (Drs, Dro2, Dro3, Dro4, Dro5, Dro6 and Drs-1). Previous studies showed that there are different expression patterns between the seven drosomycin genes. Previous studies have shown that there are copy number variations in Dro2, Dro3, Dro4 and Dro5 among several species closely related to D. melanogaster. These variations are expected to have evolved in a short evolutionary time by repeated gene duplications and deletions. In this study, therefore, to infer the mechanisms of the frequent gene duplications and deletions, I compared the nucleotide sequences within a species using sixteen strains of D. lutescens because this species has a relatively large number of drosomycin gene copies. Previous studies showed that D. lutescens has three copies of Dro3 (Dro3-1, Dro3-2 and Dro3-3), in a genome region containing Dro2, Dro3, Dro4 and Dro5, named ‘Dro2-5 region' in this study. Determining ten nucleotide sequences of Dro2-5 region from D. lutescens, I found 3 different types of sequence length variants (standard, short and long alleles). Comparing the long and short allele nucleotide sequences to the standard allele by the dot plot analysis, I found that the long allele has an insertion of 810 bp upstream Dro3-1 and short allele has and deletion of 711 bp including the entire coding region of Dro3-1. These results suggested variability of copy number of Dro3 within species as well as between species. In addition, comparing amino acid sequences within D. lutescens, it is suggested that Dro3-1, Dro3-3, Dro4 and Dro5 are functionally conserved within species but Dro3-2 lost its function in two lines due to disruptions of three disulfide bonds required for the function of drosomycin. This suggests variability of Dro3 function within species. Furthermore, I found that there are more insertions and deletions in upstream Dro3-1 and intergenic regions between Dro3-3 and Dro4, whereas there are less insertions and deletions upstream Dro3-2 and upstream Dro3-3, suggesting conservation necessary for the regulation of gene expression. Dro3-2 and Dro3-3 possibly have more important role than Dro3-1. Comparing the sequences surrounding insertions and deletions, I found that more insertions and deletions have occurred between T and C (or G and A on the opposite strand). I also found that insertions and deletions of 10 bp or longer tend to have occurred in low G+C content regions, suggesting that the G+C content has an effect on the generation of insertion and deletion. It is suggested that some insertion and deletion event has a target sequence. Therefore, I further searched in Dro2-5 region of the nine species of the D. melanogaster subgroup for the fragments of transposable elements (TE), which often contain repetitive sequences responsible for insertions and deletions. As the result, I found that species having many copies of drosomycin gene tend to have many TE fragments, suggesting that the TE fragments may have an effect to generate the copy number variations.
首都大学東京, 2016-03-25, 修士（理学）