Gene silencing mechanisms mediated by Aubergine piRNA complexes in Drosophila male gonad - PubMed (original) (raw)

FIGURE 4.

AT-chX-1 shows strong complementarity to vas mRNA. (A) The length of Chromosome X is 22,422,827 nt (accession number: AE014298). The A and B regions (shown in purple) where AT-chX-1 and -2 are coded span from 21,765,976 to 21,766,249 (274 nt) and from 21,773,609 to 21,773,781 (173 nt), respectively. The distance of the A and B regions is 7360 nt. AT-chX-1 in both regions corresponds to the sequences from 21,766,034 to 21,766,058 and from 21,773,667 to 21,773,691, respectively. AT-chX-2 in the A and B regions is from 21,766,122 to 21,766,147 and 21,773,775 to 21,773,780, respectively. The A and B regions are flanked by baggins{}2584 and baggins{}2585, and baggins{}2427 and R1{}2611, respectively. CG17686 and CG14621 are protein-coding genes located close to the A and B regions. (B) Schematic drawing of the vas mRNA. Nucleotide sequences of a partial vas mRNA (black) (173 nt; a region showing strong complementarity to the B region in A [∼81% identity] is shown) (accession number: NM165103), AT-chX-1 (red), and AT-chX-2 (blue) are indicated. Lowercase letters show nucleotides mismatched between vas mRNA and AT-chX-1 or AT-chX-2. Along with AT-chX-1, AT-chX-2 is also found in the Aub-associated small RNA identification experiment, but appeared just once. In contrast, AT-chX-1 appeared 103 times in 425 piRNA clones (∼24%). (C) Association of AT-chX-1 with Aub was confirmed by Northern blotting. The probe used was a DNA oligo containing a sequence complementary to AT-chX-1. (D) The protein levels of VAS are increased in the aub mutant fly testes. Ovary and testis lysates of wild-type (wt), aub − _/_+ and aub − /aub − were each probed with anti-Aub, anti-VAS, and anti-tubulin. In testes, VAS protein levels in aub − /aub − are about twofold compared with those in aub −/+ and wt, whereas they are not changed in the ovary lysates with or without Aub. This figure shows representative data from three individual experiments. Anti-tubulin was used for normalization. (wt) yellow white; (aub −/+) aubHN2/+; (aub − /aub −) aubHN2/aubQC42.