Identification of five new genes on the Y chromosome of Drosophila melanogaster

Abstract

The heterochromatic state of the Drosophila Y chromosome has made the cloning and identification of Y-linked genes a challenging process. Here, we report application of a procedure to identify Y-linked gene fragments from the unmapped residue of the whole genome sequencing effort. Previously identified Y-linked genes appear in sequenced scaffolds as individual exons, apparently because many introns have become heterochromatic, growing to enormous size and becoming virtually unclonable. A TBLASTN search using all known proteins as query sequences, tested against a blastable database of the unmapped fragments, produced a number of matches consistent with this scenario. Reverse transcription-PCR and genetic methods were used to confirm those that are expressed, Y-linked genes. The five genes reported here include three protein phosphatases (Pp1-Y1, Pp1-Y2, and PPr-Y), an occludin-related gene (ORY), and a coiled-coils gene (CCY). This brings the total to nine protein-coding genes identified on the Drosophila Y chromosome. ORY and CCY may correspond, respectively, to the fertility factors ks-1 and ks-2, whereas the three protein phosphatases represent novel genes. There remains a strong functional coherence to male function among the genes on the Drosophila Y chromosome.

Figure 1

Y-linked protein phosphatases (Pp1-Y1 and Pp1-Y2 genes). (A) TBLASTN using a Drosophila PPP1 as the query sequence. (B and C) BLASTN searches using cDNA of Pp1-Y1 and Pp1-Y2, respectively, as query sequences. The numbers above the bars are the abridged accession nos. (AE003043 was abridged to 3043 and so on). The fragments labeled “RT-PCR” had no armU match and were sequenced de novo.

Figure 2

Phylogeny of the Drosophila serine–threonine protein phosphatase-catalytic subunits (neighbor-joining with Poisson correction and complete deletion; interior branch test for 1,000 replicates; ref. 15). The subfamilies are indicated in the figure. Chlam Pp1 is the Chlamydomonas protein phosphatase that controls flagellar beating (ref. ; AAD38856). The bar indicates the number of amino acid substitutions per site.

Figure 3

PPr-Y gene. (A) TBLASTN using CG13125 gene product as the query sequence. (B) BLASTN using the cDNA of the long splice variant of PPr-Y as the query sequence.

Figure 4

ORY gene. (A) TBLASTN using CG6059 gene product as the query sequence. (B) BLASTN using the cDNA of ORY as the query sequence.