Transcriptome profiling of the spermatheca identifies genes potentially involved in the long-term sperm storage of ant queens

Ayako Gotoh,Katsushi Yamaguchi,Shuji Shigenobu,Satoru Kobayashi,Fuminori Ito,Kazuki Tsuji

doi:10.1038/s41598-017-05818-8

Abstract

Females of social Hymenoptera only mate at the beginning of their adult lives and produce offspring until their death. In most ant species, queens live for over a decade, indicating that ant queens can store large numbers of spermatozoa throughout their long lives. To reveal the prolonged sperm storage mechanisms, we identified enriched genes in the sperm-storage organ (spermatheca) relative to those in body samples in Crematogaster osakensis queens using the RNA-sequencing method. The genes encoding antioxidant enzymes, proteases, and extracellular matrix-related genes, and novel genes that have no similar sequences in the public databases were identified. We also performed differential expression analyses between the virgin and mated spermathecae or between the spermathecae at 1-week and 1-year after mating, to identify genes altered by the mating status or by the sperm storage period, respectively. Gene Ontology enrichment analyses suggested that antioxidant function is enhanced in the spermatheca at 1-week after mating compared with the virgin spermatheca and the spermatheca at 1-year after mating. In situ hybridization analyses of 128 selected contigs revealed that 12 contigs were particular to the spermatheca. These genes have never been reported in the reproductive organs of insect females, suggesting specialized roles in ant spermatheca.

Highlights

Reproductive success is crucial for sexual organisms, and a great diversity of reproductive strategies have been evolved in each species and sex, including copulation behaviours, sperm competition, investment for gamete production, efficiency of fertilization, and parental care
After removing the 14,894 ORFs with hits to bacterial sequences, assuming them as bacterial contaminations, and 12,005 ORFs with extremely low expression (RPKM values in all 23 samples of less than 1), the remaining 53,525 ORFs with a N50 of 1230 bp were defined as a reference gene set of C. osakensis
This study is the first report on large-scale gene expression profiling of the spermatheca to understand the molecular mechanisms of long-term sperm storage in ant queens

Summary

Introduction

Reproductive success is crucial for sexual organisms, and a great diversity of reproductive strategies have been evolved in each species and sex, including copulation behaviours, sperm competition, investment for gamete production, efficiency of fertilization, and parental care. Ants evolved the ability of queens to store sperm for an extremely longer time than honeybees (see above), they are useful for studies of the prolonged sperm storage mechanisms. The sperm pump comprises muscular layers that are located at the distal portion of the spermathecal duct and may regulate sperm migration into the spermathecal reservoir after mating or sperm release prior to insemination These morphologies are well characterized, little is known of the molecular functions of the spermatheca in ant queens. In the present study, we screened candidate genes that contribute to sperm storage functions using various differential gene expression analyses of the spermatheca and the body samples from Crematogaster osakensis ant queens using the RNA sequencing method (Fig. 1).

Methods

Results

Conclusion