Cellular energetics of long-term survival and storage in honeybee sperm

Ellen Paynter

Research output: ThesisDoctoral Thesis

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Abstract

Social insects are characterised by their colony structure and caste system. In many social insect species, reproductive females mate with a number of males only during a short period of time at the beginning of their life, and then store the sperm in a specialised organ called a spermatheca, and do not mate again. They use this sperm sparingly over their entire lifetime to produce workers for the colony. Sperm in the spermatheca must remain viable during the life of the queen, a period of up to 20 years in some species, and retain its capacity for fertilisation. The molecular mechanisms facilitating this long-term survival of sperm are largely unknown. However, it has been shown that sperm storage is costly for the queen, as her immune defence decreases with increasing numbers of sperm stored. Previous studies give indications that energy metabolism is of importance for sperm during storage. The honeybee, Apis mellifera, is a well-studied model species with many techniques and resources developed for its research. It is used as a model here to examine sperm survival and storage. This thesis investigates a number of key questions pertaining to sperm function, its maintenance and adaptations which occur during storage.

In the first chapter, a method for assessing sperm viability using flow cytometry was optimised and validated. This technique vastly decreases the time required for this assessment, and increases the accuracy of the measurements obtained. This method was then utilised in the following chapter.

In the second chapter, sperm metabolism was investigated using a number of techniques; initially metabolite profiling of the seminal fluid via GC-MS over time to analyse the composition of the fluid that the sperm are provided with during ejaculation. This work identified a number of putative substrates for sperm energy metabolism. These compounds were then verified or discounted as substrates, by measuring the rate of metabolism of the compounds in single ejaculates. Of note, glyceraldehyde-3-phosphate was identified as a compound that is metabolised rapidly by honeybee sperm, and yields more ATP than other metabolised compounds. It was also found to maintain sperm viability.

The third chapter investigated the possibility of sperm protein synthesis during storage. It was once assumed in all species that proteins are synthesised during spermatogenesis, and mature sperm were translationally silent, more recent literature has demonstrated that during capacitation, proteins important for fertilisation are synthesised in mammalian sperm. The work presented in Chapter 3 constitutes new evidence that protein synthesis does occur in mature sperm not undergoing capacitation, possibly as an adaptation to the extreme demands of long-term storage. The overrepresentation of members of key metabolic pathways in the newly synthesised protein population offered insights into the adaptation of energy production to facilitate long-term storage.

The results from the three chapters together give insight into the molecular mechanisms involved in sperm survival and storage through increased knowledge of sperm metabolism, adaptations occurring during storage and energetic costs for the honeybee.
Original languageEnglish
QualificationDoctor of Philosophy
Supervisors/Advisors
  • Baer, Boris, Supervisor
  • Millar, Harvey, Supervisor
Publication statusUnpublished - 2015

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