A study of X-chromosome meiotic drive in the Palearctic fly Drosophila subobscura. Thesis submitted in accordance with the requirements of the University of Liverpool for the degree of Doctor in Philosophy (or other degree as appropriate) by Rudi Verspoor. 5th of April, 2017 *fighter picture credit Tom Simpson A-NC-ND 2.0, www.flickr.com Abstract This thesis examines a particular selfish genetic element (SGE), X-chromosome meiotic drive (XCMD), in the species Drosophila subobscura. XCMD is a system where the X-chromosome kills or disables Y-chromosome sperm to enhance their own transmission to the next generation. This also results in those males producing female biased broods. This selfish enhancement of their own transmission results in conflict with the rest of the genome that can be a potent force in evolution. The first chapters deal with sex and mating behaviour and how XCMD and other SGEs are linked to it. Chapter three focusses on the marking techniques and mating behaviour in three species of Drosophila. This work was completed while establishing the XCMD system from wild populations Chapter four presents case studies of how SGEs are intrinsically linked to sex. Chapter five examines XCMD in D. subobscura and reveals that this species is completely monandrous. This shows that polyandry does not play a role in preventing the spread of XCMD in this species, unlike in a number of other taxa which have XCMD. I also demonstrate weak female choice against XCMD in this chapter. In chapter six and seven I examine the XCMD phenotype when it is expressed in different population genetic backgrounds. I test for evidence of suppression and incompatibilities, when XCMD is exposed in four different populations (Tunisia, Morocco, Spain, and UK). I find evidence of suppression in North Africa, but no suppression in Europe. I also find evidence for severe incompatibilities specific to XCMD on European genetic backgrounds, which are absent in North African backgrounds. These results are consistent with genetic conflict causing rapid evolution in North Africa between XCMD and suppressors, which results in XCMD specific hybrid incompatibilities in naïve European populations. My final chapter evaluates how the testes proteomes of two species, D. subobscura and D. pseudoobscura, differ between XCMD and non-XCMD individuals. This ongoing work identifies some putative candidate genes that could be involved in the network that results in XCDM in these species. Interestingly, very few strong candidate genes overlapped in the two species, supporting the idea that separate genes and mechanisms are responsible for the two XCMD systems. Contents List of abbreviations .............................................................................................................. 3 1 - Acknowledgements .......................................................................................................... 4 2- General introduction ......................................................................................................... 5 2.1 - What are selfish genetic elements and why should we study them? ......................................... 7 2.2 - Selfish genetic elements in the age of genetic modification ....................................................... 9 2.3 - X-chromosome meiotic drive and the Drosophila subobscura system ..................................... 11 2.4 - Overview of the thesis chapters ................................................................................................ 13 2.5 - References ................................................................................................................................ 17 3 - Dyeing insects for behavioural assays: the mating behaviour of anaesthetised Drosophila .......................................................................................................................... 21 3.1 - Abstract ..................................................................................................................................... 23 3.2 - Introduction ............................................................................................................................... 24 3.3 - Protocol ..................................................................................................................................... 26 3.4 - Representative Results ............................................................................................................. 31 3.5 - Discussion ................................................................................................................................. 34 3.7 - References ................................................................................................................................ 37 4 – Sex and Selfish Genetic Elements ................................................................................ 41 4.1 - Abstract ..................................................................................................................................... 43 4.2 – Introduction .............................................................................................................................. 44 4.3 - Short term impacts of mating behaviour on SGEs : Sperm-killing meiotic drive and polyandry ........................................................................................................................................... 48 4.4 - Long term impacts of mating systems on SGEs: Transposable elements, sex and mating systems ................................................................................................................................. 53 4.5 - Long term impacts of SGEs on mating systems: The case of cytoplasmic male sterility ............................................................................................................................................... 58 4.6 - Short term impacts of SGEs on mating systems: Endosymbiont manipulation ........................ 62 4.9 - References ................................................................................................................................ 65 5 - The ability to gain matings, not sperm competition, reduces the success of males carrying a selfish genetic element in a fly. ..................................................................... 73 5,1 - Abstract ..................................................................................................................................... 75 5.2 - Introduction ............................................................................................................................... 76 5.3 - Methods .................................................................................................................................... 81 5.4 - Results ...................................................................................................................................... 87 5.5 - Discussion ................................................................................................................................. 91 5.6 - Conclusions .............................................................................................................................. 95 5.7 - Acknowledgements ................................................................................................................... 96 5.8 - References ................................................................................................................................ 97 1 6 - Incipient reproductive isolation prevents the spread of a meiotic drive element .......... 105 6.1 - Abstract ................................................................................................................................... 107 6.2 - Main text ................................................................................................................................. 108 6.3 - Methods .................................................................................................................................. 117 6.4 - References .............................................................................................................................. 120 6.5 - Supplementary information ..................................................................................................... 124 7 - A single evolution of an X-chromosome meiotic drive system has spread across multiple populations of Drosophila subobscura ............................................................ 133 7.1 - Abstract ................................................................................................................................... 134 7.2 - Introduction ............................................................................................................................. 136 7.3 Methods .................................................................................................................................... 141 7.4 - Results .................................................................................................................................... 144 7.5 - Discussion ............................................................................................................................... 148 7.6 References ................................................................................................................................ 153 7.7 Supplementary material ...........................................................................................................