Transcriptional study of photoperiodic response in aphids and characterization of candidate photoperiodic photoreceptors
Acyrthosiphon pisum (Hemiptera: Aphididae) is a highly seasonal aphid model species. During the favourable warm seasons, only viviparous parthenogenetic females (virginoparae) are produced. In autumn, when days shorten (i.e. when the photeperiod is shorter), A. pisum senses the difference in night length and the parthenogenetic females start producing males and, later, sexual females (also known as “oviparae”). Some naturally occurring mutants do not respond to photoperiodic shortening and reproduce asexually all year long, these are called “anholocyclic” strains. After mating with males, sexual females lay overwintering diapausing eggs that hatch in spring, resetting the cycle. The molecular pathways involved in the switch to sexual reproduction and the molecule (or molecules) used to sense day length (the photoperiodic photoreceptor) are unknown or poorly understood. The most widely accepted model in photoperiodic response is composed of three main elements: an input pathway, a core (with a clock and a counter) and an output pathway. Some of the molecular pathways and genes involved in photoperiodism are shared with the circadian clock. Previous studies have found that certain regions of the protocerebrum in the aphid brain are relevant for the photoperiodic response. Among them, the pars lateralis (pl) in the protocerebrum expresses the core clock genes period and timeless. Close to the pl, the pars intercerebralis (pi) expresses insulin-like proteins (ILPs), which have been proposed to be the so called “virginoparin”, a substance that triggers virginoparae development in embryos. In this thesis, we aim to shed to shed light on the mechanisms and genes that trigger photoperiodic response in aphids, with special interest in the photosensitive process by studying the light-sensitive opsins and cryptochromes. We used a transcriptomic study to compare the differential expression between aphids exposed to short day (SD) photoperiods that induce male (SD14) and sexual females (SD10). Moreover, we focused on the analysis of opsins and cryptochromes as cadidates in photoperiodic photoreception. We compared gene expression using RT-qPCR with an anholocyclic strain under different photoperiods and along the day. We sequenced opsin and cryptochromes transcripts and localized their expression with PCR and in situ hybridization in the brain. We found that aphids exposed to SD14 and SD10 differentially express genes related to hormone, neuropeptides and neurotransmitter biosynthesis, such as serotonin, juvenile hormone, dopamine and melatonin. Consistently with previous findings by our group, we found several genes involved in the insulin pathway or insulin-like peptides (ILPs). We also found enrichment of pathways responsible for light perception and brain remodelling. Some specific transcripts drew our attention, such as the three takeout-like genes which are underexpressed under SD14 and that could be related to sexual progeny production. We characterized the transcripts and identified the site of expression of the opsin repertoire and cryptochromes in the brain and/or eyes. In relation with photoperiodic photoreception, the genes Ap-SWO4, Ap-C-Opsin and cryptochrome 1 (cry1) (and probably cry2) are expressed in the pars lateralis (like the core clock genes tim and per). Under SD conditions, we found that opsin genes and cry1 are overexpressed and that sexual females express cry1 rhythmically. Our work supports the role of several pathways in the photoperiodic response, as well as the participation of opsins and cryptochromes in the photoperiodic response, mainly in photosensitivity.