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Title: Characterisation of dstpk61 in Drosophila development and the role of extramacrochaetae in Drosophila oogenesis
Author: Papadia, Sofia
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2005
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This investigation focused on two genes, dstpk6l and emc; the former was identified in a screen for genes encoding sex-specific transcripts and we aimed at understanding its potential role in oogenesis and spermatogenesis, while the latter was identified in a screen for genes with interesting expression patterns in follicle cells and we aimed at elucidating its role in oogenesis, mainly in dorsal appendage formation and also in the early stages, during the potential oocyte determination or cyst envelopment by the follicle cells. The serine/threonine kinase dstpk6l is a key component of the insulin/IRS signalling pathway and is the Drosophila homologue of mammalian PDK1. It regulates cell growth and survival by phosphorylating a number of target kinases, most notably PKB/Akt, an anti-apoptotic protein kinase which is activated by combined PDK1 and rictor-mTOR complex phosphorylation. The dstpk6l locus produces several different transcripts and four predicted isoforms varying at the N’-terminus. Some transcripts were differentially expressed in male and female tissues. We demonstrate by RT-PCR and microarray analysis that transcript E51 which encodes the second longest dstpk6l isoform is highly enriched in male testes and present in very low levels in other tissues, whereas transcripts for the other three isoforms are present in male and female gonads and carcasses, with E40 and E09 being slightly ovary-enriched. The levels of expression of these transcripts vary depending on the age, nutritional/hormonally induced stress and the mating status of the flies. dstpk6l is expressed in follicle and germline cells in oogenesis, as shown by mRNA in situ hybridisation and antibody staining with an isoform­ specific antibody we generated for E40, and in spermatogenesis mRNA in situ hybridisation and RT-PCR shows dstpk6l E51 mRNA in testes rather than in accessory glands. The gene extramacrochaetae (emc) encodes a bHLH transcription factor which negatively regulates other HLH proteins by forming inactive heterodimers with them. We suggest that emc lies downstream of grk, Egfr and fs(1)K10 in the GRK/EGFR signalling pathway in the late stages of oogenesis and has a role in dorsal appendage formation, possibly involving fine-tuning of the Br-C expression area in the follicle cells that will give rise to the dorsal appendages. This research opens up the possibility of further study such as establishing the role of the dstpk6l E51 transcript in testes. Once expressed in vitro, different Dstpk6l protein isoforms can be used to identify the substrates of each one, which may shed light on why transcripts E09 and E40 are enriched in ovaries and E51 in testes. For emc, its role in the germline can be investigated and the mechanism by which it regulates dorsal appendage formation, as well as its interactions with other genes that participate in the Grk/EGFR pathway investigated.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available