The Taiman transcriptional coactivator engages Toll signals to promote apoptosis and inter-tissue invasion in Drosophila Öffentlichkeit
Byun, Phil K. (Fall 2018)
Abstract
Tissue morphogenesis and remodeling is a tightly choreographed phenomenon fundamental to the development of multicellular organisms. Among the numerous developmental cues that can guide morphogenesis, steroid hormones stand out as distinct from local morphogen gradients for their ability to cause organism-wide transcriptional changes in response to systemic hormonal pulses. In Drosophila, the steroid hormone ecdysone (Ec) controls a number of tissue morphogenic events such as fusion of the thoracic discs into an intact dorsal thorax, activation and movement of hemocytes and immune cells, and overall cell growth. Ec exerts these effects by binding to its cognate receptor, the Ec receptor (EcR). Activation of EcR homologs in humans, such as the estrogen and androgen receptors, is often associated with invasive cancers. We have discovered that ectopic expression of an EcR co-activator called taiman (tai) can transform a normal wing epithelial to invade and penetrate the neighboring thorax. This unique and novel phenotype can be modified using alleles of known genetic interactors of tai such as EcR, yorkie (yki; the nuclear effector of the Hippo pathway), pvf2 and pvf3 (PDGF/VEGF related proteins 2 and 3). To ascertain a more complete landscape of the transcriptional changes induced by tai expression in invasive wing cells, we performed two different screens: (1) a genetic suppressor screen using genetic deficiencies (deletions) that tile across the entire Drosophila 2nd chromosome, (2) and an RNA-sequencing (RNA-seq) analysis of transcripts altered in Tai-expressing pupal wing cells relative to control wings cells. These parallel screens revealed that the Toll and immune deficient (IMD) pathways, which control expression of innate immunity and apoptotic genes, are active in Tai-expressing cells. Each of these pathways is activated by binding of ligand to cell-surface receptors: the Toll family of receptors is bound by secreted, processed Spätzle (Spz) ligands, and transmembrane peptidoglycan recognition proteins (PGRPs) serve as IMD receptors. We find that Tai expression in wing cells elicits a systemic Toll/IMD response in the absence of a pathogen, a phenomenon referred to as “sterile inflammation” that is often associated with locally invasive Drosophila tumors. Based upon published work linking Toll and IMD pathways to competitive killing of neighboring cells by faster growing “super-competitors” that overexpress dMyc, we posited that Tai-expressing wing cells express immune ligands, specifically the Spz proteins, and “kill” their way through the thoracic epidermis and into underlying tissue by activating Toll in these cells. Consistent with this hypothesis, a Toll/IMD reporter is activated in thoracic cells adjacent to invasive Tai-expressing wing cells and this correlates spatially with elevated apoptosis. Moreover, loss of function alleles of factors that act downstream of the Toll receptor dominantly suppress Tai-driven wing invasion. Intriguingly, a strong loss-of-function allele of the IMD pathway inhibitor caspar (casp) dominantly suppressed Tai invasion, implying that elevated IMD activity can prevent invasion. Upon further investigation, I found that Tai-expression hyper-sensitizes wing cells to casp dosage, and that casp heterozygosity causes Tai-expressing cells to undergo apoptosis, which in turn prevents them from invading thoracic tissue. These data led to my model that Tai-expressing cells elevate expression of Spz proteins, which kill neighboring cells, and Casp, which protects Tai cells from Spz-mediated death. When Casp is reduced, Tai-expressing cells succumb to Toll-driven cell death. In summary, these studies show a novel inter-tissue invasion model driven by an EcR co-activator Taiman that non-autonomously induces Toll-mediated killing of neighboring cells but the differing threshold for IMD activation protects Tai-expressing cells from apoptotic fate. Similar mechanism of local invasion is seen in human cancers where pro-inflammatory signals have been linked to invasive behavior of cancer cells, including breast and prostate. In the future, this novel aspect of Tai function may provide insight into immune-based interactions that contribute to the competitive advantage of human tumors overexpressing Tai homologs.
Table of Contents
Table of Contents
Chapter 1: An Introduction 1
Cellular movement and tissue invasion as a Biological Question 1
Features of Drosophila melanogaster 5
Border cell migration and cancer metastasis 6
Taiman and Ecdysone Receptor 9
Tissue Fusion 12
The Hippo pathway 13
Apoptosis and caspases in Drosophila 16
Innate immunity in Drosophila 18
Sterile Inflammation 22
Chapter 2: The Taiman transcriptional coactivator engages Toll
signals to promote apoptosis and inter-tissue invasion in Drosophila 24
Introduction 25
Results 28
Discussion 61
Material and Methods 67
Chapter 3: A deficiency screen for uncovering dominant modifiers of
Tai-driven wing invasion 71
Introduction 72
Results 74
Discussion 82
Material and Methods 83
Chapter 4: Requirement of steroid hormone production 84
in the wing epithelium for proper growth
Introduction 85
Results 89
Discussion 98
Material and Methods 100
Chapter 5: Future directions & concluding remarks 101
Additional deficiency screens 102
Reactive oxygen species 104
Autonomous vs. Non-autonomous 106
Additional binding partners of Tai in the nucleus 107
Role of shd and local steroid level in wound regeneration 109
Concluding remarks 112
Reference 113
Appendix 126
About this Dissertation
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Primary PDF
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The Taiman transcriptional coactivator engages Toll signals to promote apoptosis and inter-tissue invasion in Drosophila () | 2018-12-14 15:57:16 -0500 |
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Supplemental Files
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Mapped reads from HTS RNA-seq analysis of en>GFP and en>tai,GFP larval wing discs (Alphabetical gene list with corresponding read frequency (FPKM; fragments per kilobase mapped) for en>GFP (Sample/value 1) and en>tai,GFP (Sample/value 2) RNA samples. Fold change (GFP vs. tai,GFP) is presented in log base 2 (log[2]∆).) | 2018-12-14 14:15:13 -0500 |
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