Investigating the Role of TOR Signalling during Drosophila Oogenesis: Lessons from Follicle and Border Cells

Halder, Sudipta (2022) Investigating the Role of TOR Signalling during Drosophila Oogenesis: Lessons from Follicle and Border Cells. PhD thesis, Indian Institute of Science Education and Research Kolkata.

Text (PhD thesis of Sudipta Halder (14RS074)) 14RS074.pdf - Submitted Version Restricted to Repository staff only Download (4MB)

Abstract

The sequence of events following fertilization and subsequent development of a single embryo into an adult individual is conserved across metazoans. Cell division, growth, differentiation, cell death, cell shape changes and cell migration are indispensable processes both for embryonic development and also for maintenance of adult tissues and homeostasis. These processes are collectively referred to as morphogenesis that aids in pattern and adult form generation. During morphogenesis epithelial cells emerge as one of the most ubiquitous cell types found across all tissues. The mechanisms of epithelial morphogenesis generate the diversity of epithelial cells from pre-existing epithelia or from the mesenchymal cells. The diverse functions of epithelial cells emerge from the shapes in which they are found in – Cuboidal, Columnar and Squamous. Epithelial cells can often undergo cell shape transition from one shape to another to perform specialized functions. In addition, epithelial cells can also gain migratory ability via epithelial to mesenchymal transitions, to undergo cell migration and help in wound healing. Perturbations in these processes during embryonic development or during adult maintenance is associated with congenital disorders and tumorigenesis. However, the mechanisms of cell shape morphogenesis and cell migration are not completely understood. In this thesis I have investigated the role of Target of Rapamycin (TOR) kinase during the cell shape transition of cuboidal into squamous cell and in the collective migration of border cells. TOR is a conserved central regulator of cellular growth and maintains a balance between anabolism and catabolism by integrating nutrient availability and growth factor signaling. Tumor cell growth, proliferation and spreading is known to be fueled by TOR activity. On the other hand, tumor and cancer formation occurs when normal morphogenesis mechanisms become uncoordinated. Thus, understanding regulators and mechanisms of morphogenesis is critical to study pathological conditions. However, the role of TOR during normal morphogenesis events has remained relatively understudied. I have employed the follicular epithelial layer of Drosophila egg chambers during Drosophila oogenesis, as a model to study mechanisms of cell shape change and cell migration with respect to TOR signaling. The follicular epithelium encapsulates germline cells – 15 nurse cell and an oocyte. Here, the follicular layer remains cuboidal in shape until mid-oogenesis, when it undergoes morphogenesis into three populations – anterior squamous cells, posterior columnar cells and a subset of epithelial-turned-mesenchymal or epithelia derived cluster of 6-10 migratory cells called border cells. I observed that TOR dependent endocytosis is critical for the transition of cuboidal cells into squamous cells. TOR depletion not only affects endocytosis but also the timely removal of Fasciclin 2, a cuboidal cell specific lateral adhesion molecules, which is a critical step during cuboidal to squamous cell shape change. This modulation of squamous cell formation seems to be independent of S6K driven protein synthesis and REPTOR driven transcriptional machinery below TOR, but instead dependent on TOR regulated endocytosis. Additionally, my data also suggest that TOR might be negatively regulating Par-1 function in cuboidal cells to promote endocytosis and Fas2 removal to aid in cuboidal to squamous cell shape transition. With respect to cell migration, I show that TOR is required for the collective movement of border cells. Cytokine stimulation of the anterior cells, promote migratory fate in a subset of anterior follicle cells, called the border cells. The border cells detach from the epithelia and undergo directed migration towards the oocyte to complete border cell migration. I found that TOR is required in the border cells for delamination of the cluster as well as during the migratory phase by stabilization of forward protrusions and retraction thus controlling overall speed of the cluster. In contrast to cell shape change, I observed that REPTOR dependent transcriptional activity below TOR negatively regulates border cell delamination and migration. In sum, my thesis provides evidence for a role of TOR in regulating specific morphogenesis events of cell shape change transition and collective cell movement using distinct mechanisms of endocytic and transcriptional regulation, respectively.

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