Cell and mechanobiology of Asymmetric Cell Division

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PROJECT SUMMARY Generating cells with different fates, functions and behaviors is critically important for the development and maintenance of tissues, organs, and multicellular organisms. Cellular diversity can be generated through Asymmetric Cell Division (ACD). Stem cells utilize ACD to create differentiating sibling cells while maintaining the stem cell in the process. In addition to the asymmetric partitioning of proteins or RNAs, other mechanisms such as mechanical cues, sibling cell size asymmetry or organelle asymmetry could potentially also contribute to binary cell fate decisions. Here, I propose to use asymmetrically dividing Drosophila neuroblasts, the neural stem cells of the developing fly central nervous system, to investigate the cell and mechanobiology of ACD in vivo. Recently, we discovered that Non-muscle Myosin II-dependent cortical flows, induced through both polarity- and spindle-dependent cues, are implicated in the generation of sibling cell size asymmetry. I will investigate how cortical flows are induced and modulated with spatiotemporal precision to achieve reproducible sibling cell size asymmetry. Our recent discovery of Protein Kinase N (PKN), and the Rho GTPase pathway as inducers of cortical flows will provide molecular entry points. I will also investigate how cell size asymmetry contributes to cell fate decisions, using RNA sequencing, immunohistochemistry, and long-term live cell imaging in vivo. A second project encompassed in this research direction is aimed at investigating the molecular mechanisms and function of molecular centrosome asymmetry, which is manifested in biased microtubule organizing center (MTOC) activity in interphase. We identified new proteins and mechanisms, such as Kinesins, Pp4 and dynamic centriolar protein localization in mitosis, regulating centrosome asymmetry. Centrosome segregation is highly stereotypic in stem cells, but whether and how centrosome asymmetry affects cell fate decisions, remains to be resolved. We will use fly neural stem cells to investigate the mechanisms and functions of centrosome asymmetry during ACD. I am particularly interested in investigating whether centrosome asymmetry provides a mechanism for biased cell fate determinant segregation, either via asymmetric RNA or sister chromatid segregation. I will also investigate whether biased MTOC activity impacts transcriptional regulation via chromatin organization. This research program will benefit from several novel and innovative tools, consisting of live cell imaging, superresolution microscopy, RNA sequencing and acute protein mislocalization and perturbation systems (nanobody, optogenetics), which my lab implemented to probe cytoskeletal dynamics with high spatial and/or temporal precision in vivo. ACD is an evolutionary conserved mechanism and the proposed research program is medically significant because defects in ACD can cause neurodevelopmental disorders or cancer.