Browsing by Subject "Organelles"
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Item Characterization of the Roles of Intrinsically Disordered Regions from RNA-Binding Proteins in Phase Separation(2016-05-27) Lin, Yuan; Ross, Elliott M.; Liu, Qinghua; Tu, Benjamin; Rosen, Michael K.Eukaryotic cells organize complex biochemical reactions through compartmentalization. While many intracellular compartments are enclosed by membranes, others are not. Messenger ribonucleoprotein (mRNP) granules are membrane-less organelles that enrich RNA and RNA-binding proteins containing intrinsically disordered regions (IDRs). I demonstrate that IDRs, coupled with RNA binding domain and RNA, can phase separate in vitro, producing dynamic liquid droplets. Over time, these droplets mature into more stable states, as assessed by slowed fluorescence recovery after photobleaching and resistance to salt. Maturation often coincides with the formation of fibrous structures. Pathological mutation within IDRs leads to the acceleration of maturation. Different disordered domains can co-assemble into phase-separated droplets. In the case of the IDR from FUS (fused in sarcoma), I show that tyrosine residues are important in promoting phase separation. Either mutation of these aromatic residues or phosphorylation of the IDR disassembles liquid droplets. I further discover that the disassembly is due to the disruption of aromatic interactions mediated by critical tyrosine residues and therefore an increase in the overall solubility of proteins. Taken together, these studies demonstrate a plausible mechanism by which interactions between IDRs, coupled with RNA binding, could contribute to mRNP granule assembly in vivo by promoting phase separation. Progression from dynamic liquids to stable fibers may be regulated to produce cellular structures with diverse physiochemical properties and functions. Misregulation of maturation could contribute to diseases that are associated with aberrant mRNP granules. Posttranslational modifications of IDRs could modulate the assembly and disassembly of mRNP granules by altering the solubility of IDRs.Item Compositional Control of Phase-Separated Cellular Bodies(2018-04-04) Banani, Salman Ferozali; Yu, Hongtao; Thomas, Philip J.; Chen, Zhijian J.; Rosen, Michael K.Cellular bodies such as P bodies and PML nuclear bodies (PML NBs) appear to be phase separated liquids organized by multivalent interactions among proteins and RNA molecules. Although many components of various cellular bodies are known, general principles that define body composition are lacking. We have proposed a model for the formation of cellular bodies that is based on the polymerization-driven phase separation of key scaffold components of cellular bodies. We modeled cellular bodies using several engineered multivalent proteins and RNA. \textit{In vitro} and in cells these scaffold molecules form phase separated liquid droplets that are strongly enriched with the scaffold molecules. Analytical theories of polymerization suggest the resulting second phase contains large polymers of the multivalent scaffolds. Low valency client molecules partition differently into these structures depending on the stoichoimetric ratio of the scaffolds, with a sharp switch in recruitment across the phase diagram diagonal. Composition can switch rapidly through changes in scaffold concentration or valency. Natural PML NBs and P bodies show analogous partitioning behavior, suggesting how their compositions could be controlled by levels of PML SUMOylation or cellular mRNA concentration, respectively. Indeed, the engineered polySUMO/polySIM engineered scaffolds recruit many of the natural PML NB clients in a manner that depends on the SUMO:SIM stoichiometric ratio. Together, these data suggest a conceptual framework for considering the composition and control thereof of cellular bodies assembled through heterotypic multivalent interactions.Item Organelle Extravaganza: A Portable Science Suitable for High School Biology Students(2009-01-16) Hulsey, Jennifer Leigh; Calver, Lewis E.The goal of this thesis project was to create a portable Science Suitcase on organelles designed to supplement the current high school biology curriculum in the Dallas Independent School District. The Science Suitcase includes a narrated animation, three laboratory experiments, and an interactive game that can be borrowed by teachers and brought into their classrooms. The Science Suitcase will help enhance students' interest in science, bridge the gap between ninth grade biology and Advanced Placement biology, and help students meet the state of Texas science education requirements based on the National Science Education Standards.