Inferring the Physics of mRNA Trafficking in Neuronal Systems

Massachusetts Institute Of Technology, Cambridge MA

Investigators

Abstract

Active transport mechanisms of messenger RNAs (mRNAs) are core to neuronal network development and function. Fluorescence imaging is a powerful approach to resolving the physical basis of mRNA transport using direct reporters of mRNA location and copy number in live cells. However, resolving these mechanisms requires quantitative, physics-based approaches that model ribosome-mRNA associations, copy numbers, and recruitment to synaptic and cytoskeletal sites where local translation is needed. The present project integrates live-cell imaging with physics-based modeling and inference of stochastic molecular transport and copy number variations to characterize the molecular basis of neuronal synapse development that is core to brain development and function in living systems. Educational initiatives advanced by the PI include undergraduate and graduate curriculum enhancements including a discussion based seminar course on the physics of living systems that is taught jointly between the Departments of Biological Engineering, Physics, and Biology at MIT. The PI is additionally active in developing and maintaining free web servers that distribute worldwide physics-based inference procedures developed by his group. The PI participates in outreach to under-represented minorities through teaching in an annual workshop organized by the Department of Biology over the inter-activity period in January. Educational and research activities of the PI are translated to undergraduate students through MIT's Undergraduate Research Opportunities Program, as well as through host visitations of international students from foreign countries. The objective of this project is to understand the translational dynamics of messenger RNAs that are regulated by active transport mechanisms in neuronal cells. Neurons consist of highly elongated axonal and dendritic processes that extend hundreds to thousands of cell bodies away from the nucleus, where transcription occurs. Consequently, neuronal plasticity in development and learning requires synaptic and cytoskeletal proteins to be synthesized locally within these extended processes that cannot be reached by passive mRNA transport mechanisms alone. To achieve this function, mRNAs are actively trafficked by molecular motors to synaptic sites to enable local protein production. In this project the PI will apply single-molecule live-cell imaging together with physics-based modeling of mRNA and ribosomal transport to understand the molecular basis of neuronal mRNA transport. The PI will develop stochastic modeling and inference procedures to infer the association dynamics of mRNAs and ribosomes, as well as their physical association with filamentous actin networks, microtubules, and synaptic proteins to identify cellular landmarks that regulate mRNA translation. Fluorescence fluctuation analysis is performed to infer copy numbers of mRNAs and ribosomes in ribonucleoprotein complexes, which are cross-validated using multiplexed super-resolution fluorescence imaging in fixed neuronal samples with exchangeable DNA probes. This work will help resolve the physical basis of mRNA recruitment and trafficking in the formation and turnover of synapses that are central to neuronal development and plasticity. This project is being jointly supported by the Physics of Living Systems program in the Division of Physics and the Cellular Dynamics and Function Cluster in the Division of Molecular and Cellular Biosciences.