In recent years there has emerged a striking realization that liquid-liquid phase separation of proteins and nucleic acids is responsible for the formation of various intracellular membraneless organelles. Examples of organelles formed by phase separation are nucleoli and Cajal bodies in the nucleus and stress granules and P granules in the cytoplasm. The phase separation of protein-RNA composites, in particular, is being appreciated for crucial roles of connecting gene regulatory processes with the phenotypic complexity of eukaryotes. Despite the appar- ent biological signi?cance and extensive experimental efforts, our understanding of the mechanisms which link protein-RNA phase separation with the transcriptional and catalytic processes is still lacking. The fundamental challenges stem from (i) The molecular heterogeneity and conformational ?exibility of RNA and proteins, which contain low complexity disordered regions (ii) The juxtaposition of molecular and cellular scales (iii) Presence of non-equilibrium effects due to biochemical reactions, ATP driven processes, and irreversible bio-polymer ?uxes. The current theoretical and computational paradigms often lack optimal spatio-temporal resolution and the right combination of physical insights for confronting the complex experimental data in a comprehensive and integrative manner. Here, I propose using multi-scale computational tools developed in our lab combined in conjunction with data-driven approaches for revealing general mechanistic principles of protein-RNA phase separation and its link with the functional regulatory processes. The proposal consists of three directions. In the ?rst direction, we focus on hierarchical coarse-graining of proteins and RNA for studying thermodynamic driving forces of liquid-liquid phase separation in in vitro via molecular dynamics techniques. In the second direction, we employ ?nite-element and reaction-diffusion simulations trained by molecular models and experimental data for studying the connec- tion of liquid-liquid phase separation with transcriptional and catalytic reactions, which is characteristic of in vivo conditions. In the third direction, we assess the impact of protein-RNA phase separation generic gene regulatory networks by using stochastic dynamics simulations. The speci?c systems chosen for the study are experimen- tally well-characterized RNA binding proteins FUS, TDP-43, Tau, hnrpa1, and hnrpa2. These systems are known for forming liquid protein-RNA condensates under usually regulated conditions and aggregated structures when misregulated, thereby leading to major neurodegenerative diseases. The completion of the proposed research program will elucidate the nature of the protein-RNA phase sepa- ration its link with functional biochemical reactions and provide much-needed insights for developing intervention strategies for halting protein aggregation into diseases inducing cellular bodies.
The proposed research will reveal general mechanistic principles of protein-RNA liquid-liquid phase separation, the misregulation of which is known to be a signi?cant source of neurodegenerative diseases. Our study will indicate novel preventative strategies guided by simulations and theory, which could be used to control the protein- RNA condensates and combat processes that lead to protein aggregation linked with mental disorders.
