Defining the control and consequences of the biophysical properties of the nucleus

• Applicant: Dr. Tamas Szoradi
• Subject Area: Biophysics ; Biochemistry ; Cell Biology
• Term: from 2018 to 2021
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 417630603

The properties of the cell interior are crucial for the organization and efficiency of biochemical reactions, but the physical nature of the cytoplasm or the nucleus remains poorly understood, particularly at the mesoscale (10 nm – 1 µm). A salient feature of the cell interior is the high degree of macromolecular crowding. The diffusive and interactive behaviour of molecules is significantly altered by crowded environments, as evidenced by the requirement for crowding agents to drive many biochemical reactions in vitro. Furthermore, several nuclear structures such as nucleoli, Cajal bodies, and heterochromatin, have recently been described as phase-separated liquid droplets. Phase separation is strongly favoured by crowding. Consequently, the formation, dynamics and size of these compartments are likely to be strongly influenced by macromolecular crowding. Therefore, crowding is predicted to have a major impact on nuclear processes at the scale of both molecules and sub-nuclear compartments. Since it remains a mystery how these physical properties are regulated, the goal of my proposed research project is to define the control and physiological consequences of crowding in the nucleus. I will address this question using a new technology developed in my host laboratory. The group of Liam Holt recently developed Genetically Encoded Multimeric nanoparticles (GEMs) that greatly facilitate the characterization of the mesoscale properties of the cell. GEMs are self-assembling fluorescent nanoparticles that are encoded in a single gene. Key biophysical parameters including the degree of macromolecular crowding can be inferred from the motion of these probes. Preliminary experiments in the Holt lab revealed that inhibition of the TORC1 complex increased the mobility of nuclear-targeted GEMs. This observation suggests that TORC1 signalling regulates the mesoscale biophysical properties of the nucleus, which reveals a new, and so-far uncharacterized role for the TORC1 pathway. Therefore, my research objective is to determine the mechanisms by which the TORC1 pathway regulates the biophysical properties of the nucleus and to determine the consequences of this regulation. First, I will characterize the impact of TORC1 signalling on nuclear crowding and dynamics. I will use GEMs to investigate the effects of TORC1 inhibition on crowding in the nucleus and nucleolus and I will study chromosome dynamics at multiple heterochromatic and euchromatic loci. Second, I will take advantage of the GEM system and the power of yeast genetics to investigate mechanisms for the control of nuclear crowding by TORC1. Finally, I will determine the consequences of perturbations to nuclear crowding on transcription, splicing and mRNA transport. Through these experiments, we will begin to elucidate how the biophysical properties of the nucleus are controlled. Moreover, this project improves our understanding of the physiological relevance of macromolecular crowding in the nucleus.

• DFG Programme: Research Fellowships
• International Connection: USA
• Host: Dr. Liam Holt