Search bioRxiv⌕ Search

Biology subjects

Michaels, T.

Publications and source records attributed to Michaels, T..

2 recordsLinked to original sources

Spatiotemporal properties of glutamate input support direction selectivity in the dendrites of retinal starburst amacrine cells

AO_SCPLOWBSTRACTC_SCPLOWThe asymmetric summation of kinetically distinct glutamate inputs across the dendrites of retinal "starburst" amacrine cells is proposed to underlie their direction selective properties, but experimentally verifying input kinetics has been a challenge. Here, we used two-photon glutamate sensor (iGluSnFR) imaging to directly measure the input kinetics across individual starburst dendrites. We found that signals measured from proximal dendrites were relatively sustained compared to those measured from distal dendrites. These differences were observed across a range of stimulus sizes and appeared to be shaped mainly by excitatory rather than inhibitory network interactions. Temporal deconvolution analysis suggests that the steady-state vesicle release rate was [~] 3 times larger at proximal sites compared to distal sites. Using a connectomics-inspired computational model, we demonstrate that input kinetics play an important role in shaping direction selectivity at low stimulus velocities. Together, these results provide direct support for the space-time wiring model for direction selectivity.

neuroscience↗

Tie-lines reveal interactions driving heteromolecular condensate formation

Phase separation of biomolecules give rise to membraneless organelles that contribute to the spatiotemporal organisation of the cell. In most cases, such biomolecular condensates contain multiple components, but the manner in which interactions between components control the stability of condensates remained challenging to elucidate. Here, we develop an approach to determine tie-line gradients in ternary liquid-liquid phase separation (LLPS) systems, based on measurements of the dilute phase concentration of only one component. We show that the sign of the tie-line gradient is related to the cross-interaction energy between the polymers in the system and discriminates between competitive and cooperative phase separation. Using this approach, we studied the interaction between protein Fused in Sarcoma (FUS) and polyethylene glycol (PEG) polymer chains, and measured positive tie-line gradients. Our results show that PEG drives LLPS through an associative interaction with FUS and is not an inert crowder. We further studied the interaction between PolyA RNA (3.0{+/-}0.5kDa) and the protein G3BP1, and using the tie-line gradient as a reporter for the stoichiometry of polymers in the condensate we determined a G3BP1-to-PolyA RNA molar ratio of 1:4 in the dense phase. Our framework for measuring tie-line gradients opens up a route for the characterisation of interaction types and compositions in ternary LLPS systems.

biophysics↗