Malvin Forson, BA, MPhil
CUNY Graduate Center
Studies on the Dynamic and Structural Basis of Opto-thermo Response in BcLOV4
Non-invasive tools for precise spatiotemporal control of cellular behavior in living systems are critically needed to advance our understanding of protein function and targeted therapeutic development. Two types of candidates for such control, proteins which sense either temperature or light, are particularly well understood in specific systems – but the molecular basis to integrate them within a single tool is poorly understood.
Here, we report the characterization of bimodal light & temperature-responsive behavior in BcLOV4, a fungal protein found in Botrytis cinerea. Prior work showed that this protein, which contains two Per-ARNT-Sim (PAS) domains that are often used as environmental sensors, aggregates and translocates to the plasma membrane in response to blue light. Here we studied how the residence time of BcLOV4 on the membrane is modulated by temperature, using a combination of in vitro (Hydrogen/Deuterium exchange Mass Spectrometry [HDX-MS], Limited Proteolysis Mass Spectrometry) and cellular techniques. We show that BcLOV4 generates activation dynamics as a single protein through competitive intramolecular interactions between two sensory domains under the actuation of temperature and light. Temperature-sensing was encoded in a modular domain that could be tuned by mutations. HDX-MS revealed an N-terminal motif in this domain that unusually exhibits increased protection at 37°C relative to 25°C, suggesting a thermally driven conformational stabilization.
Non-invasive tools for precise spatiotemporal control of cellular behavior in living systems are critically needed to advance our understanding of protein function and targeted therapeutic development. Two types of candidates for such control, proteins which sense either temperature or light, are particularly well understood in specific systems – but the molecular basis to integrate them within a single tool is poorly understood.
Here, we report the characterization of bimodal light & temperature-responsive behavior in BcLOV4, a fungal protein found in Botrytis cinerea. Prior work showed that this protein, which contains two Per-ARNT-Sim (PAS) domains that are often used as environmental sensors, aggregates and translocates to the plasma membrane in response to blue light. Here we studied how the residence time of BcLOV4 on the membrane is modulated by temperature, using a combination of in vitro (Hydrogen/Deuterium exchange Mass Spectrometry [HDX-MS], Limited Proteolysis Mass Spectrometry) and cellular techniques. We show that BcLOV4 generates activation dynamics as a single protein through competitive intramolecular interactions between two sensory domains under the actuation of temperature and light. Temperature-sensing was encoded in a modular domain that could be tuned by mutations. HDX-MS revealed an N-terminal motif in this domain that unusually exhibits increased protection at 37°C relative to 25°C, suggesting a thermally driven conformational stabilization.
