Biophysical Society Bulletin | February 2024

Communities

KG: We also sought to understand the precise nature of the exocytosis/endocytosis cycle in various cell types, which continues to be probed and debated to this day with ever more precise measurements and better molecular understanding. One of the big technical advances around that time was the ability to image fluorescence and measure capacitance simul taneously in living cells. Mast cells were particularly useful for this, as the imaging capability made it possible to visualize the individual vesicles and the release of their contents. Bastien Gomperts at University College London found that permeabi lized mast cells released their histamine contents in response to stimulation with GTP- γ S. He had visited Erwin Neher’s lab in Göttingen, and together with Julio Fernandez they showed that mast cells degranulate when GTP- γ S was introduced via the whole-cell patch pipette, leading to a huge stepwise capacitance increase reflecting sequential fusion of individual vesicles. Shortly afterwards, experiments were performed using beige mouse mast cells, which have giant granules and enabled the first measurements of fusion pore conductance to be performed. People were also working with chromaffin cells and other excit able endocrine cells that undergo exocytosis in response to de polarization-evoked calcium influx, similar to neurons. In these cells you can trigger exocytosis with voltage-clamp pulses and measure capacitance changes to observe the precise timing of fusion relative to depolarization and calcium influx. Other tech niques that were becoming important around this time were triggering exocytosis through photorelease of caged calcium, which clarified the timing and concentration dependance of calcium-triggered exocytosis and endocytosis; electrochemical microelectrodes to measure the time course of release of cat echolamines from single vesicles via an expanding fusion pore; and total internal reflection fluorescence microscopy to image the movements and release from individual granules. ML: And you know, Bob was right. Within the first couple of years, the Subgroup expanded from just electrophysiologists

to also include fluorescence imaging groups like those of Wolf Almers (Oregon Health Science University) and Ron Holz (University of Michigan), structural biology groups, and others. The founding of this Subgroup gave experts in these different techniques a chance to discuss our scientific questions togeth er and come up with innovative new experiments. JK: How else has the Subgroup changed since 2003? ML: Oh, there’s been an explosion of different approaches. At first, it was just patch clamp electrophysiologists. Now there is much more live-cell imaging. It’s gotten so routine that it’s easy to forget how much biophysical advances underlie all of the different imaging techniques. KG: And the molecular biology and structural biology tools that have come onto the scene in the last 20 years have enabled so much more diversity in terms of approaches. JK: I agree. Of course, electron microscopy has been integral to the endocytosis and exocytosis field since the work of John Heuser and Tom Reese . But the more recent cryo-electron microscopy structures of fusion, fission, and protein complexes have provided unprecedented structural resolution into what these proteins are doing to help membranes fuse, bud, and pull apart. JK: What do you most look forward to about this Subgroup? ML: The most important things are to get together at the BPS Annual Meeting every year and have a great dinner, great talks at the Subgroup symposium, and a great keynote speaker. KG: Yes, there simply is no better place to interact with those who approach membrane fusion and fission using rigorous, quantitative biophysical approaches. It is also great to see the field become more diverse geographically and in terms of gen der, although we have much more work to do in this area.

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February 2024

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