Obara-Nixon-Abell-2023-Nature

Compiled code used for Obara + Nixon-Abell et al., 2023, Nature (Obara + Nixon-Abell et al. 2023, https://www.biorxiv.org/content/10.1101/2022.09.03.505525v1)

Each of the types of analysis in the paper use a single batch of code, which is maintained in a specific github repository, linked below.

Codes used for figure assembly, coloring, video generation, etc. are NOT maintained in these github repositories, since they are not generally applicable. You can find them instead at the following link in Figshare packaged with the data they were applied to:

Generalized code for every other part of the paper is freely availble in the following repositories, which are complete with their own readme.txt files that will guide a user through their use:

General sptPALM and tracking code: https://github.com/cjobara/sptPALM_general

ERMCS-specific code: https://github.com/cjobara/ERMCS_specific

Local Diffusion maps via inferenceMAP: https://github.com/cjobara/LocalDiffusion

Nonparametric Bayesian Analysis: https://github.com/cjobara/UrsaAnalyticsCode

FIB-SEM analysis and local curvatrue in 2D: https://github.com/cjobara/FIB-SEM_curvature

If you have any trouble, please reach out to Chris Obara for help: cjobara@gmail.com or obarac@janelia.hhmi.org.

Cheers,

Chris

Paper details:

Motion of VAPB molecules reveals ER-mitochondria contact site subdomains

To coordinate cellular physiology, eukaryotic cells rely on the rapid exchange of molecules at specialized organelle-organelle contact sites. Endoplasmic reticulum-mitochondrial contact sites (ERMCSs) are particularly vital communication hubs, playing key roles in the exchange of signaling molecules, lipids, and metabolites. ERMCSs are maintained by interactions between complementary tethering molecules on the surface of each organelle. However, due to the extreme sensitivity of these membrane interfaces to experimental perturbation, a clear understanding of their nanoscale organization and regulation is still lacking. Here, we combine 3D electron microscopy with high-speed molecular tracking of a model organelle tether, VAPB, to map the structure and diffusion landscape of ERMCSs. We uncovered dynamic subdomains within VAPB contact sites that correlate with ER membrane curvature and undergo rapid remodeling. We show that VAPB molecules enter and leave ERMCSs within seconds, despite the contact site itself remaining stable over much longer time scales. This metastability allows ERMCSs to remodel with changes in the physiological environment to accommodate metabolic needs of the cell. An amyotrophic lateral sclerosis (ALS)-associated mutation in VAPB perturbs these subdomains, likely impairing their remodeling capacity and resulting in impaired inter-organelle communication. These results establish high speed single molecule imaging as a new tool for mapping the structure of contact site interfaces and reveal that the diffusion landscape of VAPB at contact sites is a crucial component of ERMCS homeostasis.