Fig. 6 | Tracking mitochondria in ALS motor axons

This item is part of the Figshare Project:
Early mitochondrial dysfunction revealed across FUS- and TARDBP-ALS at single cell resolution

From Data Availability Statement for the forthcoming paper in Nature Communications entitled:

Single-cell RNA sequencing reveals early mitochondrial dysfunction unique to motor neurons shared across FUS- and TARDBP-ALS

"We have deposited all raw and processed RNA sequencing data generated in this study on the NCBI Gene Expression Omnibus (GEO) under the accession number GSE226482. The C9orf72-ALS bulk RNA sequencing data was retrieved directly from the authors of the study.

[Items under this Figshare Project contain:] "Scans of fluorescent western blots, raw imaging files from confocal microscopy, the analysis files from Opera Phenix, qPCR data sets, and Seahorse assay result files."

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[Item specific description:]

Motor neurons were generated from iPSC by differentation according to Nijssen et al, 2019. BioProtocol (PMID: 33654821). Embryoid bodies containing motor neuron progenitors were attached whole at day 9, followed by maturation during which axons grow radially outward.

From day 27 onwards and as indicated, mitochondria were labelled with TMRM for 30 min prior to live cell imaging to record time-lapses.

These files are images and time-lapse videos of mitochondrial movement in iPSC-derived motor axons based on TMRM-stained mitochondria. The file format is .czi, through the ZEN software (Zeiss). Zeiss recommends using ImageJ and the ImageJ-based Fiji software package.

For analysis of mitochondrial movement, individual mitochondria were traced in Fiji using the TrackMate 7 extension with the StarDist detector and a Sparse LAP tracker (as in ‘mitotracker.py’). The mitochondrial traces were then annotated and analysed in R (version 4.2.0) using our custom package ‘mitotrackR’ (https://github.com/schwi24/mitotrackR).

Information about the microscopy file format CZI can be found here: https://www.zeiss.com/microscopy/en/products/software/zeiss-zen/czi-image-file-format.html