READ ME File For 'Dataset in support of the Southampton doctoral thesis "Increasing axonal expression of alpha9 integrin to promote axon regeneration: exosome-mediated delivery and axon-targeting of the alpha9 integrin subunit."'

Dataset DOI: https://doi.org/10.5258/SOTON/D3110

ReadMe Author: LLOYD STEELE-NICHOLSON, University of Southampton ORCID ID https://orcid.org/0000-0002-6001-3386

These datasets support the thesis entitled "Increasing axonal expression of alpha9beta1 integrin to promote axon regeneration: exosome medited delivery and axon-targeting of the alpha9 integrin subunit."
AWARDED BY: Univeristy of Southampton
DATE OF AWARD: [2024]

DESCRIPTION OF THE DATA
Datasets including tif and pngs of representative immunofluorescence images, PDF reports of Nanosight data, and individual prism files containing details of raw values and statistical tests used for statistical comparisons. Data in organised by the corresponding chapter and figure to which it contributed.

Raw files relating to all datasets may be requested from Dr Melissa Andrews, University of Southampton.

This dataset contains:
Chapter 3: Immunofluorescence images of HEK-293 cells, embryonic astrocytes and adult mouse dorsal root ganglion neurons stained with antibodies against relevant targets at indicated concentrations. Raw western blots of exosome isolates and cell lysates used in production of western blot figures, probing cell lysates/exosome isolates with indicated antibodies. Representative images and Prism files containing raw values as output DRG outgrowth assays on inhibitory (tenascin-C) and growth-promoting (laminin) extracellular matrix substrates. PDF files as output of nanoparticle tracking analysis experiments carried out on the Nanosight 3000. Tif and PNG files can be opened in ImageJ/Fiji. Prism files can be opened using Prism.

Chapter 4: Immunoflourescence images and Prism files showing neurite/axon outgrowth and localisation of alpha9 integrin in PC12 and primary embryonic cortical neurons transiently expressing axon-targeting motif-modified alpha9 integrin constructs. Cells were transfected transiently with alpha9 integrin constructs and localisation measured after 96 hours (PC12) or 48h (DIV8 cortical neurons). Outgrowth of neurites in PC12 cells or axons in cortical neurons was also measured and is detailed in Prism files.

Chapter 5: Immunofluorescence images, western blots and Prism files showing neurite/axon outgrwoth and localisation of alpha9 integrin in PTEN KO or KD PC12 cells and embryonic cortical neurons, respectively. Cell lines were generated by transduction with lentiCRISPR LV vectors targeting rat/mouse/human PTEN gene. Immunfluorescence images were generated by probing western blots of lysates and/or fixed cells with antibodies against different targetes (PTEN, p-S6, beta1 integrin, Akt, p-Akt, FAK, p-FAK). Neurite/axon outgrowth assays were performed 48h after transfection with alpha9 integrin/GFP constructs (PC12 cells) or 4-5 days after transduction (cortical neurons). Localisation was measured 96h after transfection (PC12 cells) or 3-4 days after transduction (cortical neurons).

More details on how these data were collected can be found in general materials & methods chapter and the chapter specific materials & methods sections.


Date of data collection: [October 2019 to May 2023]

Information about geographic location of data collection: 
All data collected at Life Sciences Building 85, Highfield campus, University of Southampton, Southampton, SO17 1BJ, United Kingdom.

Licence:
CC-BY.

Related projects/Funders:
No related projects. Funded by Gerald Kerkut Charitable Trust and School of Biological Sciences, University of Southampton.

Related publication:
Steele-Nicholson LJ, Andrews MR. Axon-Targeting Motifs: Mechanisms and Applications of Enhancing Axonal Localisation of Transmembrane Proteins. Cells. 2022 Mar 9;11(6):937. doi: 10.3390/cells11060937. PMID: 35326388; PMCID: PMC8946247.

Date that the file was created: June, 2024.