Global proteomics and affinity mass spectrometry analysis of human Schwann cells indicates that loss of NF1 alters cellular and mitochondrial metabolism - Global

ABSTRACT: Detailed global proteomics analysis for NF1 deficient cells and tissues is lacking. We investigated proteomes from immortalized human Schwann cells with (+/+) and without (-/-) NF1 to evaluate potential biomarkers and targets in NF1 deficient cells. We identified over 1900 proteins in both cell lines and find 148 proteins with differential expression levels based on genotype. Following Ingenuity Pathway analysis (IPA), we found multiple pathways were altered including decrease in “oxidative phosphorylation,” increases in “mitochondrial dysfunction”, and “glycolysis”, as well as changes in “Myelination Signaling Pathway.” Next, we stably transfected NF1 -/- cells with tagged mNf1 cDNAs (either WT or variant) and again evaluated the global proteome. We identified an overall trend of metabolic differences pertaining to oxidative phosphorylation, mitochondria dysfunction, and glycolysis in the mutant cDNA expressing cells compared to WT cDNA cells. We then validated differential expression of the following proteins: LAMC1, CYB5R3, and SOD2. Additionally, we used the encoded Strep tag on the cDNA to affinity purity NF1 Protein-Protein interactors from the Schwann cells expressing WT and variant cDNAs. We were able to identify 98 PPIs and 9 overlap with prior HEK293 cell datasets while 89 are unique to Schwann cells. GO analysis indicates many of these PPIs play a role in protein-transporting ATP synthase complex or are involved with intermediate filaments. While we were able to show that some of these bind differentially to variant isoforms, variations within NF1 did not significantly impact ability to bind partners. Finally, we show that loss of NF1 impacts Schwann cell mitochondrial respiration via Seahorse validating our proteomics data and indicating that NF1 plays a significant role in in mitochondrial metabolism that results in proteomics changes in Schwann cells.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Cell Culture

DISEASE(S): Neurofibromatosis 1

SUBMITTER: james mobley  

LAB HEAD: James Mobley

PROVIDER: PXD052439 | Pride | 2025-05-07

REPOSITORIES: Pride