Tagless LysoIP for immunoaffinity enrichment of native lysosomes from clinical samples.

Autor:	Saarela D; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Lis P; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Gomes S; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Nirujogi RS; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Dong W; Aligning Science Across Parkinson's (ASAP) Collaborative Research Network, Chevy Chase, United States of America., Rawat ES; Aligning Science Across Parkinson's (ASAP) Collaborative Research Network, Chevy Chase, United States of America., Glendinning S; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Zeneviciute K; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Bagnoli E; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Fasimoye R; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Lin C; Aligning Science Across Parkinson's (ASAP) Collaborative Research Network, Chevy Chase, United States of America., Nyame K; Aligning Science Across Parkinson's (ASAP) Collaborative Research Network, Chevy Chase, United States of America., Boros FA; Department of Molecular Neurology, University Hospital Erlangen, Friedrich-Alexander-University Erlangen-Nürnberg, Erlangen, Germany., Zunke F; Department of Molecular Neurology, University Hospital Erlangen, Friedrich-Alexander-University Erlangen-Nürnberg, Erlangen, Germany., Lamoliatte F; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Elshani S; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Jaconelli M; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Jans JJ; Department of Metabolic Diseases, Wilhelmina Children's Hospital, University Medical Center Utrecht, University Utrecht, Utrecht, Netherlands., Huisman MA; Department of Metabolic Diseases, Wilhelmina Children's Hospital, University Medical Center Utrecht, University Utrecht, Utrecht, Netherlands., Posern C; Department of Pediatrics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany., Westermann LM; Department of Pediatrics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany., Schulz A; Department of Pediatrics, University Medical Center Hamburg-Eppendorf, Hamburg, Germany., van Hasselt PM; Department of Metabolic Diseases, Wilhelmina Children's Hospital, University Medical Center Utrecht, University Utrecht, Utrecht, Netherlands., Alessi DR; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom., Abu-Remaileh M; Aligning Science Across Parkinson's (ASAP) Collaborative Research Network, Chevy Chase, United States of America., Sammler EM; Medical Research Council Protein Phosphorylation and Ubiquitylation Unit, School of Life Sciences, University of Dundee, Dundee, United Kingdom.
Jazyk:	angličtina
Zdroj:	The Journal of clinical investigation [J Clin Invest] 2024 Dec 26. Date of Electronic Publication: 2024 Dec 26.
DOI:	10.1172/JCI183592
Abstrakt:	Lysosomes are implicated in a wide spectrum of human diseases including monogenic lysosomal storage disorders (LSDs), age-associated neurodegeneration and cancer. Profiling lysosomal content using tag-based lysosomal immunoprecipitation (LysoTagIP) in cell and animal models has substantially moved the field forward, but studying lysosomal dysfunction in human patients remains challenging. Here, we report the development of the 'tagless LysoIP' method, designed to enable the rapid enrichment of lysosomes, via immunoprecipitation, using the endogenous integral lysosomal membrane protein TMEM192, directly from clinical samples and human cell lines (e.g., induced pluripotent stem cell derived neurons). Isolated lysosomes were intact and suitable for subsequent multimodal omics analyses. To validate our approach, we applied the tagless LysoIP to enrich lysosomes from peripheral blood mononuclear cells derived from fresh blood of healthy donors and patients with CLN3 disease, an autosomal recessive neurodegenerative LSD. Metabolic profiling of isolated lysosomes revealed massive accumulation of glycerophosphodiesters (GPDs) in patients' lysosomes. Interestingly, a patient with a milder phenotype and genotype displayed lower accumulation of lysosomal GPDs, consistent with their potential role as disease biomarkers. Altogether, the tagless LysoIP provides a framework to study native lysosomes from patient samples, identify disease biomarkers, and discover human-relevant disease mechanisms.
Databáze:	MEDLINE
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