TY - JOUR
T1 - Tagless LysoIP for immunoaffinity enrichment of native lysosomes from clinical samples
AU - Saarela, Daniel
AU - Lis, Paweł
AU - Gomes, Sara
AU - Nirujogi, Raja
AU - Dong, Wentao
AU - Rawat, Eshaan S.
AU - Glendinning, Sophie
AU - Zeneviciute, Karolina
AU - Bagnoli, Enrico
AU - Fasimoye, Rotimi
AU - Lin, Cindy
AU - Nyame, Kwamina
AU - Boros, Fanni A.
AU - Zunke, Friederike
AU - Lamoliatte, Fred
AU - Elshani, Sadik
AU - Jaconelli, Matthew
AU - Jans, Judith J. M.
AU - Huisman, Margriet A.
AU - Posern, Christian
AU - Westermann, Lena M.
AU - Schulz, Angela
AU - van Hasselt, Peter M.
AU - Alessi, Dario
AU - Abu-Remaileh, Monther
AU - Sammler, Esther
N1 - Copyright:
© 2024, Saarela et al.
PY - 2023/12/26
Y1 - 2023/12/26
N2 - 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.
AB - 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.
U2 - 10.1172/JCI183592
DO - 10.1172/JCI183592
M3 - Article
SN - 0021-9738
JO - Journal of Clinical Investigation
JF - Journal of Clinical Investigation
ER -