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10.1038/s44318-025-00660-5 http://scihub22266oqcxt.onion/10.1038/s44318-025-00660-5 41381733!?!41381733 Warning: file_get_contents(https://eutils.ncbi.nlm.nih.gov/entrez/eutils/elink.fcgi?dbfrom=pubmed&id=41381733&cmd=llinks): Failed to open stream: HTTP request failed! HTTP/1.1 429 Too Many Requests in C:\Inetpub\vhosts\kidney.de\httpdocs\pget.php on line 215       EMBO+J 2025 ; ? (?): ? Nephropedia Template TP {{tp|p=41381733|t=2025. Integrating endogenous TurboID and data-independent acquisition mass spectrometry for in vivo proximity labeling |pdf=|usr=}}{{41381733}} gab.com Text Integrating endogenous TurboID and data-independent acquisition mass spectrometry for in vivo proximity labeling JO: EMBO J http://www.kidney.de/pget.php?&tw=1&pmid=41381733 #FOAvip Proximity labeling has emerged as a powerful approach for identifying protein-protein interaction networks within living systems, particularly those involving weak or transient associations. Here, we present a comprehensive revised proximity labeling workflow, integrating TurboID labeling of endogenously expressed fusion proteins and data-independent acquisition (DIA) mass spectrometry (MS). We benchmark this pipeline with a study of five conserved Caenorhabditis elegans proteins-NEKL-2, NEKL-3, MLT-2, MLT-3, and MLT-4- that form two NEKL-MLT kinase-scaffold subcomplexes involved in membrane trafficking and actin regulation. Profiling of NEKL-MLT interactomes across 23 experiments validated our approach through the identification of known NEKL-MLT binding partners and conserved nekl-mlt genetic interactors, including the discovery of several novel functional interactors. Importantly, inclusion of methodological variations, stringent controls, and filtering strategies enhanced sensitivity and reproducibility, defining a set of intuitive quantitative metrics for routine assessment of experimental quality. We show that DIA-based interactome workflows produce physiologically relevant findings, even in the presence of experimental noise and variability across biological replicates. Our study underscores the utility of DIA mass spectrometry in proximity labeling applications and highlights the value of incorporating internal controls, quantitative metrics, and biological validation to enhance confidence in candidate interactors. Twit Text FOAVip Integrating endogenous TurboID and data-independent acquisition mass spectrometry for in vivo proximity labeling ????EMBO J http://www.kidney.de/pget.php?&tw=1&pmid=41381733 #FOAvip Twit Text # Free Paper on # # # # # LINK http://www.kidney.de/pget.php?&tw=1&pmid=41381733 #FOAvip English Wikipedia <ref>{{Cite pmid|41381733}}</ref> Integrating endogenous TurboID and data-independent acquisition mass spectrometry for in vivo proximity labeling #MMPMID41381733 Fay DS; Balasubramaniam B; Harrington SM; Edeen PT EMBO J 2025[Dec]; ? (?): ? PMID41381733show ga Proximity labeling has emerged as a powerful approach for identifying protein-protein interaction networks within living systems, particularly those involving weak or transient associations. Here, we present a comprehensive revised proximity labeling workflow, integrating TurboID labeling of endogenously expressed fusion proteins and data-independent acquisition (DIA) mass spectrometry (MS). We benchmark this pipeline with a study of five conserved Caenorhabditis elegans proteins-NEKL-2, NEKL-3, MLT-2, MLT-3, and MLT-4- that form two NEKL-MLT kinase-scaffold subcomplexes involved in membrane trafficking and actin regulation. Profiling of NEKL-MLT interactomes across 23 experiments validated our approach through the identification of known NEKL-MLT binding partners and conserved nekl-mlt genetic interactors, including the discovery of several novel functional interactors. Importantly, inclusion of methodological variations, stringent controls, and filtering strategies enhanced sensitivity and reproducibility, defining a set of intuitive quantitative metrics for routine assessment of experimental quality. We show that DIA-based interactome workflows produce physiologically relevant findings, even in the presence of experimental noise and variability across biological replicates. Our study underscores the utility of DIA mass spectrometry in proximity labeling applications and highlights the value of incorporating internal controls, quantitative metrics, and biological validation to enhance confidence in candidate interactors. ?Integrating endogenous TurboID and data-independent acquisition mass s(epmc).pdf DeepDyve Pubget Overpricing