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. 2023 Apr 12;616(7958):764–773. doi: 10.1038/s41586-023-05927-7

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Extended Data Fig. 11 — a. BioID2 that is targeted to the plasma membrane biotinylates proteins near the plasma membrane. b. Label-free based quantification comparison of significant proteins (Log₂FC > 1 and FDR < 0.05 versus GFP controls) detected in the cytosolic Astro BioID2 and plasma membrane Astro Lck-BioID2 reveal plasma membrane enriched proteins. Top half of the volcano plot shows 238 unique Lck-BioID2 proteins when compared to cytosol. The top four most abundant proteins for Lck-BioID2 are shown. Lower half of volcano plot shows comparison of 144 proteins that were common in both cytosolic BioID2 and Lck-BioID2. The five highest enriched proteins for Lck-BioID2 (Log₂FC > 2) are shown. Magenta label shows protein that was validated with IHC in panel d. Red label shows Dlgap3/SAPAP3 is enriched in the astrocyte plasma membrane. c. Heat map shows the rank-rank hypergeometric overlap (RRHO) of the RNA and protein rank for the 270 plasma membrane proteins. Each pixel represents the significance of overlap between the two datasets in –log₁₀(P-value). Red pixels represent highly significant overlap. Color scale denotes the range of P-values at the negative log₁₀ scale (bin size = 100). d. IHC analysis of Slc4a4 (Nbc1) protein in tdTomato and Lck-GFP labeled astrocytes shows co-localization within the astrocyte territory. Scale bar represents 20 μm. e. Co-localization analysis using Pearson’s r co-efficient shows high co-localization between Lck-GFP and Slc4a4 (Nbc1). The mean and SEM are shown (n = 8 tdTomato+ cells from 4 mice; Two-tailed paired t-test). f. Scale-free STRING analysis protein-protein association map of the 270 unique and enriched biotinylated proteins identified in astrocyte plasma membrane with Astro Lck-BioID2 . Node size represents the enrichment of each protein vs the GFP control (log2(BioID2/GFP)). Edges represent putative interactions from the STRING database. Bar graphs show the functional enrichment analysis of the 270 proteins using “Biological process”, “Cellular component”, and “Molecular function” terms from Enrichr.