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Cited by 871Advanced Proteomics Techniques and ApplicationsSingle-cell and spatial transcriptomicsMass Spectrometry Techniques and Applications

Cell type- and brain region-resolved mouse brain proteome

Open access
Sharma, Kirti, Schmitt, Sebastian, Bergner, Caroline G, Tyanova, Stefka, Kannaiyan, Nirmal, Manrique-Hoyos, Natalia, Kongi, Karina, Cantuti, Ludovico +5 more·2015·Nature Neuroscience
DOI 10.1038/nn.4160PMC PMC7116867
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Abstract

Brain transcriptome and connectome maps are being generated, but an equivalent effort on the proteome is currently lacking. We performed high-resolution mass spectrometry-based proteomics for in-depth analysis of the mouse brain and its major brain regions and cell types. Comparisons of the 12,934 identified proteins in oligodendrocytes, astrocytes, microglia and cortical neurons with deep sequencing data of the transcriptome indicated deep coverage of the proteome. Cell type-specific proteins defined as tenfold more abundant than average expression represented about a tenth of the proteome, with an overrepresentation of cell surface proteins. To demonstrate the utility of our resource, we focused on this class of proteins and identified Lsamp, an adhesion molecule of the IgLON family, as a negative regulator of myelination. Our findings provide a framework for a system-level understanding of cell-type diversity in the CNS and serves as a rich resource for analyses of brain development and function.

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Crossref · 10.1038/nn.4160

Cites

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Nothing to recompute.

Claims the paper makes

31 of 48 cited sources checked · 13 supported · 0 flagged · accuracy not yet measured

  • Premisecitation

    High-resolution mass spectrometry technologies now allow in-depth coverage of nearly complete proteomes of mammalian cells.

    “this situation has changed with the development of more sensitive and powerful high-resolution mass spectrometry (MS) technologies that allow in-depth coverage of nearly complete proteomes of mammalian cells 13–17.”
    Quote in the paperSubstantial claim2 of 5 sources checked
    • cites [13] Beck, M., Claassen, M. & Aebersold, R. Comprehensive proteomics. Curr. Opin. Biotechnol. 22, 3–8 (2011). source
    • cites [14] Mann, M., Kulak, N.A., Nagaraj, N. & Cox, J. The coming age of complete, accurate, and ubiquitous proteomes. Mol. Cell 49, 583–590 (2013). source
    • cites [15] Nagaraj, N. et al. Deep proteome and transcriptome mapping of a human cancer cell line. Mol. Syst. Biol. 7, 548 (2011). source
    • cites [16] Wilhelm, M. et al. Mass-spectrometry-based draft of the human proteome. Nature 509, 582–587 (2014). source
    • cites [17] Kim, M.S. et al. A draft map of the human proteome. Nature 509, 575–581 (2014). source

      The Abstract and Discussion report that high-resolution mass spectrometry identified proteins corresponding to about 84% of human protein-coding genes, supporting in-depth coverage of the human proteome. But that coverage was pooled across 30 tissue and cell types; the Discussion describes sampling individual cell types more extensively as future work. The paper therefore does not establish nearly complete coverage of individual mammalian-cell proteomes. (LLM, conf=0.84)

      Experimental — accuracy not yet measured
  • Premisecitation

    A concerted effort to resolve the brain proteome is missing.

    “Thus, protein expression analyses are required, but a concerted effort to resolve the brain proteome is missing12.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [12] Kitchen, R.R., Rozowsky, J.S., Gerstein, M.B. & Nairn, A.C. Decoding neuroproteomics: integrating the genome, translatome and functional anatomy. Nat. Neurosci. 17, 1491–1499 (2014). source

      In “Improving the compatibility of proteomics with functional genomics” (page unavailable), the paper says brain-wide protein-localisation and abundance data are far less mature than RNA data and calls for a large-scale quantitative resource. That supports a gap in resolving the brain proteome, but not the stronger assertion that a concerted effort is missing: the same section names the HUPO Brain Proteome Project, and “Challenges in assessing the proteome(s) of the CNS” describes existing studies. (LLM, conf=0.83)

      Experimental — accuracy not yet measured
  • Findingown data

    After excluding low-abundance transcripts, the number of genes was reduced to 12,933.

    “When we excluded these transcripts from our analysis, a standard procedure in RNA-Seq analysis28, the number of genes was reduced to 12,933 (Fig. 1f).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [28] Hebenstreit, D. et al. RNA sequencing reveals two major classes of gene expression levels in metazoan cells. Mol. Syst. Biol. 7, 497 (2011). source

      The citation is attached to the description of excluding low-abundance transcripts as a standard RNA-seq procedure, not to the citing authors’ count of 12,933 genes. The Results and discussion section describes detection limits at low expression levels, but the passages found do not say that excluding those transcripts is standard practice (searched for filtering and exclusion of low-expression genes). (LLM, conf=0.72)

      Experimental — accuracy not yet measured
  • Premisecitation

    iBAQ values yield protein copy number estimates of each protein in the proteome.

    “These iBAQ values yield protein copy number estimates of each protein in the proteome33.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [33] Wiśśniewski, J.R. et al. Extensive quantitative remodeling of the proteome between normal colon tissue and adenocarcinoma. Mol. Syst. Biol. 8, 611 (2012). source

      The Results section (“Estimation of absolute copy number of proteins”; page unavailable) says the paper estimates protein copy numbers using LFQ intensities and the total protein approach (TPA). It reports that iBAQ gave similar values in a validation calculation, but does not say that iBAQ values themselves yield copy-number estimates for each protein in the proteome. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
  • Interpretationreasoning

    The data indicate great depth of coverage of the proteome of glia and neurons.

    “Taken together with previous estimates of the size of cellular proteomes14,15,29, our data indicate that we have achieved a great depth of coverage of the proteome of glia and neurons.”
    Quote in the paperSubstantial claim2 of 3 sources checked
    • cites [14] Mann, M., Kulak, N.A., Nagaraj, N. & Cox, J. The coming age of complete, accurate, and ubiquitous proteomes. Mol. Cell 49, 583–590 (2013). source
    • cites [15] Nagaraj, N. et al. Deep proteome and transcriptome mapping of a human cancer cell line. Mol. Syst. Biol. 7, 548 (2011). source
    • cites [29] Beck, M. et al. The quantitative proteome of a human cell line. Mol. Syst. Biol. 7, 549 (2011). source
  • May combine two claims. 35
  • Classified as probably not a substantial claim. 28

  • May combine two claims.

    Premisecitation

    Messenger RNA and protein quantities often correlate poorly, emphasizing post-transcriptional control of protein synthesis and degradation.

    “Several studies have reported that quantities of messenger RNA and proteins often correlate poorly, emphasizing the importance of post-transcriptional processes that control protein synthesis and degradation11.”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [11] Vogel, C. & Marcotte, E.M. Insights into the regulation of protein abundance from proteomic and transcriptomic analyses. Nat. Rev. Genet. 13, 227–232 (2012). source

  • Classified as probably not a substantial claim.

    Premisecitation

    Previous studies of human cancer cell lines found proteins tend to be ubiquitously expressed, with differences in expression level rather than presence or absence.

    “Previous studies of different human cancer cell lines have found that proteins tend to be ubiquitously expressed with differences in the level of expression of the proteins rather than in their absence or presence18–20.”
    Quote in the paperSubstantial claim3 of 3 sources checked
    • cites [18] Geiger, T., Wehner, A., Schaab, C., Cox, J. & Mann, M. Comparative proteomic analysis of eleven common cell lines reveals ubiquitous but varying expression of most proteins. Mol. Cell Proteomics 11, M111.014050 (2012). source
    • cites [19] Lundberg, E. et al. Defining the transcriptome and proteome in three functionally different human cell lines. Mol. Syst. Biol. 6, 450 (2010). source
    • cites [20] Pontén, F. et al. A global view of protein expression in human cells, tissues, and organs. Mol. Syst. Biol. 5, 337 (2009). source

      The Results sections on 45 human cell lines and confocal microscopy report that most analyzed proteins were detected across cell lines, while 66% differed in expression level among the three lines examined by confocal microscopy. This supports the expression-level-versus-presence distinction, but our copy describes the lines as human cell lines without establishing the claim’s specific characterization of them as cancer cell lines. (LLM, conf=0.82)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Astrocytes provide nutrients, maintain extracellular ion balance, recycle neurotransmitters, shape synaptic circuits and regulate the blood-brain barrier.

    “Astrocytes, in contrast, provide nutrients to the nervous tissue, maintain the extracellular ion balance, recycle neurotransmitters, shape synaptic circuits and regulate the blood-brain barrier24.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [24] Molofsky, A.V. et al. Astrocytes and disease: a neurodevelopmental perspective. Genes Dev. 26, 891–907 (2012). source

      “Astrocyte function during normal development” says astrocytes maintain extracellular homeostasis, and “Maturing astrocytes promote synaptogenesis in developing postnatal brains” describes their role in synapse formation. “Other possible developmental functions of astrocytes” says they regulate blood–brain barrier permeability. The passages do not establish the more specific claims that astrocytes provide nutrients or recycle neurotransmitters. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Low abundance transcripts appear to result from rare stochastic transcription and are thought to often be non-functional mRNAs.

    “These low abundance transcripts appear to be the result of rare stochastic transcription and are thought to often result in non-functional mRNAs28.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [28] Hebenstreit, D. et al. RNA sequencing reveals two major classes of gene expression levels in metazoan cells. Mol. Syst. Biol. 7, 497 (2011). source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    A relatively small number of proteins make up a high proportion of total protein mass in ultra-deep proteome analysis.

    “As previously observed in ultra-deep proteome analysis, a relatively small number of proteins make up for a high proportion of the total protein mass15,34.”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [15] Nagaraj, N. et al. Deep proteome and transcriptome mapping of a human cancer cell line. Mol. Syst. Biol. 7, 548 (2011). source
    • cites [34] Marguerat, S. et al. Quantitative analysis of fission yeast transcriptomes and proteomes in proliferating and quiescent cells. Cell 151, 671–683 (2012). source

      In “Coordinated Expression at mRNA and Protein Levels” (page not provided), the paper reports that the 20% most abundant proteins account for 81.3% of protein molecules in proliferating cells. This supports the unequal-abundance point, but the reported proportion is based on molecule counts, not total protein mass. (LLM, conf=0.85)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Findingown data

    The non-filtered RNA-Seq data showed a bimodal distribution, with one third of transcripts having abundances lower than 1 RPKM.

    “As observed previously, the abundance of the non-filtered data, expressed as reads per kilobase of exon per million fragments mapped (RPKM), revealed a bimodal distribution (Fig. 1e), where one third of the transcripts have abundances lower than 1 RPKM15,28.”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [15] Nagaraj, N. et al. Deep proteome and transcriptome mapping of a human cancer cell line. Mol. Syst. Biol. 7, 548 (2011). source
    • cites [28] Hebenstreit, D. et al. RNA sequencing reveals two major classes of gene expression levels in metazoan cells. Mol. Syst. Biol. 7, 497 (2011). source

      The Results and discussion describe two overlapping expression groups in RNA-seq data, but the RNA-seq distribution has a low-expression shoulder; the paper calls the microarray distribution clearly bimodal. Our copy does not report that one third of transcripts have abundances below 1 RPKM. Its figure of approximately 30% refers instead to genes with no detected reads at a sequencing depth of 25 million reads. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Findingown data

    The correlation between proteomes and corresponding transcriptomes was only 0.4 to 0.45, capturing important differences between transcriptome and proteome.

    “As the correlation between the proteomes and the corresponding transcriptomes was only 0.4 to 0.45,our analysis captured important differences between transcriptome and proteome30,31 (Fig. 2c).”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [30] de Sousa Abreu, R., Penalva, L.O., Marcotte, E.M. & Vogel, C. Global signatures of protein and mRNA expression levels. Mol. Biosyst. 5, 1512–1526 (2009). source

      The section ‘Steady-state concentrations of protein molecules per mRNA and their correlates’ (page not provided) reports that several measurements have R² around 0.4 and that much of the variation in protein levels is not explained by mRNA levels. It does not give 0.4–0.45 as a general correlation coefficient: that figure is approximately a squared Pearson correlation, and the reported values vary across organisms. The citing authors’ statement about what their own analysis captured is their inference, not a finding attributed to this source. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [31] Maier, T., Guell, M. & Serrano, L. Correlation of mRNA and protein in complex biological samples. FEBS Lett. 583, 3966–3973 (2009). source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Findingown data

    Transfection of full-length Lsamp, Neurotrimin and Opcml tagged with EGFP into HEK293T cells and addition of soluble Fc fusion proteins showed homo- and heterophilic interactions as noted previously.

    “Next, we transfected plasmids encoding for full-length Lsamp, Neurotrimin and Opcml tagged with EGFP into HEK293T cells, added soluble Fc fusion proteins and observed homo- and heterophilic interactions as noted previously41 (Supplementary Fig. 10).”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [41] Gil, O.D., Zanazzi, G., Struyk, A.F. & Salzer, J.L. Neurotrimin mediates bifunctional effects on neurite outgrowth via homophilic and heterophilic interactions. J. Neurosci. 18, 9312–9325 (1998). source

      The section “Ntm forms homomeric complexes: potential role in promoting adhesion” reports homophilic binding between neurotrimin-expressing CHO cells and soluble Ntm-Fc. The paper also suggests heterophilic mechanisms in its neurite-outgrowth experiments, but does not establish the homo- and heterophilic interactions among Lsamp, Neurotrimin, and Opcml implied here. The HEK293T/EGFP experiment is the citing authors’ own work, not an experiment attributed to this source. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Lsamp belongs to the IgLON family of adhesion molecules, which also comprises Ntm, Opcml and Negr1.

    “Lsamp belongs to the IgLON family of adhesion molecules, which also comprises Ntm (neurotrimin), Opcml (opiod-binding cell adhesion molecule) and Negr1 (neuronal growth regulator1)37–39.”
    Quote in the paperNot a substantial claim1 of 3 sources checked
    • cites [37] Pimenta, A.F. et al. The limbic system-associated membrane protein is an Ig superfamily member that mediates selective neuronal growth and axon targeting. Neuron 15, 287–297 (1995). source
    • cites [38] Schofield, P.R. et al. Molecular characterization of a new immunoglobulin superfamily protein with potential roles in opioid binding and cell contact. EMBO J. 8, 489–495 (1989). source
    • cites [39] Struyk, A.F. et al. Cloning of neurotrimin defines a new subfamily of differentially expressed neural cell adhesion molecules. J. Neurosci. 15, 2141–2156 (1995). source

      The Discussion (page unavailable in our copy) identifies LAMP, neurotrimin and OBCAM as members of a subfamily of Ig-like cell adhesion molecules, supporting the Lsamp, Ntm and Opcml portion of the claim. It does not identify Negr1 as a member or use the name IgLON. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Mutations in the Col11A1 gene are associated with a type of Stickler syndrome, a connective tissue disorder characterized by ocular, skeletal, orofacial and auditory defects.

    “Mutations in the gene are associated with a type of Stickler syndrome, a connective tissue disorder characterized by ocular, skeletal, orofacial and auditory defects35.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [35] Snead, M.P. & Yates, J.R. Clinical and molecular genetics of Stickler syndrome. J. Med. Genet. 36, 353–359 (1999). source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Findingcitation

    Necl1-Fc and Necl4-Fc bound to oligodendrocytes and axons, respectively, as expected.

    “As positive controls, we used Necl1-Fc and Necl4-Fc42,43, which, as expected, bound to oligodendrocytes and axons, respectively.”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [42] Spiegel, I. et al. A central role for Necl4 (SynCAM4) in Schwann cell-axon interaction and myelination. Nat. Neurosci. 10, 861–869 (2007). source
    • cites [43] Maurel, P. et al. Nectin-like proteins mediate axon Schwann cell interactions along the internode and are essential for myelination. J. Cell Biol. 178, 861–874 (2007). source

      The Results section, “Necl proteins mediate adhesion to neurons and Schwann cells” (page not provided), reports Necl-4-Fc binding to DRG neurites, supporting the axon portion. It reports Necl-1-Fc binding to Schwann cells, not oligodendrocytes; the oligodendrocyte portion is not established by this paper. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Ncam1 negatively regulates myelination.

    “Ncam1 (Fig. 6b),a protein that negatively regulates myelination36.”
    • Also stated in: § Discussion · plain
      “Ncam1, a protein that negatively regulates myelination36.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [36] Charles, P. et al. Negative regulation of central nervous system myelination by polysialylated-neural cell adhesion molecule. Proc. Natl. Acad. Sci. USA 97, 7585–7590 (2000). source

      The Discussion reports that PSA-NCAM acts as a negative signal for myelination, and the Results show increased myelination after masking or removing its PSA moieties. The citing sentence attributes this effect to Ncam1 generally, but the paper establishes it specifically for polysialylated NCAM, not NCAM regardless of its polysialylation state. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Findingcitation

    The identified 17,634 transcripts from RNA sequencing showed a bimodal distribution, with one-third of transcripts of low abundance, possibly due to stochastic transcription.

    “The identified 17,634 transcripts from the RNA sequencing showed a bimodal distribution, where one-third of the transcripts were of low abundance, possibly as a result of stochastic transcription28.”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [28] Hebenstreit, D. et al. RNA sequencing reveals two major classes of gene expression levels in metazoan cells. Mol. Syst. Biol. 7, 497 (2011). source

      The Results and discussion describe two expression classes and an emerging low-expression peak after correcting for RNA-seq detection limits. They say the microarray distribution is clearly bimodal, however, while the uncorrected RNA-seq distribution has a low-expression shoulder. The passages do not substantiate 17,634 identified transcripts, a one-third low-abundance fraction, or stochastic transcription as the explanation for that fraction. (LLM, conf=0.87)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Findingown data

    The study observed proteome-transcriptome Pearson correlation coefficients between 0.40 and 0.45, poorer than previously reported for cell lines but in the range of recent human tissue comparisons.

    “We observed differences in the proteome and transcriptome with Pearson correlation coefficients between 0.40 and 0.45, somewhat poorer than those previously reported for cell lines15, but in the range of recent comparisons of human tissues16.”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [15] Nagaraj, N. et al. Deep proteome and transcriptome mapping of a human cancer cell line. Mol. Syst. Biol. 7, 548 (2011). source
    • cites [16] Wilhelm, M. et al. Mass-spectrometry-based draft of the human proteome. Nature 509, 582–587 (2014). source

  • May combine two claims.

    Findingown data

    Oligodendrocytes expressed the smallest number (341 proteins) of cell type-specific proteins, and cortical neurons the largest (669 proteins).

    “We found that oligodendrocytes expressed the smallest number (341 proteins) of cell type-specific proteins, defined by the greater than tenfold fold enrichment criterion, and cortical neurons the largest (669 proteins).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Between different cell types, proteome correlations ranged from 0.48 to 0.84 and transcriptome correlations from 0.25 to 0.86.

    “Between different cell types, correlations were much lower, from 0.48 to 0.84 for the proteome (Fig. 2a) and from 0.25 to 0.86 for the transcriptome (Supplementary Fig. 3).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    This corresponds to an average of 11,970 proteins per system, with 10,529 proteins identified in all five cell types and brain.

    “This corresponds to an average of 11,970 different proteins in each of these systems, of which 10,529 proteins were identified in all five cell types and brain (Fig. 1b,c).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Cell surface proteins were highly overrepresented among the most abundant brain-versus-liver-enriched proteins, and four out of the top five GOCC terms were related to membranes.

    “Similar to the comparison between the different cell types, GO-based enrichment analysis revealed that cell surface proteins were highly overrepresented in the group of the most abundant brain-versus liver-enriched proteins and four out of the top five GOCC terms were related to membranes (Supplementary Table 16).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Microglia-specific proteins had functions in membrane trafficking, lipid catabolism and cell surface signaling, whereas astrocyte-specific proteins had functions in actin-related processes and cilia.

    “The cell type-specific proteins in microglia had functions in membrane trafficking (Rab32, Myo1f), lipid catabolism (Apobr, Lipa) and signaling at the cell surface (CD180, Dock2, Nckap1l,Gpnmb, Fermt3), whereas proteins with designated functions in actin-related processes (Cnn1, Fmn2, Nebl, Pdlim7, Synpo2) and cilia (Rsph1, Ccdc39) were enriched in astrocytes.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In all four cell types, ∼30-40 of the most abundant proteins accounted for ∼25% of total protein molecules, and some were cell type specific.

    “In fact, in all four cell types, ∼ 30–40 of the most abundant proteins accounted for ∼25% of the total protein molecules, and some of these were cell type specific (for oligodendrocytes Mbp, Plp, Cnp1; for cortical neurons Atp1b1, Crmp1, Gpm6a; for astrocytes Gfap, Gstm1, Prdx6; and for microglia Gm5483; Fig. 3b).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The large majority of proteins were expressed in all four cell types, and only about 15% of quantified proteins were cell type specific (>10-fold more abundant in one cell compared with all others).

    “The large majority of proteins were expressed in all four cell types, whereas only about 15% of the quantified proteins could be considered to be cell type specific (>10-fold more abundant in one cell compared with all others; Supplementary Table 6).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The top 50 cell type-specific proteins included many well-established markers and, based on PubMed search, more than one-third of the 50 proteins in various glial types had not previously been studied in the respective cell type.

    “The top 50 cell type-specific proteins (the most enriched and abundant proteins) included many well-established cell type-specific markers, but, based on PubMed search, also a relatively large number of proteins that have previously not been studied in the context of the respective cell type (more than one-third of the 50 proteins in the various glial types).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    The 12,934 proteins identified in different CNS cell types correspond to 11,690 ENSEMBL genes, representing about 60% of all protein coding genes in the genome.

    “The 12,934 proteins identified in different CNS cell types correspond to 11,690 ENSEMBL genes, which represents ∽60% of all protein coding genes in the genome.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingreasoning

    Microglia were the most diverging cell type, and astrocytes and oligodendrocytes were closest to each other.

    “Consistent with their hematopoietic origin, microglia were the most diverging cell type; and, as expected from their shared lineage, astrocytes and oligodendrocytes were closest to each other (Fig. 2b).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    PCA and hierarchical clustering showed that the optic nerve, cerebellum and brain stem were the most diverging brain regions, while the remaining seven regions clustered more tightly.

    “As judged by principal component analysis (PCA) and hierarchical clustering, the optic nerve, the cerebellum and the brain stem were the most diverging brain regions, whereas the remaining seven regions clustered more tightly (Fig. 5a,b).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Necl1-Fc promoted growth and spreading of oligodendrocytes, whereas cells did not attach and grow on Lsamp-Fc coated coverslips.

    “Whereas Necl1-Fc, used as a positive control, promoted growth and spreading of oligodendrocytes, cells did not attach and grow on Lsamp-Fc coated coverslips (Fig. 8f,g).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Antibody staining confirmed expression of Lsamp in oligodendrocytes and neurons, but not in astrocytes or microglia.

    “Using antibody staining, we confirmed the expression of Lsamp in oligodendrocytes and neurons, but not in astrocytes or microglia (Fig. 7e,f).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    In total, 2,901 brain region-enriched proteins (>4-fold difference) were identified, with the thalamus expressing the smallest and the optic nerve the largest numbers (154 and 1,179, respectively).

    “In total, we identified 2,901 brain region-enriched proteins (>4-fold difference) and found that the thalamus expressed the smallest and the optic nerve the largest numbers of brain region-enriched proteins (154 and 1,179, respectively; Supplementary Tables 17 and 18).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Immunofluorescence staining showed that Col11A1 can be used as a new marker for immature oligodendrocytes, and immunohistochemistry revealed a weak association of Col11A1 with white matter tracts.

    “Immunofluorescence staining of cultured cells showed that Col11A1 can be used as a new marker for immature oligodendrocytes and immunohistochemistry of brain sections revealed a weak association of Col11A1 with white matter tracts (Supplementary Fig. 8).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Proteins of the extracellular matrix were identified, with Col11A1 being the most highly expressed and enriched protein.

    “These analyses identified proteins of the extracellular matrix (Col11A1, Col1A1, Col1A2, Col4A1, Col3A1, Col5A2), of which Col11A1 was the most highly expressed and enriched protein.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Findingown data

    Most IgLON family members were enriched in neurons and oligodendrocytes, except Ntm, which was only enriched in neurons.

    “Notably, most family members were enriched in neurons and oligodendrocytes (except of Ntm, which was only enriched in neurons; Fig. 6c).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    Prph, Sncg and Serpinb1b were most abundant and enriched in the brain stem; Omp and S100a5 in the olfactory bulb; Ppp1r1b, Actn2 and Pde10a in the striatum; and Plcb4, Prkcd and Synpo2 in the thalamus.

    “Prph, Sncg and Serpinb1b were most abundant and enriched in the brain stem, Omp and S100a5 in the olfactory bulb, Ppp1r1b, Actn2 and Pde10a in the striatum, and Plcb4, Prkcd and Synpo2 in the thalamus (Fig. 5c).”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Microglia are glial cells with functions mainly related to immune response and maintaining brain homeostasis.

    “Microglia cells constitute the third class of glial cells with functions mainly related to immune response and maintaining brain homeostasis25.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [25] Hanisch, U.K. & Kettenmann, H. Microglia: active sensor and versatile effector cells in the normal and pathologic brain. Nat. Neurosci. 10, 1387–1394 (2007). source

  • May combine two claims.

    Premisecitation

    Glia constitute 50-90% of cells in the human brain and are highly specialized toward cell type-selective tasks.

    “Glia are not only abundant (constituting 50–90% of the cells in the human brain), but are also highly specialized toward cell type-selective tasks, and therefore ideally suited for these analyses21.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [21] Barres, B.A. The mystery and magic of glia: a perspective on their roles in health and disease. Neuron 60, 430–440 (2008). source

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Oligodendrocytes primarily synthesize membrane and insulate axons by enwrapping them with a multilamellar myelin sheath.

    “Oligodendrocytes are primarily engaged in membrane synthesis, as their function is to insulate axons by enwrapping them with a multilamellar myelin sheath22,23.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [22] Sherman, D.L. & Brophy, P.J. Mechanisms of axon ensheathment and myelin growth. Nat. Rev. Neurosci. 6, 683–690 (2005). source
    • cites [23] Aggarwal, S., Yurlova, L. & Simons, M. Central nervous system myelin: structure, synthesis and assembly. Trends Cell Biol. 21, 585–593 (2011). source

  • May combine two claims.

    Study design and conductown data

    Between biological replicates of the same cell types, Pearson's correlation coefficients were about 0.96, demonstrating excellent technical and biological reproducibility.

    “Between biological replicates of the same cell types, Pearson’s correlation coefficients were about 0.96, demonstrating excellent technical and biological reproducibility (Fig. 2a).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductown data

    Cell type-specific markers for astrocytes, oligodendrocytes, microglia and neuronal progenitors were reproducibly enriched at least 16-fold, confirming proteome quality.

    “The expression levels of cell type-specific markers for astrocytes (Aqp4, Gfap, Aldh1l1), oligodendrocytes (Cnp1, Mbp, Plp1), microglia (Iba1, Tlr2, Tlr7) and neuronal progenitors (Bcl11a, Bcl11b, Ebf3) showed that almost all of these markers were reproducibly enriched at least 16-fold, also confirming the quality of the proteome analysis (Supplementary Fig. 6a).”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductown data

    Expression levels of well-described markers for astrocytes, oligodendrocytes, cortical neurons, cerebellar granule neurons and microglia were reproducibly enriched at least 16-fold, confirming proteome quality.

    “The expression levels of well-described markers for astrocytes (Aquaporin 4, Gfap, Glast and Aldh1l1), oligodendrocytes (Cnp1, Sox10, Mbp, Mag, Mog, Plp1), cortical neurons (neurofilament, Snap25, synaptotagmin I, Gabra 2), cerebellar granule neurons (Synaptotagmin I, Snap25, Sv2b, Gprin1, L1cam) and microglia (Iba1, Tlr2, Tlr7, CD86) in our triplicate proteome analysis revealed that almost all of”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Study design and conductown data

    The deep primary cell proteomes faithfully reproduce known cellular functions and markers, providing a solid foundation for global characterization and new discoveries.

    “Together these analyses of our deep primary cell proteomes faithfully reproduce known and characteristic cellular functions as well as their markers, providing a solid foundation for their global characterization and for new discoveries.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisenone given

    All four IgLONs are GPI-anchored glycoproteins with three Ig domains implicated in cell adhesion and growth guidance in neurons, but not in glia.

    “All four IgLONs are glycosylphosphati-dylinositol (GPI)-anchored glycoproteins with three Ig domains that have been implicated in cell adhesion and growth guidance in neurons, but not in glia.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data

    The analysis filtered for 8,147 protein groups that were quantified in all replicates for at least one cell type or its differentiation stage.

    “We filtered for 8,147 protein groups that were quantified in all replicates for at least one cell type or its differentiation stage.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    All statistical analyses were performed on at least three different biological replicates.

    “All statistical analyses were performed on at least three different biological replicates.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S2 (checkable).

    Interpretationown data

    MS-based proteomics can be implemented more broadly into neuroscience to help resolve the full molecular complexity of the brain at the protein level, with cellular or even subcellular resolution.

    “our data and the rapidity with which it can now be generated, suggest that MS-based proteomics can be implemented more broadly into neuroscience to help resolve the full molecular complexity of the brain at the protein level, with cellular or even subcellular resolution.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    The one-dimensional annotation matrix algorithm used a two-sided Wilcoxon-Mann-Whitney test with Benjamini-Hochberg FDR threshold of 0.02 to adjust for multiple hypothesis testing.

    “it tests and computes the difference of every protein annotation from the overall intensity distribution using the two-sided Wilcoxon-Mann-Whitney test where the multiple hypothesis testing is adjusted by applying a Benjamini-Hochberg FDR threshold of 0.02.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Differentially expressed proteins were identified by an ANOVA based test at permutation-based FDR cutoff of 0.05.

    “Differentially expressed proteins were identified by an ANOVA based test at permutation-based FDR cutoff of 0.05.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Protein identification required at least one unique or razor peptide per protein group.

    “Protein identification required at least one unique or razor peptide per protein group.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data

    Two-sample Welch t-tests with a 2% permutation-based FDR filter were used to identify proteins with significant differential expression in a cell type compared to others.

    “We performed two-sample welch t-tests to identify proteins with a significant differential expression in a cell type in comparison to others employing a 2% permutation-based FDR filter.”
    Quote in the paperSubstantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    Only cell cultures of at least 95% purity were included in further experiments.

    “only cultures of at least 95% purity were included in further experiments.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Blinded quantification was performed by determining areas of the myelinated and unmyelinated axons.

    “Blinded quantification was performed by determining areas of the myelinated and unmyelinated axons.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Study design and conductnone given

    The sample size was similar or higher than those generally employed in the field.

    “The sample size was similar or higher to those generally employed in the field.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    PLL coating was used as a positive control to verify the quality of the primary oligodendrocyte preparation.

    “PLL coating (100 μg ml-1) was used as a positive control to verify the quality of the primary oligodendrocyte preparation.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductown data

    The annotation matrix ANOVA used a permutation-based FDR cutoff of 0.001 to identify annotation terms significantly different between at least two groups.

    “The replicates were grouped based on cell type and differentiation stage before applying an ANOVA test on the annotation matrix with a permutation-based FDR cutoff of 0.001 to identify annotation terms that are significantly different between at least two groups.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S2 (checkable).

    Interpretationown data

    These technological advances can be used broadly in the proteomic study of diverse and rare brain cell types.

    “We believe that these technological advances can be used broadly in the proteomic study of diverse and rare brain cell types.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationown data

    The results identify a previously unknown negative regulator of myelination in specific fiber tracts of the CNS and exemplify the utility of the proteome resource.

    “Apart from identifying a previously unknown negative regulator of myelination in specific fiber tracts of the CNS, these results exemplify the utility of our proteome resource.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Commonly used filtering criteria for RNA-Seq reduced the number to 12,933, indicating that proteome coverage is close to completion, at least in terms of detection of each functionally expressed gene.

    “Commonly used filtering criteria forRNA-Seq reduced the number to 12,933, indicating that our proteome coverage is close to completion, at least in terms of detection of each functionally expressed gene.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    The required false discovery rate was set to 1% at the peptide and protein level.

    “the required FDR was set to 1% at the peptide and protein level.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Lsamp mediates homophilic and heterophilic interactions with other IgLON family members.

    “Lsamp is known to mediate homophilic, as well as heterophilic, interactions with other members of the IgLON family48.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [48] Gil, O.D. et al. Complementary expression and heterophilic interactions between IgLON family members neurotrimin and LAMP. J. Neurobiol. 51, 190–204 (2002). source

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Study design and conductnone given

    Animal experiments were performed according to the Lower Saxony State regulations for animal experimentation.

    “All animal experiments were performed according to the Lower Saxony State regulations for animal experimentation.”
    Quote in the paperNot a substantial claim
58 of 58

  • The paper relies on this without support.

    How proteomes differ among specialized cells in tissue is not known.

    “Proteins are the main functional components in all cells, but how the proteomes differ in specialized cells in tissue is not known.”

  • The paper relies on this without support.

    The molecular nature of axon-glia interaction is unknown.

    “Given that the molecularnature of axon and glia interaction is unknown”

  • The authors state this as an assumption.

    Data distribution was assumed to be normal but not formally tested.

    “Data distribution was assumed to be normal but this was not formally tested.”
3 of 3

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Protein expression data of isolated mouse brain cells was generated by Sharma and colleagues using MACS to isolate microglia, astrocytes, oligodendrocytes, and neurons from young (P8) C57/BL6 mice (3 biological replicates), analysed by mass spectrometry.

    “Protein expression data of isolated mouse brain cells was generated by Sharma and colleagues [14] using Magnetic-Activated Cell Sorting (MACS) to isolate microglia, astrocytes, oligodendrocytes, and neurons from the brains of young (P8) C57/BL6 mice (3 biological replicates), analysed by mass spectrometry.”

    From Selective suppression of oligodendrocyte-derived amyloid beta rescues neuronal dysfunction in Alzheimer’s disease (2024) · cites this paper as [14]

    The Results section, “Proteome analysis of acutely isolated CNS cell types” (page unavailable), confirms magnetic sorting of microglia, astrocytes, oligodendrocytes and PSA-NCAM+ neuronal progenitors from P8 mice, with three biological replicates and quantified proteins. The claim calls the fourth population neurons rather than neuronal progenitors. The Online Methods identify C57BL/6 mice for the separate P60 brain-region analysis but do not specify the strain of the P8 mice. (LLM, conf=0.88)

    Experimental — accuracy not yet measured

  • BCAS1 was one of the most enriched proteins in oligodendrocytes in the previously generated proteome database.

    “We examined our previously generated proteome database for oligodendrocyte specific proteins and noticed that BCAS1 was one of the most enriched proteins in oligodendrocytes (21).”

    From BCAS1 expression defines a population of early myelinating oligodendrocytes in multiple sclerosis lesions (2017) · cites this paper as [21]

    The Results section describes a cell-type-resolved proteome and ranks proteins by enrichment in oligodendrocytes, supporting the identification of the previously generated database. But the passages naming enriched proteins do not mention BCAS1, and searches for BCAS1 and its expanded name found no passage establishing that it was among the most enriched. (LLM, conf=0.78)

    Experimental — accuracy not yet measured

  • The transcriptome is often not an accurate indicator of protein abundance, especially in complex brain tissue.

    “The transcriptome, however, is often not an accurate indicator of protein abundance (Liu et al., 2016), especially in complex brain tissue (Sharma et al., 2015).”

    From Deep Multilayer Brain Proteomics Identifies Molecular Networks in Alzheimer’s Disease Progression (2020) · cites this paper as Sharma et al., 2015

  • At the protein level, oligodendrocytes contain high levels of APP, BACE1, and PSEN1.

    “we exploited publicly available proteomics data derived from isolated mouse brain cells [14] and found that, at the protein level, oligodendrocytes indeed contain high levels of APP, BACE1, and PSEN1 (S1 Fig).”

    From Selective suppression of oligodendrocyte-derived amyloid beta rescues neuronal dysfunction in Alzheimer’s disease (2024) · cites this paper as [14]

  • mRNA-protein correlation coefficients reach no higher than 0.47 even in acutely isolated brain cell types.

    “mRNA-protein correlation coefficients reach no higher than 0.47 even in acutely isolated brain cell types (Sharma et al., 2015).”

    From A Multi-network Approach Identifies Protein-Specific Co-expression in Asymptomatic and Symptomatic Alzheimer’s Disease (2017) · cites this paper as Sharma et al., 2015

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Different starting counts for the RNA-seq data

    Minor

    The Results give 19,501 expressed gene symbols before filtering, while the Discussion gives 17,634 identified transcripts for the RNA-seq analysis before describing the same filtered total of 12,933. The paper does not explain why these starting counts differ.

  • Conflicting final harvest day for cortical neurons

    Minor

    The Results say cortical neurons were analyzed at 5, 10 and 15 days in vitro, but the culture protocol says they were cultivated until days 5, 10 and 16.

2 of 2

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Authors
Sharma, K., Schmitt, S., Bergner, C. G., Tyanova, S., Kannaiyan, N., Manrique-Hoyos, N., Kongi, K., Cantuti, L., Hanisch, U.-K., Philips, M.-A., Rossner, M. J., Mann, M., & Simons, M.
Affiliations
  • Max Planck Institute of Biochemistry (DE)
  • Max Planck Institute of Experimental Medicine (DE)
  • Ludwig-Maximilians-Universität München (DE)
  • University of Tartu (EE)
  • University of Göttingen (DE)
Corresponding author
  • Mann, Matthias
  • Simons, Mikael
ORCID
Venue
Nature Neuroscience
Publisher
Springer Science and Business Media LLC
Published
2015-11-02
Volume
18
Issue
12
Pages
1819-1831
ISSN
  • 1097-6256 (ISSN-L)
  • 1546-1726
DOI
10.1038/nn.4160
PMC
PMC7116867
OpenAlex
W1955394435
Cited by
871
References
61
Citations by year
2026: 41 · 2025: 54 · 2024: 73 · 2023: 84 · 2022: 87 · 2021: 77 · 2020: 92 · 2019: 81 · 2018: 85 · 2017: 65 · 2016: 44 · 2015: 1
FWCI
28.41
Type
journal article
Language
en
Open access
Licence
www.springer.com/tdm (tdm)
Locations
  • Nature Neuroscience (publishedVersion)
  • PubMed (publishedVersion)
  • PubMed Central (submittedVersion)
  • GRO (submittedVersion)
  • MPG.PuRe (Max Planck Society) (submittedVersion)
Topics
  • Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
  • Physical Sciences › Chemistry › Spectroscopy › Mass Spectrometry Techniques and Applications
Keywords
oligodendrocytes, astrocytes, microglia, cortical neurons, cell surface proteins, adhesion molecules, LSAMP, myelination
MeSH terms
Animals, Brain, Humans, Male, Mice, Inbred C57BL, Neurons

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Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-07.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-07.

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