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Definition of a consensus integrin adhesome and its dynamics during adhesion complex assembly and disassembly.

Horton ER, Byron A, Askari JA, Ng DH, Millon-Frémillon A, Robertson J, Koper EJ, Paul NR, Warwood S, Knight D, Humphries JD, Humphries MJ - Nat. Cell Biol. (2015)

Bottom Line: Here, we have integrated several IAC proteomes and generated a 2,412-protein integrin adhesome.Analysis of this data set reveals the functional diversity of proteins in IACs and establishes a consensus adhesome of 60 proteins.The definition of this consensus view of integrin adhesome components provides a resource for the research community.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Cell-Matrix Research, Faculty of Life Sciences, University of Manchester, Manchester M13 9PT, UK.

ABSTRACT
Integrin receptor activation initiates the formation of integrin adhesion complexes (IACs) at the cell membrane that transduce adhesion-dependent signals to control a multitude of cellular functions. Proteomic analyses of isolated IACs have revealed an unanticipated molecular complexity; however, a global view of the consensus composition and dynamics of IACs is lacking. Here, we have integrated several IAC proteomes and generated a 2,412-protein integrin adhesome. Analysis of this data set reveals the functional diversity of proteins in IACs and establishes a consensus adhesome of 60 proteins. The consensus adhesome is likely to represent a core cell adhesion machinery, centred around four axes comprising ILK-PINCH-kindlin, FAK-paxillin, talin-vinculin and α-actinin-zyxin-VASP, and includes underappreciated IAC components such as Rsu-1 and caldesmon. Proteomic quantification of IAC assembly and disassembly detailed the compositional dynamics of the core cell adhesion machinery. The definition of this consensus view of integrin adhesome components provides a resource for the research community.

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Overlap and comparison of IAC proteomes in the meta-adhesome. (a) Pairwise overlaps of FN-enriched proteins identified in the seven proteomic datasets and the literature-curated adhesome4 are displayed as a hierarchically clustered heatmap. K562, human chronic myelogenous leukaemia cells11; MEF, mouse embryonic fibroblast cells (this study); A375, human malignant melanoma cells14; HFF, human foreskin fibroblast cells13; MKF1, mouse kidney fibroblast cells15; MKF2 and MKF3, mouse kidney fibroblast cells16. Details of the proteomic datasets are provided in Supplementary Table 1. (b) The number of proteomic datasets in which proteins in the meta-adhesome are identified (dataset occurrence) is displayed as a pie chart. Numbers of proteins identified are indicated for each segment (proportions of the meta-adhesome are shown in parentheses). (c) Line graph showing the cumulative proportion of the meta-adhesome in at least x proteomic datasets, where x is the minimum (min.) dataset occurrence category. Numbers of proteins identified are indicated for each data point. (d) Protein-protein interaction network model of the meta-adhesome. The 2,412 meta-adhesome proteins were mapped onto a curated database of reported protein-protein interactions. The largest connected graph component is displayed, comprising 11,430 interactions (grey lines; edges) between 2,035 proteins (circles; nodes). Node size and colour are proportional to the number of proteomic datasets in which a protein was identified. Locations of proteins identified in all seven datasets are indicated. (e) Line graph showing the proportion of identified proteins that are in the literature-curated adhesome. Numbers of literature-curated adhesome proteins identified are indicated for each data point. (f) The number of reported protein-protein interactions (degree) for each protein is plotted according to the number of proteomic datasets in which it was identified. Box-and-whisker plot shows the median (line), mean (plus sign), 25th and 75th percentiles (box) and 5th and 95th percentiles (whiskers) (n = 1,117, 518, 238, 102, 33, 25 and 10 mapped proteins identified in 1–7 datasets, respectively, with degree ≥ 1). *P < 0.05, **P < 0.01, ****P < 0.0001; Kruskal–Wallis test with Dunn’s post hoc correction (see Supplementary Table 15 for statistics source data).
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Figure 1: Overlap and comparison of IAC proteomes in the meta-adhesome. (a) Pairwise overlaps of FN-enriched proteins identified in the seven proteomic datasets and the literature-curated adhesome4 are displayed as a hierarchically clustered heatmap. K562, human chronic myelogenous leukaemia cells11; MEF, mouse embryonic fibroblast cells (this study); A375, human malignant melanoma cells14; HFF, human foreskin fibroblast cells13; MKF1, mouse kidney fibroblast cells15; MKF2 and MKF3, mouse kidney fibroblast cells16. Details of the proteomic datasets are provided in Supplementary Table 1. (b) The number of proteomic datasets in which proteins in the meta-adhesome are identified (dataset occurrence) is displayed as a pie chart. Numbers of proteins identified are indicated for each segment (proportions of the meta-adhesome are shown in parentheses). (c) Line graph showing the cumulative proportion of the meta-adhesome in at least x proteomic datasets, where x is the minimum (min.) dataset occurrence category. Numbers of proteins identified are indicated for each data point. (d) Protein-protein interaction network model of the meta-adhesome. The 2,412 meta-adhesome proteins were mapped onto a curated database of reported protein-protein interactions. The largest connected graph component is displayed, comprising 11,430 interactions (grey lines; edges) between 2,035 proteins (circles; nodes). Node size and colour are proportional to the number of proteomic datasets in which a protein was identified. Locations of proteins identified in all seven datasets are indicated. (e) Line graph showing the proportion of identified proteins that are in the literature-curated adhesome. Numbers of literature-curated adhesome proteins identified are indicated for each data point. (f) The number of reported protein-protein interactions (degree) for each protein is plotted according to the number of proteomic datasets in which it was identified. Box-and-whisker plot shows the median (line), mean (plus sign), 25th and 75th percentiles (box) and 5th and 95th percentiles (whiskers) (n = 1,117, 518, 238, 102, 33, 25 and 10 mapped proteins identified in 1–7 datasets, respectively, with degree ≥ 1). *P < 0.05, **P < 0.01, ****P < 0.0001; Kruskal–Wallis test with Dunn’s post hoc correction (see Supplementary Table 15 for statistics source data).

Mentions: Comparative analyses identified cell-type-, negative-control- and biochemical-isolation-methodology-specific variations in IAC composition (Fig. 1a, Supplementary Fig. 1). Individual IAC proteomes contained hundreds of proteins (602 ± 250, mean ± s.d.; range, 314–1,023) and identified up to a third of literature-curated adhesome4 components (20.9 ± 7.1%, mean ± s.d.; range, 9.1–32.3%) (Fig. 1a, Supplementary Fig. 1c). This variation is likely to result from the context under which the IACs were observed20. Over half of the proteins in the meta-adhesome (1,359; 56.3%) were identified uniquely in a single dataset (Fig. 1b). These proteins represent low-abundance or context-specific adhesome components, or those difficult to detect by MS. The number of proteins identified in the meta-adhesome decreased exponentially as the stringency in dataset number increased (Fig. 1b,c). Four hundred and forty-eight proteins were detected in at least three datasets (Fig. 1c), more than the 63 proteins previously found in common between three published IAC proteomes21. Only 10 proteins were enriched in all seven datasets (labelled in Fig. 1d). We hypothesised that a restricted set of robustly detected proteins may represent a context-independent core of IAC components20. Indeed, the proportion of identified proteins that were literature-curated adhesome4 components increased with dataset occurrence (Fig. 1e, Supplementary Fig. 1a), suggesting that robustly detected proteins are more likely to represent canonical adhesion proteins.


Definition of a consensus integrin adhesome and its dynamics during adhesion complex assembly and disassembly.

Horton ER, Byron A, Askari JA, Ng DH, Millon-Frémillon A, Robertson J, Koper EJ, Paul NR, Warwood S, Knight D, Humphries JD, Humphries MJ - Nat. Cell Biol. (2015)

Overlap and comparison of IAC proteomes in the meta-adhesome. (a) Pairwise overlaps of FN-enriched proteins identified in the seven proteomic datasets and the literature-curated adhesome4 are displayed as a hierarchically clustered heatmap. K562, human chronic myelogenous leukaemia cells11; MEF, mouse embryonic fibroblast cells (this study); A375, human malignant melanoma cells14; HFF, human foreskin fibroblast cells13; MKF1, mouse kidney fibroblast cells15; MKF2 and MKF3, mouse kidney fibroblast cells16. Details of the proteomic datasets are provided in Supplementary Table 1. (b) The number of proteomic datasets in which proteins in the meta-adhesome are identified (dataset occurrence) is displayed as a pie chart. Numbers of proteins identified are indicated for each segment (proportions of the meta-adhesome are shown in parentheses). (c) Line graph showing the cumulative proportion of the meta-adhesome in at least x proteomic datasets, where x is the minimum (min.) dataset occurrence category. Numbers of proteins identified are indicated for each data point. (d) Protein-protein interaction network model of the meta-adhesome. The 2,412 meta-adhesome proteins were mapped onto a curated database of reported protein-protein interactions. The largest connected graph component is displayed, comprising 11,430 interactions (grey lines; edges) between 2,035 proteins (circles; nodes). Node size and colour are proportional to the number of proteomic datasets in which a protein was identified. Locations of proteins identified in all seven datasets are indicated. (e) Line graph showing the proportion of identified proteins that are in the literature-curated adhesome. Numbers of literature-curated adhesome proteins identified are indicated for each data point. (f) The number of reported protein-protein interactions (degree) for each protein is plotted according to the number of proteomic datasets in which it was identified. Box-and-whisker plot shows the median (line), mean (plus sign), 25th and 75th percentiles (box) and 5th and 95th percentiles (whiskers) (n = 1,117, 518, 238, 102, 33, 25 and 10 mapped proteins identified in 1–7 datasets, respectively, with degree ≥ 1). *P < 0.05, **P < 0.01, ****P < 0.0001; Kruskal–Wallis test with Dunn’s post hoc correction (see Supplementary Table 15 for statistics source data).
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Related In: Results  -  Collection

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Figure 1: Overlap and comparison of IAC proteomes in the meta-adhesome. (a) Pairwise overlaps of FN-enriched proteins identified in the seven proteomic datasets and the literature-curated adhesome4 are displayed as a hierarchically clustered heatmap. K562, human chronic myelogenous leukaemia cells11; MEF, mouse embryonic fibroblast cells (this study); A375, human malignant melanoma cells14; HFF, human foreskin fibroblast cells13; MKF1, mouse kidney fibroblast cells15; MKF2 and MKF3, mouse kidney fibroblast cells16. Details of the proteomic datasets are provided in Supplementary Table 1. (b) The number of proteomic datasets in which proteins in the meta-adhesome are identified (dataset occurrence) is displayed as a pie chart. Numbers of proteins identified are indicated for each segment (proportions of the meta-adhesome are shown in parentheses). (c) Line graph showing the cumulative proportion of the meta-adhesome in at least x proteomic datasets, where x is the minimum (min.) dataset occurrence category. Numbers of proteins identified are indicated for each data point. (d) Protein-protein interaction network model of the meta-adhesome. The 2,412 meta-adhesome proteins were mapped onto a curated database of reported protein-protein interactions. The largest connected graph component is displayed, comprising 11,430 interactions (grey lines; edges) between 2,035 proteins (circles; nodes). Node size and colour are proportional to the number of proteomic datasets in which a protein was identified. Locations of proteins identified in all seven datasets are indicated. (e) Line graph showing the proportion of identified proteins that are in the literature-curated adhesome. Numbers of literature-curated adhesome proteins identified are indicated for each data point. (f) The number of reported protein-protein interactions (degree) for each protein is plotted according to the number of proteomic datasets in which it was identified. Box-and-whisker plot shows the median (line), mean (plus sign), 25th and 75th percentiles (box) and 5th and 95th percentiles (whiskers) (n = 1,117, 518, 238, 102, 33, 25 and 10 mapped proteins identified in 1–7 datasets, respectively, with degree ≥ 1). *P < 0.05, **P < 0.01, ****P < 0.0001; Kruskal–Wallis test with Dunn’s post hoc correction (see Supplementary Table 15 for statistics source data).
Mentions: Comparative analyses identified cell-type-, negative-control- and biochemical-isolation-methodology-specific variations in IAC composition (Fig. 1a, Supplementary Fig. 1). Individual IAC proteomes contained hundreds of proteins (602 ± 250, mean ± s.d.; range, 314–1,023) and identified up to a third of literature-curated adhesome4 components (20.9 ± 7.1%, mean ± s.d.; range, 9.1–32.3%) (Fig. 1a, Supplementary Fig. 1c). This variation is likely to result from the context under which the IACs were observed20. Over half of the proteins in the meta-adhesome (1,359; 56.3%) were identified uniquely in a single dataset (Fig. 1b). These proteins represent low-abundance or context-specific adhesome components, or those difficult to detect by MS. The number of proteins identified in the meta-adhesome decreased exponentially as the stringency in dataset number increased (Fig. 1b,c). Four hundred and forty-eight proteins were detected in at least three datasets (Fig. 1c), more than the 63 proteins previously found in common between three published IAC proteomes21. Only 10 proteins were enriched in all seven datasets (labelled in Fig. 1d). We hypothesised that a restricted set of robustly detected proteins may represent a context-independent core of IAC components20. Indeed, the proportion of identified proteins that were literature-curated adhesome4 components increased with dataset occurrence (Fig. 1e, Supplementary Fig. 1a), suggesting that robustly detected proteins are more likely to represent canonical adhesion proteins.

Bottom Line: Here, we have integrated several IAC proteomes and generated a 2,412-protein integrin adhesome.Analysis of this data set reveals the functional diversity of proteins in IACs and establishes a consensus adhesome of 60 proteins.The definition of this consensus view of integrin adhesome components provides a resource for the research community.

View Article: PubMed Central - PubMed

Affiliation: Wellcome Trust Centre for Cell-Matrix Research, Faculty of Life Sciences, University of Manchester, Manchester M13 9PT, UK.

ABSTRACT
Integrin receptor activation initiates the formation of integrin adhesion complexes (IACs) at the cell membrane that transduce adhesion-dependent signals to control a multitude of cellular functions. Proteomic analyses of isolated IACs have revealed an unanticipated molecular complexity; however, a global view of the consensus composition and dynamics of IACs is lacking. Here, we have integrated several IAC proteomes and generated a 2,412-protein integrin adhesome. Analysis of this data set reveals the functional diversity of proteins in IACs and establishes a consensus adhesome of 60 proteins. The consensus adhesome is likely to represent a core cell adhesion machinery, centred around four axes comprising ILK-PINCH-kindlin, FAK-paxillin, talin-vinculin and α-actinin-zyxin-VASP, and includes underappreciated IAC components such as Rsu-1 and caldesmon. Proteomic quantification of IAC assembly and disassembly detailed the compositional dynamics of the core cell adhesion machinery. The definition of this consensus view of integrin adhesome components provides a resource for the research community.

Show MeSH
Related in: MedlinePlus