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Panorama of ancient metazoan macromolecular complexes

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ABSTRACT

Macromolecular complexes are essential to conserved biological processes, but their prevalence across animals is unclear. By combining extensive biochemical fractionation with quantitative mass spectrometry, we directly examined the composition of soluble multiprotein complexes among diverse metazoan models. Using an integrative approach, we then generated a draft conservation map consisting of >1 million putative high-confidence co-complex interactions for species with fully sequenced genomes that encompasses functional modules present broadly across all extant animals. Clustering revealed a spectrum of conservation, ranging from ancient Eukaryal assemblies likely serving cellular housekeeping roles for at least 1 billion years, ancestral complexes that have accrued contemporary components, and rarer metazoan innovations linked to multicellularity. We validated these projections by independent co-fractionation experiments in evolutionarily distant species, by affinity-purification and by functional analyses. The comprehensiveness, centrality and modularity of these reconstructed interactomes reflect their fundamental mechanistic significance and adaptive value to animal cell systems.

No MeSH data available.


Physical validation of complexesa, Verification of complexes from tagged human cell lines and transgenic worms (see Extended Methods). Inset reports spectral counts obtained in replicate AP/MS analyses of indicated bait protein (header). MIB2-VPS4 complex confirmed by co-IP (Extended Data Fig. 6a). b, Conserved complexes significantly overlap large-scale AP/MS data reported for human cell lines (BioGrid pre-pub 166968, Huttlin et al., 2015) to a comparable extent as literature reference sets39,42, using 3 measures of complex-level agreement (see Extended Methods, Extended Data Fig. 6b); ***, p-value < 0.001, determined by shuffling (gray distributions). c, Agreement of inferred molecular weights (MW) of human protein complexes with size exclusion chromatography (SEC) profiles (data in c, d from ref. 43). d, Co-elution of human Commander complex subunits by SEC consistent with an approx. 500 kDa particle.
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Figure 4: Physical validation of complexesa, Verification of complexes from tagged human cell lines and transgenic worms (see Extended Methods). Inset reports spectral counts obtained in replicate AP/MS analyses of indicated bait protein (header). MIB2-VPS4 complex confirmed by co-IP (Extended Data Fig. 6a). b, Conserved complexes significantly overlap large-scale AP/MS data reported for human cell lines (BioGrid pre-pub 166968, Huttlin et al., 2015) to a comparable extent as literature reference sets39,42, using 3 measures of complex-level agreement (see Extended Methods, Extended Data Fig. 6b); ***, p-value < 0.001, determined by shuffling (gray distributions). c, Agreement of inferred molecular weights (MW) of human protein complexes with size exclusion chromatography (SEC) profiles (data in c, d from ref. 43). d, Co-elution of human Commander complex subunits by SEC consistent with an approx. 500 kDa particle.

Mentions: We used multiple approaches to assess the accuracy (Fig. 4) and functional significance (Fig. 5) of the predicted complexes. First, we performed affinity purification-mass spectrometry (AP/MS) experiments on select novel complexes from the ‘new’, ‘old’ and ‘mixed’ age clusters, validating most associations in both worm and human (Fig. 4a, Extended Data Fig. 6a). We next performed a global validation by comparing our derived complexes to a newly reported large-scale AP/MS study of 23,756 putative human protein interactions detected in cell culture (BioGrid pre-publication 166968, Huttlin EL et al., downloaded Feb. 10, 2015), and observed a partial, but exceptionally significant, overlap to a degree comparable to literature-derived complexes (Fig. 4b, Extended Data Fig. 6b).


Panorama of ancient metazoan macromolecular complexes
Physical validation of complexesa, Verification of complexes from tagged human cell lines and transgenic worms (see Extended Methods). Inset reports spectral counts obtained in replicate AP/MS analyses of indicated bait protein (header). MIB2-VPS4 complex confirmed by co-IP (Extended Data Fig. 6a). b, Conserved complexes significantly overlap large-scale AP/MS data reported for human cell lines (BioGrid pre-pub 166968, Huttlin et al., 2015) to a comparable extent as literature reference sets39,42, using 3 measures of complex-level agreement (see Extended Methods, Extended Data Fig. 6b); ***, p-value < 0.001, determined by shuffling (gray distributions). c, Agreement of inferred molecular weights (MW) of human protein complexes with size exclusion chromatography (SEC) profiles (data in c, d from ref. 43). d, Co-elution of human Commander complex subunits by SEC consistent with an approx. 500 kDa particle.
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Related In: Results  -  Collection

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Figure 4: Physical validation of complexesa, Verification of complexes from tagged human cell lines and transgenic worms (see Extended Methods). Inset reports spectral counts obtained in replicate AP/MS analyses of indicated bait protein (header). MIB2-VPS4 complex confirmed by co-IP (Extended Data Fig. 6a). b, Conserved complexes significantly overlap large-scale AP/MS data reported for human cell lines (BioGrid pre-pub 166968, Huttlin et al., 2015) to a comparable extent as literature reference sets39,42, using 3 measures of complex-level agreement (see Extended Methods, Extended Data Fig. 6b); ***, p-value < 0.001, determined by shuffling (gray distributions). c, Agreement of inferred molecular weights (MW) of human protein complexes with size exclusion chromatography (SEC) profiles (data in c, d from ref. 43). d, Co-elution of human Commander complex subunits by SEC consistent with an approx. 500 kDa particle.
Mentions: We used multiple approaches to assess the accuracy (Fig. 4) and functional significance (Fig. 5) of the predicted complexes. First, we performed affinity purification-mass spectrometry (AP/MS) experiments on select novel complexes from the ‘new’, ‘old’ and ‘mixed’ age clusters, validating most associations in both worm and human (Fig. 4a, Extended Data Fig. 6a). We next performed a global validation by comparing our derived complexes to a newly reported large-scale AP/MS study of 23,756 putative human protein interactions detected in cell culture (BioGrid pre-publication 166968, Huttlin EL et al., downloaded Feb. 10, 2015), and observed a partial, but exceptionally significant, overlap to a degree comparable to literature-derived complexes (Fig. 4b, Extended Data Fig. 6b).

View Article: PubMed Central - PubMed

ABSTRACT

Macromolecular complexes are essential to conserved biological processes, but their prevalence across animals is unclear. By combining extensive biochemical fractionation with quantitative mass spectrometry, we directly examined the composition of soluble multiprotein complexes among diverse metazoan models. Using an integrative approach, we then generated a draft conservation map consisting of &gt;1 million putative high-confidence co-complex interactions for species with fully sequenced genomes that encompasses functional modules present broadly across all extant animals. Clustering revealed a spectrum of conservation, ranging from ancient Eukaryal assemblies likely serving cellular housekeeping roles for at least 1 billion years, ancestral complexes that have accrued contemporary components, and rarer metazoan innovations linked to multicellularity. We validated these projections by independent co-fractionation experiments in evolutionarily distant species, by affinity-purification and by functional analyses. The comprehensiveness, centrality and modularity of these reconstructed interactomes reflect their fundamental mechanistic significance and adaptive value to animal cell systems.

No MeSH data available.