Macromolecular Connectivity Landscape of Mammalian Brain

Pourhaghighi, Mohammad Reza; Ash, Peter E.A.; Phanse, Sadhna; Goebels, Florian; Małolepsza, Edyta; Tsafou, Kalliopi; Nathan, Aparna; Chen, Siwei; Zhang, Yingying; Wierbowski, Shayne D.; Boudeau, Samantha; Hu, Lucas; Moutaoufika, Mohamed T.; Malty, Ramy H.; Cormar, Graham; Guo, Hongbo; Abdullatif, Ali Al; Apicco, Daniel J.; Becker, Lindsay A.; Gitler, Aaron D.; Youssef, Ahmed; Bryant, Camron D.; Parkinson, John; Lage, Kasper; Babu, M. Madan; Yu, Haiyuan; Wolozin, Benjamin; Bader, Gary D.; Emili, Andrew

doi:10.2139/ssrn.3293682

Cited by 2 publications

(6 citation statements)

References 61 publications

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“…If we apply an even more stringent filter, and require that proteins are found in all soluble fractionations, or all xCF-MS fractionations, or in the cross-linking set, we still recover 8,684 proteins, covering 69.2% of mouse brain proteins and 56.3% of the transmembrane proteins. This represents a substantial increase in coverage over prior brain CF/MS data (Pourhaghighi et al, 2018) , and saturation analysis suggests that further experiments using our broad range of conditions are not likely to increase proteome coverage ( Figure 2B ).…”

Section: Native Fractionation Achieves High Coverage Of the Mouse Bramentioning

confidence: 75%

“…By combining proteomics methods, we reveal protein interactions that span the range of cellular functions in the brain. Standard CF-MS is well-suited to determining soluble protein complexes (Havugimana et al, 2012;Kirkwood et al, 2013;Pourhaghighi et al, 2018;Wan et al, 2015) . Thus, similar to previous studies, we recover conserved, soluble protein complexes such as the proteasome, Cop9 signalosome, and the T-complex/TriC/CCT molecular chaperone ( Supplemental Figure 2 ).…”

Section: Co-fractionation Broadly Recovers Neural Protein Complexesmentioning

confidence: 99%

“…This pipeline allows the determination of protein interactions from co-fractionation profiles of candidate protein pairs, and is therefore called co-fractionation mass-spectrometry (CF-MS). A recent study used CF-MS to measure protein interactions from the mouse brain (Pourhaghighi et al, 2018) , resulting in the large-scale first map of soluble protein complexes from the brain. However, one draw-back of standard CF-MS pipelines is their reliance on the solubility of native protein complexes.…”

Section: Introductionmentioning

confidence: 99%

“…This integrative proteomics approach allowed us to improve on current methods and create a map of the functional protein neighborhoods of the mouse brain. Our combined approach allowed us to do so without the addition of external datasets, such as mRNA co-expression, which Pourhaghighi et al used to boost the performance of their method (Pourhaghighi et al, 2018) . This map of functional neighborhoods is therefore the first large-scale map of protein complexes in the mammalian brain that relies solely on proteomics of endogenous proteins isolated directly from the mammalian brain.…”

Section: Introductionmentioning

confidence: 99%

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Mapping Functional Protein Neighborhoods in the Mouse Brain

Liebeskind

Young

Halling

et al. 2020

Preprint

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New proteomics methods make it possible to determine protein interaction maps at the proteome scale without the need for genetically encoded tags, opening up new organisms and tissue types to investigation. Current molecular and computational methods are oriented towards protein complexes that are soluble, stable, and discrete. However, the mammalian brain, among the most complicated and most heavily studied tissue types, derives many of its unique functions from protein interactions that are neither discrete nor soluble. Proteomics investigations into the global protein interaction landscape of the brain have therefore leveraged non-proteomics datasets to supplement their experiments. Here, we develop a novel, integrative proteomics pipeline and apply it to infer a global map of functional protein neighborhoods in the mouse brain without the aid of external datasets. By leveraging synaptosome enrichment and interactomics methods that target both soluble and insoluble protein fractions, we resolved protein interactions for key neural pathways, including those from refractory subcellular fractions such as the membrane and cytoskeleton. In comparison to external datasets, our observed interactions perform similarly to hand-curated synaptic protein interactions while also suggesting thousands of novel connections. We additionally employed cleavable chemical cross-linkers to detect direct binding partners and provide structural context. Our combined map suggests new protein pathways and novel mechanisms for proteins that underlie neurological diseases, including autism and epilepsy. Our results show that proteomics methods alone are sufficient to determine global interaction maps for proteins that are of broad interest to neuroscience. We anticipate that our map will be used to prioritize new research avenues and will pave the way towards future proteomics techniques that resolve protein interactions at ever greater resolution.

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Section: Native Fractionation Achieves High Coverage Of the Mouse Bramentioning

confidence: 75%

Section: Co-fractionation Broadly Recovers Neural Protein Complexesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Mapping Functional Protein Neighborhoods in the Mouse Brain

Liebeskind

Young

Halling

et al. 2020

Preprint

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“…81 Regarding cytoskeletal regulation by ArhGAPs, α-tubulin regulates ArhGAP21 localization and functions 82 (Table 2, Figure 7b). Focal adhesion kinase (FAK) is an ArhGAP21 and Graf1 binding protein, and RhoA and Cdc42 may be cooperatively regulated by these RhoGAPs in dendritic organization 6,20,83,84 (Figure 7b). RhoB-knockout pyramidal neurons show complex dendritic blanching and immature spine development.…”

Section: Pdz Regions Of Arhgap21/23 Possess a Conserved Globular Stru...mentioning

confidence: 99%

A Structural Network Analysis of Neuronal ArhGAP21/23 Interactors by Computational Modeling

Kouchi

Kojima

2023

ACS Omega

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RhoGTPase-activating proteins (RhoGAPs) play multiple roles in neuronal development; however, details of their substrate recognition system remain elusive. ArhGAP21 and ArhGAP23 are RhoGAPs that contain N-terminal PDZ and pleckstrin homology domains. In the present study, the RhoGAP domain of these ArhGAPs was computationally modeled by template-based methods and the AlphaFold2 software program, and their intrinsic RhoGTPase recognition mechanism was analyzed from the domain structures using the protein docking programs HADDOCK and HDOCK. ArhGAP21 was predicted to preferentially catalyze Cdc42, RhoA, RhoB, RhoC, and RhoG and to downregulate RhoD and Tc10 activities. Regarding ArhGAP23, RhoA and Cdc42 were deduced to be its substrates, whereas RhoD downregulation was predicted to be less efficient. The PDZ domains of ArhGAP21/23 possess the FTLRXXXVY sequence, and similar globular folding consists of antiparalleled β-sheets and two α-helices that are conserved with PDZ domains of MAST-family proteins. A peptide docking analysis revealed the specific interaction of the ArhGAP23 PDZ domain with the PTEN C-terminus. The pleckstrin homology domain structure of ArhGAP23 was also predicted, and the functional selectivity for the interactors regulated by the folding and disordered domains in ArhGAP21 and ArhGAP23 was examined by an in silico analysis. An interaction analysis of these RhoGAPs revealed the existence of mammalian ArhGAP21/23-specific type I and type III Arf- and RhoGTPase-regulated signaling. Multiple recognition systems of RhoGTPase substrates and selective Arf-dependent localization of ArhGAP21/23 may form the basis of the functional core signaling necessary for synaptic homeostasis and axon/dendritic transport regulated by RhoGAP localization and activities.

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Macromolecular Connectivity Landscape of Mammalian Brain

Cited by 2 publications

References 61 publications

Mapping Functional Protein Neighborhoods in the Mouse Brain

Mapping Functional Protein Neighborhoods in the Mouse Brain

A Structural Network Analysis of Neuronal ArhGAP21/23 Interactors by Computational Modeling

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