Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Molecular basis of F-actin regulation and sarcomere assembly via myotilin

ABSTRACT: Sarcomeres, the basic contractile units of striated muscle cells, contain arrays of thin (actin) and thick (myosin) filaments that slide past each other during contraction. The Ig-like domain containing protein myotilin provides structural integrity to Z-discs—the boundaries between adjacent sarcomeres. Myotilin binds to Z-disc components, including F-actin and α-actinin-2, but the molecular mechanism of binding and implications of these interactions on Z-disc integrity are still elusive. To illuminate them we used a combination of small angle X-ray scattering, cross-linking mass spectrometry, biochemical and molecular biophysics approaches. We discovered that myotilin displays conformational ensembles in solution. We generated a structural model of the F-actin:myotilin complex that revealed how myotilin interacts with and stabilizes F-actin via its Ig-like domains and flanking regions. Mutant myotilin designed with impaired F-actin binding showed increased dynamics in cells. Structural analyses and competition assays uncovered that myotilin displaces tropomyosin from F-actin. Our findings suggest a novel role of myotilin as a co-organizer of Z-disc assembly and advance our mechanistic understanding of myotilin's structural role in Zdiscs.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Striated Muscle Cell

SUBMITTER: Friedel Drepper

LAB HEAD: Bettina Warscheid

PROVIDER: PXD024301 | Pride | 2021-02-22

REPOSITORIES: Pride

ACCESS DATA

Json Xml

Similar Datasets

Quality control at the cell surface prevents accumulation of damaged L-type CaV1.2 calcium channels

Project description:Defective ion channel turnover and clearance of damaged proteins are associated with aging and neurodegeneration. The L-type CaV1.2 voltage-gated calcium channel mediate depolarization-induced calcium signals in heart and brain. Here, we determined the interaction surface between the L-type calcium channel CaVβ subunit and actin using cross-linking mass spectrometry and protein-protein docking, and uncovered a role in replenishing damaged CaV1.2 channels. Computational and in vitro mutagenesis identified hotspots in CaVβ that decrease its affinity for actin but not for CaV1.2. Coexpression of an actin-association-deficient CaVβ mutant with the CaV1.2 channel downregulated current amplitudes with a concomitant reduction in the number of functionally available channels. Neither alterations in the single-channel properties nor changes in the total number of channels at the cell surface were found, indicating that current inhibition resulted from a build-up of conduction-defective channels. Our findings established CaVβ–actin interaction as a key player for selective monitoring and clearing corrupted CaV proteins to ensure the maintenance of a functional pool of channels and proper calcium signal transduction. The CaVβ–actin molecular model introduces a potentially druggable protein-protein interface to intervene CaV-mediated signaling processes.

2024-02-16 | PXD044630 | Pride

Quality control at the cell surface prevents accumulation of damaged L-type CaV1.2 calcium channels

2024-02-16 | PXD044723 | Pride

Molecular characterization of a complex of Apoptosis Inducing Factor 1 (AIFM1) with cytochrome c oxidase of the mitochondrial respiratory chain

Project description:Combining mass spectrometry based chemical cross-linking and complexome profiling, we analyzed the interactome of heart mitochondria. We focused on complexes of oxidative phosphorylation and found that dimeric apoptosis inducing factor 1 (AIFM1) forms a defined complex with ~10% of monomeric cytochrome c oxidase (COX), but hardly interacts with respiratory chain supercomplexes. Multiple AIFM1 inter-crosslinks engaging six different COX subunits provided structural restraints to build a detailed atomic model of the COX-AIFM12 complex. Application of two complementary proteomic approaches thus provided unexpected insight into the macromolecular organization of the mitochondrial complexome. Our structural model excludes direct electron transfer between AIFM1 and COX. Notably however, the binding site of cytochrome c remains accessible allowing formation of a ternary complex. The discovery of the previously overlooked COX-AIFM12 complex and clues provided by the structural model hint at a role of AIFM1 in OXPHOS biogenesis and in programmed cell death.

2021-09-09 | PXD025102 | Pride

RNxQuest: an extension to the xQuest pipeline enabling analysis of protein-RNA cross-linking/mass spectrometry data

Project description:The recently introduced cross-linking of isotope-labelled RNA coupled with mass spectrometry (CLIR-MS) technique enables protein-RNA cross-links to be used as precisely localized distance restraints in de novo structural modelling. The novel data type requires a bespoke data analysis approach. The new RNxQuest Python package supports this approach. Here we demonstrate the performance of the new package using a mixture of novel and published datasets.

2023-09-06 | PXD039754 | Pride

MRPS36 provides a missing link in the eukaryotic 2-oxoglutarate dehydrogenase complex for recruitment of E3 to the E2 core - Phospoproteomics data

Project description:The tricarboxylic acid (TCA) cycle, or Krebs cycle, is the central pathway of energy production in eukaryotic cells and plays a key part in aerobic respiration throughout all kingdoms of life. The enzymes involved in this cycle generate the reducing equivalents NADH and FADH2 by a series of enzymatic reactions, which are utilized by the electron transport chain to produce ATP. One of the key enzymes in this cycle is 2-oxoglutarate dehydrogenase (OGDHC), which generates NADH by oxidative decarboxylation of 2-oxoglutarate to succinyl-CoA. Notably, this enzyme consists of multiple subunits as a megadalton protein complex. Thus far, it was thought that OGDHC consists of solely three catalytically active subunits (E1, E2, E3). However in fungi and animals, the small protein MRPS36 has been proposed as a putative additional component. Based on extensive XL-MS data obtained from measurements in both, mice and bovine heart mitochondria, and from phylogenetic analyses, we provide structural evidence that MRPS36 is an exclusive and crucial member of eukaryotic OGDHC. Comparative genomics analysis and computational structure predictions reveal that in eukaryotic OGDHC, E2o does not contain a peripheral subunit-binding (PSBD) domain. Instead, our data provide compelling evidence that in eukaryotes, MRPS36 evolved as E3 adaptor protein, functionally replacing the PSBD domain. Based on our data we provide a refined structural model of the complete eukaryotic OGDHC assembly containing all its 58 subunits (~ 3.4 MDa). The model provides new insights into the protein-protein interactions within the OGDH complex and highlights putative mechanistic implications.

2023-03-10 | PXD036896 | Pride

Tissue Transglutaminase 2 has higher affinity for relaxed than for stretched fibronectin fibers

Project description:Chemical cross-linking coupled to mass spectrometry was used to study the interaction between fibronectin (Fn; full-length or gelatin-binding domain fragment = Fn 45) with tissue transglutaminase 2 in oxidized and non-oxidized form. The complexes were cross-linked with disuccinimidyl suberate (DSS) or a combination of pimelic dihydrazide (PDH) in combination with the zero-length cross-linking reagent, DMTMM.

2023-12-20 | PXD043976 | Pride

Integrative Structural Characterization of the Human Canonical and Non-canonical Cop9 Signalosome Complexes

Project description:The COP9 signalosome (CSN) is an evolutionarily conserved protein complex that functions as a deneddylase to inactivate Cullin-RING E3 ligases for controlling protein ubiquitination. CSN possesses structural flexibility that is important for its activation upon binding to a diverse array of CRLs. The canonical and non-canonical CSN complexes consist of 8 (CSN1-8) and 9 (CSN1-9) subunits, respectively. Although CSN9 is not essential for CSN assembly and function, it appears to be important in non-catalytic regulation of CRLs by CSN. Here we employed a combinatory cross-linking mass spectrometry (XL-MS) approach to generate the largest PPI maps of human CSN complexes, which significantly enhanced the precision of integrative structural modeling. The resulting integrative structures allowed us not only to elucidate architectures of both complexes, but also to assess CSN structural dynamics that was not described in the crystal structure. In addition, we have determined CSN9 docking sites and its impact on the CSN structure. While CSN9 binding did not induce global conformational changes, it triggered subunit local reorientations that may be associated with CSN9 involvement in CSN-mediated steric regulation of CRLs.

2020-02-04 | PXD014673 | Pride

CASP XL-MS target T0975: Cross-linking for the 13th Community Wide Experiment on the Critical Assessment of Techniques for Protein Structure Prediction (CASP13)

Project description:Chemical cross-linking coupled to mass spectrometry was used to provide distance restraints for different homo- and heterooligomeric complexes as part of the CASP13 experiment (https://www.predictioncenter.org/casp13/index.cgi).

2019-10-02 | PXD010385 | Pride

CASP XL-MS target H0968 (ID32): Cross-linking for the 13th Community Wide Experiment on the Critical Assessment of Techniques for Protein Structure Prediction (CASP13)

2019-10-02 | PXD010004 | Pride

CASP XL-MS target H0953 (ID 51): Cross-linking for the 13th Community Wide Experiment on the Critical Assessment of Techniques for Protein Structure Prediction (CASP13)

2019-10-02 | PXD010094 | Pride