Involvement of MURC/Cavin‐4 in store‐operated Ca <sup>2+</sup> entry in neonatal cardiomyocytes
Bibliographic record
Abstract
STIM1 is a regulatory protein of store‐operated calcium entry (SOCE) and its expressions of in the cardiomyocytes suggests that it may play a role in cardiomyocytes function. To better understand this role, we sought to identify unknown protein partners of STIM1. We fused the ERM domain of STIM1 (amino acids 219–548) to GST and used it for a GST‐pulldown assay combined with analytical liquid chromatography‐mass spectrometry. From the lysate of RIPA‐solubilized proteins from adult rat hearts, we identified the muscle‐related coiled‐coil protein (MURC), also known as Cavin‐4, as a potential candidate. Also, we show that is the HR1 domain of MURC interacts with the ERM domains of STIM1. To understand the role of this interaction, we overexpressed MURC in neonatal rat ventricular myocytes (NRVM) and determined is impact on SOCE. NRVM were loaded with the Ca2+ probe, FURA‐2AM, and incubated with nifedipine to prevent Ca 2+ entry at the plasma membrane via L‐type calcium channels. To induce intracellular store depletion and SOCE activation, 10mM caffeine and 2μM thapsigargin were added to the perfusion buffer in the absence of extracellular Ca 2+ and Ca 2+ entry was determined when Ca 2+ was re‐introduced in the extracellular medium. Using this protocol, we demonstrated that the Ca2+ entry in NRVM overexpressing MURC is 1.8‐fold higher that control bGal condition. This effect is reduced by SOCE inhibitors (SKF‐96365, 2‐APB, and YM‐58483). The Ca 2+ entry measured in NRVM overexpressing ΔHR1‐MURC was similar to the Ca 2+ entry in the bGal control. These results indicate that MURC overexpression relies on its HR1 domain to potentiate SOCE in these cells. The increase in SOCE in MURC overexpressing NRVM may be the result from an enhanced interaction between Orai1 and STIM1. To test this possibility, we assessed the extent of STIM1‐Orai1 complex formation with a co‐immunoprecipitation assay using HA‐Orai1 as the target protein. The STIM1‐Orai1 interaction was evaluated in resting conditions or upon SOCE activation. As expected, SOCE activation enhanced the interaction between STIM1 and Orai1 by 2.80‐fold compared to the control condition. MURC overexpression was also sufficient to increase the STIM1‐Orai1 complex formation in resting (1.78‐fold) and Ca 2+ store‐depleted (2.44‐fold) conditions. These results show that MURC overexpression in NRVM favors the interaction of STIM1 with Orai1. Also, we show that R140W‐MURC mutant, a missense variant of the HR1 domain associated with human dilated cardiomyopathy, exacerbates the SOCE increase in NRVM. Our study provides novel evidence that MURC regulates SOCE by interacting with STIM1 in cardiomyocytes. In addition, we identified a first potential mechanism by which the R140W mutation may contribute to Ca 2+ mishandling and the development of cardiomyopathies. Support or Funding Information Supported by the Natural Sciences and Engineering Research Council of Canada (NSERC) [M.A.M, and G.B.], CRMUS graduate student research award [J.M.], FRQS doctoral training award [J.D.], CIHR graduate scholarship [H.G.], André‐Lussier Research Chair [J.‐L.P.], and FRQS and Heart and Stroke Foundation of Canada Investigator awards [M.A‐M.]. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.
How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".