Proteolytic cleavage of the hydrophobic domain in the CaVα2δ1 subunit improves assembly and activity of cardiac CaV1.2 channels
Bibliographic record
Abstract
Voltage-gated L-type Ca V 1.2 channels in cardiomyocytes exist as heteromeric complexes with the pore-forming Ca V α1, Ca V β, and Ca V α2δ1 subunits. The full complement of subunits is required to reconstitute the native-like properties of L-type Ca 2+ currents, but the molecular determinants responsible for the formation of the heteromeric complex are still being studied. Enzymatic treatment with phosphatidylinositol-specific phospholipase C, a phospholipase C specific for the cleavage of glycosylphosphatidylinositol (GPI)-anchored proteins, disrupted plasma membrane localization of the cardiac Ca V α2δ1 prompting us to investigate deletions of its hydrophobic transmembrane domain. Patch-clamp experiments indicated that the C-terminally cleaved Ca V α2δ1 proteins up-regulate Ca V 1.2 channels. In contrast, deleting the residues before the single hydrophobic segment (Ca V α2δ1 Δ1059–1063) impaired current up-regulation. Ca V α2δ1 mutants G1060I and G1061I nearly eliminated the cell-surface fluorescence of Ca V α2δ1, indicated by two-color flow cytometry assays and confocal imaging, and prevented Ca V α2δ1-mediated increase in peak current density and modulation of the voltage-dependent gating of Ca V 1.2. These impacts were specific to substitutions with isoleucine residues because functional modulation was partially preserved in Ca V α2δ1 G1060A and G1061A proteins. Moreover, C-terminal fragments exhibited significantly altered mobility in denatured immunoblots of Ca V α2δ1 G1060I and Ca V α2δ1 G1061I, suggesting that these mutant proteins were impaired in proteolytic processing. Finally, Ca V α2δ1 Δ1059–1063, but not Ca V α2δ1 G1060A, failed to co-immunoprecipitate with Ca V 1.2. Altogether, our data support a model in which small neutral hydrophobic residues facilitate the post-translational cleavage of the Ca V α2δ1 subunit at the predicted membrane interface and further suggest that preventing GPI anchoring of Ca V α2δ1 averts its cell-surface expression, its interaction with Ca V α1, and modulation of Ca V 1.2 currents.
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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 | 0.001 |
| Insufficient payload (model declined to judge) | 0.003 | 0.001 |
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".