Cloning and characterization of the invertebrate visual iG(q)alpha: Membrane association, receptor coupling efficacy and turnover
Bibliographic record
Abstract
This study reports the cloning and characterization of invertebrate visual iGqalpha membrane association, receptor coupling efficacy and turnover. iGqalpha relays signals from rhodopsin to phospholipase C leading to membrane depolarization. When its full-length cDNA from L. pealei was cloned, its deduced amino acid sequence was 77% identical to mammalian Gqalpha. Both G proteins are highly homologous and iGqalpha contained conserved N-terminal cysteine residues that can be palmitoylated, a GTP-binding motif, three SWITCH regions and a site for effector/receptor interactions in the C-terminus. The role of G protein palmitoylation remains unclear. My findings suggest that iGqalpha is palmitoylated but that this is not important for its initial membrane targeting. The palmitoylation-deficient iGqalpha(C3,4A) however, is unable to maintain membrane attachment following receptor activation. Thus activated iGga(C3,4A) could translocate from membrane to cytosol and mimicked the translocation of iGqalpha observed in intact invertebrate photoreceptors. The activation-dependent translocation of iGqalpha may act to regulate the amount of a subunits which can interact with activated rhodopsin therefore regulating photosensitivity in the invertebrate eye. While palmitoylated iGqalpha expressed in HEK 293T cells do not show any significant changes in protein levels in the basal and stimulated conditions, non-palmitoylated iGqalpha turnover, initially has a half-life of 20 hours under basal conditions and decreases to 10 hours upon agonist stimulation. Accelerated turnover of non-palmitoylated iGqalpha is attributed to increased degradation by the ubiquitin-proteasomal pathway. My findings demonstrate not only an activation-dependent and cellular localization-dependent proteolysis of a G protein, but also for the first time implicate the ubiquitin-proteasomal pathway in the degradation and turnover of a Gqalpha protein. Lastly, the receptor-interacting N-terminal domain contains a six amino acid extension MTLESI in mammalian mGqalpha that is not present in iGqalpha. I explored the function of MTLESI by testing its role in receptor specificity and efficacy. My findings suggest that the lack of MTLESI on iGqalpha increases its efficacy in coupling to G q-linked receptors but does not decrease its specificity. Furthermore, the mechanism by which the efficacy of iGqalpha is regulated appears to be at the level of guanine nucleotide exchange as demonstrated by increased GTPgamma[35S] binding of iGqalpha compared with mGqalpha.
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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 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 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".