IDENTIFICATION AND LOCALIZATION OF NA+ K+ATPASE IN BOVINE SPERMATOZOA
Bibliographic record
Abstract
Na+ K+ATPase is a ubiquitous transmembrane protein. In sperm, Na+ K+ATPase is important for fertility and capacitation, participating in ion transport and very recently shown to function as a signaling molecule in tyrosine phosphorylation. Na+ K+ATPase is a dimer, with isoforms of its alpha (1, 2, 3, and 4; approximately 110 kDa) and beta (1, 2, and 3; approximately 55 kDa) subunits. We hypothesized that specific alpha and beta isoforms in the head are more likely to be active in sperm capacitation, whereas those present in the midpiece or tail region are more likely involved in sperm flagellar activity, so this study's objective was to identify and localize subunits of Na+ K+ATPase within bull sperm. The presence and localization of Na+ K+ATPase in fresh bull sperm was identified by immunoblotting and immunocytochemistry using mouse monoclonal antibodies against the alpha 1 and 3 isoforms, and the beta 1, 2 and 3 isoforms. The relative quantity of Na+ K+ATPase present in spermatozoa HPM from different bulls was determined by densitometry of immunoblot bands and compared to ATPase in bovine kidney membranes. Sperm and kidney specifically bound all antibodies at kDa equivalent to the supplied positive control and to additional bands; the kDa of the additional bands differed between the tissues. Deglycosylation of the B1 subunit reduced the kDa of the bands in both sperm and kidney, but the mass of the deglycosylated bands still differed between kidney and sperm. All alpha and beta isoforms were detectable in bull HPM, at significantly lower concentrations than in the kidney. The most prevalent isoforms in sperm HPM were B1 (2030-4085 units/mg protein) and A3 (1650 units/mg protein), with the least being B2, where there were multiple low intensity bands; however in kidney the most abundant were A1 (402,572 units/mg protein) and B1 (136,873 units/mg protein) with multiple bands of low intensity observed in B2. The B1 isoform showed 4 bands in sperm HPM (70, 60, 40 and 45 kDa), and one band in kidney (55 kDa). Two bands were observed in HPM from the A1 isoform (105 and 100 kDa) and one band in kidney (100 kDa). Multiple bands were observed in A3, B2 and B3 for both sperm HPM and kidney, however, rat brain controls showed a single band at the appropriate molecular mass for each isoforms. Immunofluorescence of intact fresh bull sperm confirmed that all isoforms were present in the head region of spermatozoa in which HPM's were naturally disrupted or deliberately permeabilised, however completely intact cells showed no fluorescence, suggesting that the antibody binding sites are either present on the inner surface of the HPM or on the outer surface of the acrosomal membrane. The A3 subunit was also uniformly distributed postequatorially. The unique quantity, location and structure of Na+ K+ATPase in sperm suggests that this protein has unique functions in sperm. (poster)
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How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 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) | 0.000 | 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 teacher head, 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".