Cooperative Interactions between the Two DNA Binding Domains of Pax3: Helix 2 of the Paired Domain Is in the Proximity of the Amino Terminus of the Homeodomain
Bibliographic record
Abstract
Pax3 is a transcription factor that plays an important role during neurogenesis and myogenesis, and Pax3 mutant animals display neural tube defects and lack limb muscles. Pax3 harbors two DNA binding domains, the paired domain (PD) and a paired-type homeodomain (HD). Genetic and biochemical data have (i) identified strong cooperative interactions between the PD and HD domains for DNA binding in the intact Pax3 protein and (ii) suggested an important role for the amino-terminal portions of both domains in such cooperativity. We have studied proximity relationships between the PD and HD of Pax3. For this, we have used a cross-linking strategy with the bifunctional thiol reagent bismaleimidoethane (BMOE) in 21 mutants bearing pairs of cysteine residues (DCM) inserted in strategic locations of a functional Pax3 protein otherwise devoid of endogenous cysteine residues. All 21 DCMs were characterized for protein stability, for DNA binding by the PD and HD, and for the effect of BMOE on protein binding to PD, HD, or PD-HD combined DNA targets. BMOE-induced cross-links in DCMs were detected as slower migrating species on immunoblots. Mutants bearing double cysteine insertions (I59C/S222C, S73C/Q219C, and V78C/K218C) showed the most robust cross-linking upon BMOE exposure. These cross-linking studies suggest that portions of helix 1 (I59), helix 2 (S73), and the loop between helices 2 and 3 (V78) of the PD are in the proximity of the N-terminal segment of the HD (K218, Q219, and S222) in the tertiary structure of Pax3. These results are compatible with a model in which the PD and HD are organized in an everted arrangement, with the N-terminal portion of the PD being in the proximity of the N-terminus of the HD. This arrangement may be important for the noted PD-HD cooperativity in DNA binding.
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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 | 0.000 |
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".