Characterization of initiation of DNA replication in HCT116 Ku80+- Cells
Bibliographic record
Abstract
DNA replication initiates at origins with the origin recognition complex (ORC) binding and activating initiation. Although ORC specifically binds origins in vivo, it possesses no sequence specific DNA binding activity in vitro. Thus, it is thought to interact with another protein that recruits it to the origin. The heterodimeric Ku (Ku70/Ku80) protein is one candidate. Ku is an abundant nuclear protein that has recently been implicated in the initiation of DNA replication, but the mechanism by which it performs this function is yet unknown. Here, using haploinsufficient human HCT116 Ku80 +/- cells that express 25-40% Ku80 protein, the origin activity and origin-association of other replication proteins were examined in a Ku80-deficient environment. Ku80 was not found to influence the expression of replication proteins, except for Ku70, its partner, which displayed a reduction of its protein expression to 64% of wildtype cells. Ku80+/- cells displayed a prolongation of G1 phase by 2 hours, but normal progression through S-phase. Furthermore, the activity of the early-activated replication origins of the lamin B2, beta-globin and c-myc loci decreased by 4.5-, 3.4-, and 4.3-fold, respectively. This was associated with a 2.1-, 1.5- and 1.7-fold decrease in the association of Ku80 with these origins, respectively, and a 1.5-, 2.3- and 2.5-fold decrease in that of Ku70, respectively. Moreover, the origin-association of three of the six subunits of the ORC complex, Orc-3, -4, and -6 was significantly decreased in Ku80 +/- cells, indicating decreased ORC complex formation at the origins. In contrast, the association of Orc-2 was not influenced by Ku80 deficiency, nor was the association of Ku80 with origins affected by Orc2 deficiency. Overall, these results suggest that Ku binds origins to allow assembly of the ORC complex, which is necessary for origin licensing and activation.
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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".