Analysis of candidate regulators of TBC-2 and endosome maturation in «Caenorhabditis elegans»
Bibliographic record
Abstract
The Rab5 and Rab7 GTPases are key regulators of endosome to lysosome trafficking, whose activities are positively regulated by Rab Guanine nucleotide Exchange Factors and negatively regulated by GTPase Activating Proteins (GAP). In this thesis, the nematode Caenorhabditis elegans was used as a model system to study regulation of Rab GTPase-mediated endosomal trafficking, for simple genetics and easy visualization of organelles under Differential Interference Contrast (DIC) and confocal optics. It was previously demonstrated that TBC-2 functions as a RAB-5 GAP. Loss of tbc-2 activity results in the formation of enlarged late endosomes in the intestine that require the activities of RAB-5, RAB-7 and components of the homotypic fusion and vacuole protein sorting (HOPS) complex. TBC-2 colocalizes with RAB-7 on late endosomes, and requires RAB-7 for membrane localization. My hypothesis is that RAB-7 recruits TBC-2 to late endosomes to inactivate RAB-5 and possibly RAB-7 itself, to facilitate early to late endosome maturation. The vh8 mutant was previously identified in the lab as a suppressor of the tbc-2(-) phenotype and displaces GFP::RAB-7 from vesicular membranes. It is possible that vh8 encodes for a potential RAB-7 GEF. To investigate the molecular identity of vh8, I tested candidate genes by RNAi. While none of the candidates appear to be vh8, I did find that the HOPS core complex is required for the tbc-2(-) phenotype. ARL-8, and its human homolog arl8b, have recently been shown to be important for late endosome to lysosome trafficking. To investigate whether ARL-8 is required for TBC-2 mediated early to late endosome trafficking, I performed RNAi experiments. The results of which suggest that ARL-8 functions downstream of TBC-2. ARL-8 is required for the tbc-2(-) large late endosome phenotype, but is not required for membrane localization of either TBC-2 or RAB-7.In mammalian cells, the Rac1 GTPase is able to bind Armus, a human homolog of TBC-2. That raises the possibility that RAB-7 recruits CED-10/Rac1 via TBC-2. However, I observe that in tbc-2(tm2241) mutant animals, CED-10 is still localized to vesicles. It suggests that CED-10 might be recruited to membrane via other mechanism. Active CED-10 is also crucial for phagocytosis and clearance of apoptotic cell corpses. CED-10 can be activated by the CED-5/CED-12 complex, which is proposed to be recruited and activated by PI(3,5)P2. Work from this thesis suggests that the active state might not be a requirement for CED-10 localization, as ced-5 RNAi does not change the localization of CED-10.
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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 | 0.000 | 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) | 0.000 | 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".