All 88 fetched assertions, four alternative products and 39 original reference identity pairs are retained. Every assertion was assessed individually. The review contains 23 ACCEPT, 23 MODIFY, 39 KEEP_AS_NON_CORE, two UNDECIDED and one MARK_AS_OVER_ANNOTATED decisions, with no NEW annotation. The two added references are the directly inspected mouse donor paper and this review’s compact evidence index. Positive generic interactions remain non-core unless the actual partner class supports a more precise term, following the campaign policy in projects/CLINGEN_MENDELIAN.md.
The genuine default Falcon attempt timed out; one recorded fallback failed with an insufficient-quota response. No provider report was invented. These manual notes document the actual literature work. Normal fetched and selected exact-main sources are preserved separately. Full-text flags were not treated as proof of full access: PMID:33961781 is partial, while PMID:21976065 remains an abstract in the publication cache despite separate recovery of its original publisher PDF.
CNOT1 supplies the physical framework of CCR4–NOT and recruitment interfaces for factors targeting mRNAs. The human NOT1–CAF1 structure and direct binding experiments establish scaffold work; the functional reporter in that particular study uses Drosophila S2 cells (PMID:22977175, Results and Materials and Methods). Reconstitution of human complexes independently establishes the noncatalytic modules’ contribution to efficient deadenylation (PMID:31320642, Results and Methods). GO:0140378 is therefore the single core molecular function and replaces the two broader adaptor annotations. The GO:0030674 comment expressly distinguishes complex scaffolds from other adaptors. The historical PAINT node was not inspected; this refinement is based on target experiments, not a reconstructed ancestral error.
The preserved GO:0004535 assertion already has contributes_to. The original 2011 study measures reduced poly(A) degradation by CNOT6L immunoprecipitates after CNOT1 depletion, even when exogenous CNOT6L abundance is unchanged. It also shows human FLAG-CNOT1 in cytoplasmic foci and reduced AGO2-positive P-body counts without reduced AGO2 abundance (PMID:21976065, Results and deadenylase-assay Methods; original publisher paper). These data support the complex’s contributed nuclease activity and a contextual P-body assembly role, without establishing intrinsic CNOT1 hydrolysis or direct condensate nucleation. The RNase contribution is integrated into the scaffold core rather than repeated as a second core.
TTP recruitment experiments use human CNOT1 fragments named with historical isoform-C/2371-residue coordinates, mouse Krebs extracts for deadenylation and a human HEK293 reporter for cellular output (PMID:23644599, Results). Those settings are kept distinct. Current alternative-product reconstruction gives lengths 2376, 2371, 2150 and 1551 residues from the authentic UniProt VAR_SEQ features. Neither those lengths nor engineered structural fragments establish equivalent activity of all four natural products.
Both original RNA-capture annotations can be refined to mRNA binding. The actual Castello Table S1 row 359 identifies CNOT1/ENSG00000125107 in the crosslinked HeLa mRNA interactome, with no control identification. The original Methods and supplemental Methods were inspected. The Baltz source is supported by the complete abstract and the exact CNOT1 row reproduced in the first author’s thesis, PDF page 144; the full original Methods were not recovered (PMID:22658674; PMID:22681889). These findings support RNA contact but not an autonomous RNA-recognition specificity or catalytic activity.
The interaction audit preserves literal IntAct/MINT methods, species, roles, features and record identifiers. Current database joins are corroboration, not substitutes for the cited assays; a zero join does not refute a published interaction. CNOT6/CNOT6L/CNOT7/CNOT8 associations are refined to enzyme binding using verified nuclease identities. Association in a complex is not invariably a purified binary interface. DDX6 binding has the more precise DEAD/H-box RNA-helicase term, whereas the resolved W-binding pockets are in CNOT9, not CNOT1 (PMID:24768540, selected abstract). The original U2OS network supplies separate AP-MS edges for CNOT3 and CNOT11; the imaging arm does not supply those physical contacts (PMID:40205054). ANXA2R is the actual Q3ZCQ2 partner in PMID:30833792; it is not a helicase.
One exact source association remains unresolved. PMID:21278420 directly establishes TTP-mediated recruitment and CAF1-associated decay, but the inspected main text does not resolve the preserved ZFP36L1/BRF1 pair. The linked supplemental retrieval returned an error page, which was excluded. Row 20 is UNDECIDED rather than removed or labeled a wrong-gene annotation. Its reference review records this unresolved relationship while retaining the paper’s positive TTP and decay findings.
The complete original receptor study separates ligand-dependent ER/RXR interaction from the absence of the corresponding tested RARα interaction, while RAR-dependent reporter repression is still positive (PMID:16778766, Results and Discussion). The obsolete estrogen-receptor-binding term is refined to current nuclear-receptor binding. The original RAR-binding assertion remains intact but is marked over-annotated at this assay scope; the contextual RAR-signaling repression annotation is retained non-core. This does not assert universal inability to contact a retinoic-acid receptor under all conditions.
P-body and cytosolic localization are supported by human imaging, the mouse donor study and the exact Reactome physical-entity chain, respectively (PMID:21976065; PMID:20133598; Reactome:R-HSA-429955 and R-HSA-6798044). The mouse donor experiments concern gonadal tissue and NANOS2-associated recruitment, not human germ cells. Nuclear localization is consistent with direct promoter/receptor experiments. The tear-proteomics original Table II identifies CNOT1, supporting extracellular detection as non-core without establishing secretion or an extracellular molecular function (PMID:22664934). The exact membrane-proteomics target table for PMID:19946888 was not recovered; that source-specific annotation remains UNDECIDED.
Stem-cell and trophectoderm effects remain contextual. The original study includes mouse ESC and human H1 experiments, with separate CNOT2/3 rescue and H9 scopes; no human embryo experiment is inferred (PMID:22367759). Current exact ortholog and same-term GOA records support the retained transfers where experimental grounding was found. Historical PAINT tree, alignment and IBD placement were not reconstructed, and donor count is not used to reject an inference. Human CNOT1 among its own experimental sources is legitimate grounding.
No additional process annotation is proposed. Existing terms already cover deadenylation, translational repression and miRNA-mediated silencing; direct recruitment/scaffold experiments explain what CNOT1 performs in those processes. The authenticated GO-CAM index contains no target match; absence from the index is not used as evidence for a missing process. Questions remain about natural isoform differences, the exact BRF1 source relationship and the unresolved membrane-proteomics record.
The accompanying source index contains compact raw record identifiers, table entries, source URLs and hashes, plus explicitly identified paraphrases. It is provenance, not independent scientific evidence. Full raw responses, source-text extracts, failed-access receipts and exact read ranges are retained in the temporary audit packet. No figure-pixel inspection is claimed by this owner review. Literal quotation totals are confined to the YAML: 11 words from PMID:16778766, 11 from PMID:22977175 and 12 from PMID:24768540 by whitespace tokenization; a conservative punctuation-based audit is also performed before sealing. No primary quotation is duplicated in this note or the published index.
The earlier failed supplemental retrieval above is superseded by recovery of the original publisher-linked ZIP for PMID:21278420 (DOI:10.1093/nar/gkr011). All four pages of nar-01971-a-2010-File009.pdf were rendered and visually inspected. Table S1 lists oligonucleotides; Table S2 reports GS-TTP purification, including human CNOT1 with 19 peptides versus zero in the GS control (9% coverage), with murine Zfp36 as bait. Figure S1 shows YFP-TTP constructs assayed with Not1, Caf1a or Ccr4a in HEK293 cells. These are positive TTP findings, but the supplement does not identify a BRF1/ZFP36L1 construct or resolve the exact preserved interaction assertion. Row 20 remains UNDECIDED. No source object, supporting entity, annotation action or core function changes. The normal publication cache is unchanged. Original PDF SHA256: 72c82a5ee7010ccc1d9520eba5fd42a9d1733ff0c79552cde669ed2289766f46.
This assessment preserves every original annotation, source qualifier, product, reference identity, selected cache and earlier note. It supersedes the earlier enzyme-binding refinement for the eight CNOT6/CNOT6L source rows (zero-based 16, 17, 25, 28, 31, 43, 48 and 55); the feedback listed seven, but the same assay distinction applies to the eighth source assertion. All eight positive associations remain KEEP_AS_NON_CORE under the campaign policy in projects/CLINGEN_MENDELIAN.md, supported-binding exception. None is collapsed into another source row.
The human GFP-CCR4a/b experiments recover multisubunit complexes (PMID:17452450, Methods and Results, Figure 2). The human NOT1 study separately establishes coimmunoprecipitation of CCR4a/b and direct recombinant binding/structure for CAF1/POP2 (PMID:22977175, Methods, Figures 1–3). CAF1/POP2 connect the CCR4 enzymes to the NOT module. The later exact IntAct records are spoke-expanded anti-tag coimmunoprecipitation associations (PMID:26496610 and PMID:33961781). These positive records do not distinguish a direct CNOT1–CNOT6/6L contact. The original PMID:21984185 full assay remains uninspected; its positive annotation and complex context are retained without manufacturing a negative result. This is an evidence-specific limit on the added partner-class specificity, not a rule that all coimmunoprecipitation evidence is invalid. CNOT7/CNOT8 refinements remain supported by direct recombinant assays.
The single scaffold core now explicitly contributes to poly(A)-specific ribonuclease activity (GO:0004535), consistent with the original contributes_to source qualifier. CNOT1 depletion reduces the activity of CNOT6L-containing immunoprecipitates without reducing the exogenous CNOT6L level (PMID:21976065, Figure 3 and Deadenylase assay Methods). Human reconstitution shows enhanced and more selective poly(A) degradation by the assembled complex (PMID:31320642, Results and Methods). The molecular activity of CNOT1 itself remains protein complex scaffold activity; this does not assign hydrolysis to CNOT1 or create a second core.
The RNA-capture refinements remain. In the original Castello article, UV crosslinking does “not promote protein-protein crosslinking”; the subsequent LiCl/LiDS purification dissociates most noncovalent protein interactions (PMID:22658674, printed pages 1393–1394 and Technical Aspects of the Interactome Capture Protocol). The exact Table S1 CNOT1 entry supports a high-probability mRNA-contact hit, with individual validation still appropriate. This method does not simply capture every member of an RNA-associated complex. The Baltz thesis independently reproduces the CNOT1 target row and describes irradiated/nonirradiated HEK293 controls with 500 mM LiCl and 1% lithium dodecyl sulfate in Methods section 3.5 (PMID:22681889; thesis printed pages 34–35, target table PDF page 144). This extends access to the author’s original thesis methods, not to the full journal article. Neither experiment establishes a cognate sequence, binding domain, equal activity of all products, or a separate RNA-binding core.
The tear annotation remains KEEP_AS_NON_CORE. The original Table II identifies CNOT1 in the merged tear-proteome list (PMID:22664934). Schirmer-strip sampling and detergent extraction leave cellular carryover possible; the study does not identify the cellular source or release mechanism of CNOT1. Detection does not establish secretion or extracellular function, but absence of a signal peptide does not refute detection in an extracellular sample. The current GO:0005576 definition is spatial. No CNOT1-specific artifact or contrary localization experiment was found that warrants labeling this positive source assertion overannotated.
External source access: the selected 21976065, 22658674 and 22681889 caches remain abstract-only and byte-exact. Actual full 21976065 was read at https://doi.org/10.1007/s13238-011-1092-4; the original Castello article and supplement were recovered at https://rnajc.ucsf.edu/sites/rnajc.ucsf.edu/files/CastelloArticlePlusSupplementalJune2012.pdf; the Baltz thesis is https://refubium.fu-berlin.de/bitstream/handle/fub188/9034/Dissertation_Baltz.pdf?sequence=1. The finite source index records hashes and actual read scopes. The previous BRF1 access correction remains intact.
The earlier quotation-scope statement describes the prior version and is superseded here: these notes now contain one short excerpt from the actual external Castello article. Every authored occurrence in the YAML, notes and index is covered by the cumulative source budget; no source exceeds 25 words.
Follow-up validation, calculated status and ordinary rendering passed. The 39 generic-binding advisories follow the explicit campaign exception (projects/CLINGEN_MENDELIAN.md, line 2976 at the checked source main); 31 were inherited and eight correspond to the source-specific association decisions above. DRAFT remains the calculated status. All original source-cache and history bytes remain exact.