Rv2446c Still unknown · low auto-curated

Annotation: from legacy to revised

Auto-curated: this verdict and function were generated by rules from PGAP + Pfam + Foldseek and have not been hand-reviewed.

In the literature (TB corpus sweep) named only in full text

No TB publication mentions this locus tag in its title or abstract (sweep of a ~330k-abstract PubMed TB corpus). A full-text search does however find 3 paper(s) naming this gene in their body text — see the full-text field below. The gene remains dark on function, but it is not unmentioned, and this page no longer claims that it is.

Scope of the sweep: What was actually swept: the H37Rv locus tag and the ortholog locus tags (M. bovis Mb…, M. marinum MMAR_…, M. smegmatis MSMEG_…, M. leprae ML…, M. abscessus MAB_…), plus the gene name where one exists. NOT swept: protein and enzyme names, including family shorthands such as Ldt(Mt4) or VapC28, and their orthographic variants. An absent locus-tag mention is therefore evidence about the TAG, and only indirect evidence about the protein (P18.14).

An antigen status or a virulence phenotype is NOT a molecular function: the verdict stays unchanged. This layer adds the missing context, it does not requalify the gene. This distinguishes a gene that is dark because nobody has looked from one that is dark despite having been studied. Source: PubMed (whole), MULTI-ALIAS sweep of title+abstract: the H37Rv locus tag AND every ortholog identifier (M. bovis Mb…, M. marinum MMAR_…, M. smegmatis MSMEG_…, M. leprae ML…, M. abscessus MAB_…), each hit VERIFIED against the abstract text (word-boundary regex). phase73, 2026-07-13.

Genomic-neighbour overlap (structural caveat) co-directional · 1 % of gene

co-directional overlap: ordinary (e.g. shared stop/start codons in an operon), not the Rv2438A-type artefact P20.1, derived from GFF3 gene coordinates, 2026-08-03.

Binding-pocket screen (P2Rank, geometric prediction) detector blind at this length

Read with care. This protein (123 aa) is below the size where this detector has meaningful power: on proven enzymes, only 3.8% (1/26) under 200 aa reach this layer's P2Rank confidence threshold of best_probability >= 0.70, versus 60.5% (75/124) above it (P16.3b calibration, negative control EsxA/EsxB-scale panel). Both figures are recall AT 0.70, which is also the best separation this detector reaches on that panel (Youden J = 0.416) and whose false-positive rate above 200 aa is 18.9% (81.1% specificity); the looser 0.50 that appears elsewhere in this repository would raise recall above 200 aa to 74.2% and drop specificity to 64.9%. A negative or weak pocket result here should NOT be read as evidence against a ligand-binding role -- the test essentially has no power at this length, not that the protein lacks a site. The same holds in the other direction, and that half was missing here: a STRONG score at this length is not a result either. The same calibration panel does carry that other arm below 200 aa, and the two do not separate: the 3.8% above is matched by 6.6% (5/76) of proteins annotated as non-catalytic clearing 0.70 (Fisher p = 1.00; 15.4% against 11.8% at 0.50, p = 0.73). At this length the score therefore carries no measured information in either direction -- which is a stronger statement than untested power, and the reason a strong score here is not a result. P16.3e, calibration phase72b_pocket_calibration.py, 2026-08-03.

CRISPRi vulnerability

Vulnerability index -0.47 (95% CI -0.61 to -0.31). A more negative index = more vulnerable to knockdown (better drug-target quality); indicative threshold VI ≤ -6 = highly vulnerable.

Quantitative CRISPRi knockdown, graded (finer than binary Tn-seq essentiality). Source: CRISPRi vulnerability index (Bosch 2021, supplementary Table S2 'mmc3.xlsx', bulk import via Europe PMC -- supersedes the per-gene pebble.rockefeller.edu scrape of phase46_crispri_vulnerability.py).

Caveat on this estimate. Bosch et al. flag this gene's vulnerability estimate as NOT certain (their own confidence call, not a p-value threshold). Its guide library did sample a range of repression strengths (str_span=0.682 on a 0-1 scale, thanks to at least one guide <=18 nt; n_guides=39 descriptive only), so the dose-response fit is not starved of titration the way most 'not certain' genes are -- the uncertainty here is the authors' own confidence call on the fit, not a design limitation of the guide library. Do not cite this VI as a standalone vulnerability claim without checking `certain` first.

Orthologues (reciprocal best hits across mycobacteria)

Reciprocal-best-hit orthologues (DIAMOND) against the Mycobrowser reference proteomes. A missing species is informative: e.g. a gene absent from M. leprae was likely lost in its reductive genome evolution. Locus tags link to Mycobrowser.

Curated reference (UniProt)

Functional vocabulary (eggNOG-mapper, orthology transfer)

Orthology-based transfer (eggNOG 5.0.2, diamond). EC/KO/GO/CAZy are computed annotations, not manual curation; cross-check against the primary literature before treating a specific reaction as established.

Conservation & selection (intra-MTBC, 142 601 strains, snapshot of 2026-09-29)

pN/pS from segregating SNPs (singletons removed) normalised by possible sites. Low pN/pS = purifying selection (a strong signal that a "hypothetical" is a real, constrained gene). A high pN/pS is ambiguous: relaxed constraint or positive selection (drug resistance, antigenic variation) inflate it; e.g. rpoB/katG/pncA score high here for resistance, not loss of function. A clonal disruption (one allele over a clade) suggests lineage pseudogenisation; a convergent one (many independent alleles) is typical of resistance loss-of-function.

Lineage-specific relaxation (candidate, one evidence handle) 1 lineage(s)

Candidate lineage-specific loss-of-function signal: rare/absent in the other well-sampled lineages (<5%) but risen in the listed lineage(s). ONE handle, not a verdict change (P8 two-handle policy) -- the general phenomenon of lineage-specific gene loss reflecting ecotype-specific selective pressure is already established at genome scale (Silva-Pereira et al. 2024); this signal on an individual dark gene needs an independent second handle before any requalification.

Caveat: Sample-size heterogeneity across MTBC lineages (up to 4 orders of magnitude, 16 smaller lineages excluded for n<500); essentiality is NOT resolvable per lineage (TnSeq data is H37Rv-only). For the two smallest adequate lineages (L5 n=630, L6 n=1364) the frequency floor bottoms out at the absolute minimum of 3 carrier strains -- a handful of strains from a SINGLE transmission cluster (clonal outbreak) can pass this floor without representing independent evolutionary origins of the disrupting allele; no phylogenetic correction for clonal sampling was applied. Treat low-count hits in these two lineages with extra caution.

Reference: Silva-Pereira TT, Soler-Camargo NC, Sá Guimarães AM 2024, Microbiology Spectrum, doi:10.1128/spectrum.02289-23

Outgroup conservation (beyond the MTBC) Mycobacteriaceae

Two orthogonal outgroup signals. M. canettii (the immediate outgroup) gives a deep-divergence dN/dS (a low value confirms a constrained, real gene; shown as confident only at ≥8 substitutions, else flagged low-power). Genus-wide presence/absence (tblastn vs assembled non-MTBC genomes) places the gene on the ancient-core ↔ MTBC-specific axis: a gene absent even from the closest MTBAP relatives is a candidate MTBC-specific innovation (possible host-adaptation factor, to confirm by synteny). The phylostratum extends that axis outside the genus (tblastn vs 13 reference genomes spanning Mycobacteriaceae → Corynebacteriales → Actinomycetia → outside the phylum): it is the deepest clade in which a homolog is still detected, i.e. a proxy for gene age. Read it with the null model in mind: a shallow (young) stratum can also reflect homology-detection failure for short or fast-evolving ORFs, so it is a descriptive axis, not a proof of novelty.

Essentiality (transposon mutagenesis)

Genome-wide Himar1 transposon essentiality in H37Rv (DeJesus 2017). An essential call (ES/ESD/GD) is strong, independent evidence that a "hypothetical" locus encodes a functional, selectively required gene — orthogonal to intra-species conservation.

Proteomics (mass spectrometry) not detected

Not detected in the mass-spectrometry datasets integrated by PaxDb. This is not evidence of absence, and this atlas can say which bias dominates, because it measured it on its own 482 undetected proteins. Size is the main factor: undetected proteins have a median length of 145 aa against 295 aa for detected ones, and 25.3 % of them are shorter than 100 aa against 6.0 %. Predicted localisation matters far less than usually assumed: detection rates are 89.3 % for globular, 84.9 % for transmembrane, 88.0 % for lipoproteins and 77.8 % for signal-peptide proteins — so a short protein is under-sampled whatever its predicted compartment, and hydrophobicity alone is a weak excuse. Condition-specific expression and few tryptic peptides (PE/PPE repeats) add to this.

Predicted localisation (DeepTMHMM + lipobox)

Transmembrane topology and signal peptide from DeepTMHMM (deep-learning reference predictor); lipoproteins from a (myco)bacterial lipobox motif. A sequence-based prediction of subcellular context.

Physico-chemical properties (computed, ProtParam)

Computed from the ancestral MTBC0 sequence with the ExPASy ProtParam method (Biopython). Descriptive biophysical context: a positive GRAVY flags a hydrophobic (often membrane) protein, a high instability index (>40) predicts a short in-vitro half-life, an extreme pI hints at compartment or binding partner.

Domains (Pfam, hmmscan --cut_ga)

HHpred profile-profile (no confident hit): No confident hit. Remains dark. top hits scattered generic membrane-protein hits, best 68.3%, E=28. Remote homology search on the deep UniRef30 profile (MPI Toolkit), run for the residual dark set.

3D structure (interactive — Mol*/pdbe-molstar)

No experimental structure is deposited for this gene: showing the AlphaFold DB predicted model (mean pLDDT 87.6, confident), coloured by per-residue confidence (dark blue = very high, orange/red = low — treat low-confidence regions as unreliable, especially termini and loops). Drag to rotate, scroll to zoom.

Genomic context (neighbours & predicted operon) operon of 4

Neighbours from the H37Rv annotation (- strand). The operon is predicted by co-directional intergenic distance (same strand, gaps ≤50 bp) — a transcription-unit hypothesis, not a mapped TSS. For a "hypothetical", co-transcription with a characterised operon is a concrete functional lead (complements the STRING neighborhood channel below).

Published operon evidence (two independent sources)

Two independent published operon maps, cited alongside the distance heuristic above: an association corroborating or contradicting the heuristic, never proof of shared function. REMap's RNA-seq coverage continuity is the most direct of the three signals; Roback's logistic regression and the intergenic-distance heuristic are statistical proxies.

Node colour = verdict, dashed = still uncharacterised; edge colour = evidence (green experimental, orange genomic-context, grey co-expression), width ∝ score. Click a partner to open its page; "Explore full network" to walk the graph.

Closest characterised functional partner: folC (folylpolyglutamate synthase FolC), high confidence (score 909, all STRING channels except text-mining; the partner is chosen on its genomic-context evidence). Read this as a lead toward a shared pathway or operon, not as evidence of a physical complex: when this atlas co-folded 98 such context-only pairs, their predicted interfaces were no more confident than those of length-matched control pairs.

Showing the 15 strongest of this gene's 116 partners at medium confidence (36 at high confidence). The table is an extract, not a neighbourhood: across the proteome the real median degree is 34 while the served median is 15, so no hub, degree distribution or centrality may be read on this table. None of the 116 partners rests on an interaction measured in M. tuberculosis itself.

STRING combines evidence channels (neighborhood, fusion, cooccurrence, coexpression, experimental, database, text-mining) into a 0–1000 score. The ctx badge marks edges carried by the genomic-context channels (conserved neighborhood, fusion, phylogenetic co-occurrence), which are independent of orthology and structure and the strongest signal for an unknown gene. The no text-mining column recomputes the score from data alone, so a link that does not depend on the literature is visible. Association is a function hypothesis, not proof: corroborate with the operon context and the primary literature before assigning a function.

Where the experimental evidence comes from, and why it matters here. STRING's experimental channel fuses interactions measured in M. tuberculosis with interactions measured in another organism and transferred to this pair by orthology. The two are not equivalent for this atlas, whose standing rule is that better studied elsewhere is not a function established in M. tuberculosis. Measured on the served network: of the 38 229 partner pairs shown across the atlas, 13 483 partner rows carry a non-zero experimental channel, and only 106 pairs rest on an interaction measured in M. tuberculosis itself — 0.3 % of the network. Those 106 are long-established complexes (ESX-1/ESAT-6, RNA polymerase with CarD and RbpA, the Pup–Mpa proteasome, gyrase, Emb, GarA with PknB and PknG, the Rel modules); no still-uncharacterised gene is among them. The reason is not a gap in this atlas but in the literature: there is almost no experimental interactome of this bacterium, IntAct holding 134 curated protein pairs for a ~4 000-gene proteome. A exp (transferred) badge is therefore a lead, never a measurement, and an absence of badge is not evidence that the pair does not interact.

Evidence

• Legacy H37Rv annotation: integral membrane protein
• MTBC0 PGAP product: DUF4233 domain-containing protein
• Pfam (hmmscan --cut_ga): DUF4233 PF14017.12 (E=8e-33)
• (auto-curated by rules from PGAP + Pfam + Foldseek; not hand-reviewed)

Ancestral MTBC0 protein sequence

>mtbc0_002605|Rv2446c|
MTDRSREPADPWKGFSAVMAATLILEAIVVLLAIPVVDAVGGGLRPASLGYLVGLAVLLILLTGLQRRPWAIWVNLGAQPVLVAGFAVYPGVGFIGVLFAALWVLIAYLRAEVRRRRDYRVSQ