mmpL6 Resolved · high auto-curated

Annotation: from legacy to revised

Curation note: 2026-10-02 (mtbc/Rv1557 project, back-propagation P52.2, destination C1, status: unpublished, not peer-reviewed). Disruption signal in two small basal lineages. The `conservation` layer reports no disrupting allele (disrupt_frac 0.0, pseudogene_flag false), but its frequency floor (142 of 142,601 strains) cannot see an allele fixed in a small lineage. The dedicated project checked the variant reports of two neighbouring projects on small basal animal-adapted lineages that carry many fixed loss-of-function variants (M. dassie, 7 strains, 72 fixed frameshift, stop-gained or splice variants genome-wide; M. suricattae, 17 strains, 101): no HIGH-impact variant of this gene is fixed in either lineage. The only variant of the gene among their fixed variants is a missense change, NC_000962.3:1762006 C>A (p.Asn88Lys in the numbering of the 397-aa ORF), in M. suricattae. The atlas per-clade counts agree for the strains held in the local database (M. suricattae 5, M. dassie 6, M. mungi 1, Chimpanzee bacillus 1): no insertion, deletion or nonsense variant in the window; N88K is carried by 5 of 5 M. suricattae and by none of the 6 M. dassie, and two other missense variants are polymorphic in M. dassie (2 and 1 of 6 strains). Limits: the window is the 397-aa ORF of H37Rv, the C-terminal 41% of the 967-aa full-length protein; a loss-of-function event in the 570 N-terminal residues cannot be called against H37Rv and is not covered by these reports, and absence from a list of fixed HIGH-impact variants is not proof of an intact reading frame. The two neighbouring reports are internal and not peer-reviewed. This shows no sign of disruption in the C-terminal window of these lineages; it does not bear on the 'diversifying/relaxed' label of the layer, which rests on other lineages. 2026-10-02 (mtbc/Rv1557 project, back-propagation P52.2, destination C1, status: in preparation, not peer-reviewed). Two readings of the polymorphism of this gene use different strain sets. This 397-aa ORF is the C-terminal part of MmpL6: in M. bovis AF2122/97 (TbD1 intact) the protein has 967 aa, and the 397 aa served here match its residues 571 to 967 (15 of the first 16 residues differ, at the TbD1 junction), the 570 N-terminal residues being deleted in H37Rv. The layer pools 142,601 strains and counts 8 variable sites carried by at least 142 strains (0.1%; pN/pS 2.451, 'diversifying/relaxed'). In the atlas per-clade counts (snapshot 2026-09-29), 7 of these 8 sites are carried almost exclusively by L4 (4 sites), L2 (2) or L3 (1), lineages that lack TbD1 and hold only this remnant (101,472 of the 142,601 strains); the eighth (NC_000962.3:1762409 A>G, 148 strains) is in M. bovis. The label therefore mostly describes the remnant, and it rests on 8 sites, the minimum of the layer's power gate. The dedicated project instead scanned the same window in 733 strains of TbD1-intact lineages (M. canettii, L5, L6, L7, basal and derived animal-adapted lineages; scan made before its later additions of M. microti, M. pinnipedii, M. suricattae, L1, L9 and L10) from the production SPDI files: 37 distinct variants (20 missense, 13 synonymous, 3 deletions, 1 complex change). No site is fixed in one of the two groups defined by residue 551 (derived K, 486 strains; ancestral N, 247 strains) and absent from the other (Dn = Ds = 0), so a classical McDonald-Kreitman test cannot be run. The K group is nearly monomorphic (9 sites, 8 of them seen in a single strain: 7 missense, 2 synonymous); the N group has 24 sites (13 missense, 11 synonymous), but its three commonest variants (40%, 34% and 11% of the N strains) are each confined to one clade, M. canettii for the first two and L7 for the third, so this is clade structure and not homogeneous polymorphism. A site-based Fisher test of missense against synonymous sites, K against N, gives OR 2.96, p = 0.26 (not significant, low power). The project reads the near-monomorphism of the K group as compatible with constraint on this C-terminal domain; this is an interpretation of a negative result, not a demonstrated purifying selection, and it is not peer-reviewed. The `conservation` call is kept: the two analyses use different strains, and neither is a test of selection on the full-length protein.

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

Annotated on the H37Rv protein: this gene has no 1:1 ancestral MTBC0 anchor (PE/PPE, paralogue, IS element, or otherwise unanchored CDS).

In the literature (TB corpus sweep) 6 publications

6 TB publications mention this gene. 6 publications discuss this gene (count verified against title and abstract text). Queried separately, 6 records carry a M. tuberculosis context and none a non-TB mycobacterial context. The three counts come from three distinct PubMed queries (the total is text-verified, the context counts are raw query volumes), so they neither partition the total nor need to add up to it.

This layer CITES the literature and adds context; it does not change the verdict or the function stated elsewhere in this fiche. This distinguishes a gene that is dark because nobody has looked from one that is dark despite having been studied. Source: PubMed (whole): H37Rv locus tag + GENE NAME + ortholog identifiers (Mb…, MMAR_…, MSMEG_…, ML…, MAB_…), under a mycobacterial context filter; hits verified against the abstract text. Species-context counts distinguish M. tuberculosis literature from literature on other mycobacteria. phase76/phase77, 2026-07-13.

CRISPRi vulnerability

Vulnerability index 0.95 (95% CI -1.13 to 4.00). 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, and here that call has a mechanical cause visible in the library itself: every guide targeting this gene is >=21 nt (str_span=0.298 on a 0-1 scale, near the genome floor), and guide repression strength saturates from 21 nt on -- so the library never sampled a partial knockdown for this gene, only the near-maximal repression plateau. The dose-response fit (and this VI) is therefore essentially unconstrained by data, not merely uncertain -- a limitation of the CRISPRi library design for the non-essential class (Bosch et al. 2021), not a defect in this specific measurement. n_guides=15 is descriptive only and does not predict this. Do not cite this VI as a standalone vulnerability claim.

Legacy record & comparison (Mycobrowser)

The legacy Mycobrowser record is shown for verification. Mycobrowser is no longer maintained; its EC numbers predate recent nomenclature revisions, so a class change usually reflects re-numbering, not a conflict.

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.

Outgroup conservation (beyond the MTBC) Corynebacteriales

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) detected

Detection by mass spectrometry is direct, experimental evidence that the protein product exists — orthogonal to sequence conservation and to Tn-seq essentiality, and especially decisive for a "hypothetical" locus. Read the dataset count as abundance, not as replication. Measured across this atlas (3,491 detected proteins), the number of datasets a protein appears in tracks its abundance almost monotonically: median 1.2 ppm at 1 dataset of 16, 27 ppm at 10, 392 ppm at 15, 1,325 ppm at 16 — three orders of magnitude. A high count therefore says the protein is abundant more than it says several independent experiments confirmed it. What the count does establish, at any value, is that the product exists.

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)

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

No experimental structure is deposited for this gene: showing the AlphaFold DB predicted model (mean pLDDT 78.5, 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.

Structural search (AlphaFold DB model, Foldseek vs PDB — genome-wide) pLDDT 78.5

Foldseek search of the AlphaFold DB model (mean pLDDT 78.5, gated at 70) against the PDB — a genome-wide extension of the ESMFold dark-gene search that also covers proteins beyond the single-sequence length limit. Confident structural neighbours (E < 0.01) shown.

Genomic context (neighbours & predicted operon) operon of 5

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.

Transcriptional regulation (signed TRN: ChIP-seq + TFOE)

Regulatory edges from the ISB signed transcriptional regulatory network (TF ChIP-seq binding, Minch 2015 + TF-overexpression response, Rustad 2014). An edge is regulatory evidence (binding and/or expression change), not necessarily direct. For a "hypothetical", membership in a known regulon (e.g. DosR dormancy, PhoP virulence) is a physiological-context lead, not a function.

Condition: Condition-blind by construction. These edges come from transcription-factor OVEREXPRESSION in a single laboratory condition, so a listed regulator is not a claim that it governs this gene in every circumstance. Measured counter-examples exist within this atlas (see Rv2516c, where the DosR regulon collapses under moxifloxacin while the gene itself rises). Read a regulator as 'capable of shifting this gene', and look for the condition before building a hypothesis on it.

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: Rv1558 (hypothetical protein), high confidence (score 868, all STRING channels except text-mining; the partner is chosen on its genomic-context evidence).

Showing the 15 strongest of this gene's 19 partners at medium confidence (6 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 19 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

• Annotation from H37Rv (no MTBC0 1:1 anchor; H37Rv protein used): transmembrane transport protein MmpL6
• Pfam (hmmscan --cut_ga): MMPL PF03176.22 (E=9e-105)
• (auto-curated by rules from PGAP + Pfam + Foldseek; not hand-reviewed)

Ancestral MTBC0 protein sequence

>H37Rv|Rv1557|mmpL6
MQGISVTGLVKRGWMVRSVFDTIDGIDQLGEQLASVTVTLDKLAAIQPQLVALLPDEIASQQINRELALANYATMSGIYAQTAALIENAAAMGQAFDAAKNDDSFYLPPEAFDNPDFQRGLKLFLSADGKAARMIISHEGDPATPEGISHIDAIKQAAHEAVKGTPMAGAGIYLAGTAATFKDIQDGATYDLLIAGIAALSLILLIMMIITRSLVAALVIVGTVALSLGASFGLSVLVWQHLLGIQLYWIVLALAVILLLAVGSDYNLLLISRFKEEIGAGLNTGIIRAMAGTGGVVTAAGLVFAATMSSFVFSDLRVLGQIGTTIGLGLLFDTLVVRAFMTPSIAVLLGRWFWWPQRVRPRPASRMLRPYGPRPVVRELLLREGNDDPRTQVATHR