Loquacious (loqs) encodes a family of double-stranded RNA-binding proteins related to mammalian TRBP and PACT that act as cofactors of the two Drosophila Dicer ribonucleases. The 464-residue isoform F (X2J5X6) is identical to the miRNA-pathway isoform PB except for the absence of one lysine (PB Lys384) in the linker before the third dsRNA-binding domain (DRBM). Like PB, it carries three DRBMs: DRBM1 and DRBM2 bind RNA, DRBM2 binds the pre-miRNA hairpin, and DRBM3 with its preceding linker binds Dicer-1. In the cytoplasm Loqs-PB binds Dicer-1, raises its affinity for pre-miRNAs and stimulates and directs their cleavage to mature miRNAs. Loqs-PB also associates with Dicer-1 and AGO1 in a miRNA RISC-loading complex, although loqs-null flies still assemble miRISC. A different isoform, Loqs-PD, which lacks DRBM3 and has its own C-terminus, partners Dicer-2 in endogenous and exogenous siRNA production. Through miRNA production the gene is required for female germline stem-cell maintenance, fertility and normal development.
Summary: True but too general; the supported activity is double-stranded RNA binding by the DRBMs, a term this accession already carries.
Reason: Loqs-PB binds pre-miRNA through DRBM2 (PMID:17666393), and X2J5X6 keeps all three DRBMs and the pre-miRNA-binding residues 308..309 unchanged. RNA binding is therefore correct but less specific than the evidence supports; refine it to GO:0003725 double-stranded RNA binding, which the gene already carries (ISS row).
ISS PMID:10908586 The drosophila melanogaster genome: translation factors and ...
ACCEPT
Summary: Core activity. DRBM2 of Loqs-PB binds the pre-miRNA hairpin, and all three DRBMs are intact in this isoform.
Reason: The 2000 genome-survey ISS assignment came from DRBM content. Later biochemistry confirms it directly for PB, where DRBM2 binds pre-miRNA and a DRBM2 double mutation (308..309) abolishes pre-miRNA binding (PMID:17666393). X2J5X6 is identical to PB across all three DRBMs. Loqs-PB thus provides the dsRNA-binding (cofactor) contribution to Dcr-1 processing.
IPI PMID:17666393 Functional anatomy of the Drosophila microRNA-generating enz...
MODIFY
Summary: The IPI with Dcr-1 is real, but bare protein binding is uninformative. The same paper shows that the binding increases Dcr-1 affinity for pre-miRNA and miRNA output, which is ribonuclease activator activity.
Reason: Loqs-PB binds Dcr-1 (FBgn0039016) through DRBM3 and the linker before it, and this binding raises Dcr-1 affinity for pre-miRNA and stimulates miRNA production (PMID:17666393, PMID:15985611, PMID:15918769, PMID:17928574). GO:0170054, defined as binding to and increasing the activity of a ribonuclease, describes that function exactly. The interaction itself is not in doubt; per the protein-binding policy the bare term is replaced. One caveat applies to this accession. PF lacks PB Lys384, which lies in that Dcr-1-binding linker two residues after the LLKL motif whose mutation strongly reduces Dcr-1 activity. No experiment has tested whether the missing lysine matters, so the transfer to PF is by sequence identity.
Summary: Loqs is predominantly cytoplasmic, which is where Dcr-1 processes exported pre-miRNAs.
Reason: Immunostaining in S2 cells shows Loqs mainly in the cytoplasm (PMID:15918769), consistent with its role after nuclear Drosha/Pasha cleavage and export. This is as specific as the evidence allows.
IMP PMID:18634873 RAX is required for fly neuronal development and mouse embry...
KEEP AS NON CORE
Summary: Embryos homozygous for the loqs (dRax) insertion f00791 have abnormal CNS commissural axons; this is a downstream developmental requirement.
Reason: The insertion is a gene-level allele (the same f00791 used in PMID:15918770) and disrupts miRNA biogenesis, so the CNS phenotype most likely reflects loss of miRNA-dependent regulation. It is a credible curator IMP but not a direct activity of the protein, and no experiment isolates isoform PF. Retained as non-core.
Mutant fly embryos homozygous for the insertion dRax[f00791] display highly abnormal commissural axon structure of the CNS
GO:0009047 dosage compensation by hyperactivation of X chromosome
IGI PMID:22554892 A role for siRNA in X-chromosome dosage compensation in Dros...
MARK AS OVER ANNOTATED
Summary: The evidence is a viability modifier (loqs f00791 lowers survival of roX1 roX2 males), which does not show that Loqs does any step of X-chromosome hyperactivation.
Reason: The study, read in full text, scores adult male survival in a sensitized roX mutant background; the loqs allele removes all isoforms, including the miRNA-pathway PB, and the authors list both miRNA and endo-siRNA roles of loqs. The phenotype establishes that the small-RNA pathways are needed in a sensitized background, a necessity argument. It identifies no step of MSL recruitment or X hyperactivation that Loqs performs, and the siRNA-related mechanism belongs to the PD isoform rather than this PB-like product. The annotation is not wrong as a genetic observation but overstates participation.
While loqs has a prominent role in miRNA biogenesis, an isoform of Loqs has been implicated in the biogenesis of endo-siRNA from structured loci and transposons
Summary: True but too general. Loqs regulates gene expression through pre-miRNA processing, which the gene already carries and which is a descendant of this term.
Reason: Loqs-PB is required for miRNA-directed silencing because it is needed for Dcr-1 processing of pre-miRNAs (PMID:15918769, PMID:15918770). GO:0031054 pre-miRNA processing is a descendant of regulation of gene expression via regulatory ncRNA-mediated gene silencing, and it is the specific supported process. Refine to it.
IDA PMID:20037596 Loqs and R2D2 act sequentially in the siRNA pathway in Droso...
MODIFY
Summary: Correct but broad; the cited study itself places Loqs-PB at the Dcr-1 pre-miRNA processing step.
Reason: In Marques et al., a stronger loqs mutant reduces miRNA levels, and the authors place Loqs-PB at processing of miRNA precursors by Dcr-1, not at Ago1 loading. Pre-miRNA processing (GO:0031054, already on the gene) is the specific supported process.
IDA PMID:20037596 Loqs and R2D2 act sequentially in the siRNA pathway in Droso...
UNDECIDED
Summary: Valid for the loqs gene through the Dcr-2 partner isoform Loqs-PD. Not established for this PB-like isoform F accession.
Reason: Marques et al. show Loqs is needed for Dcr-2 dsRNA processing, and later work assigns that role to Loqs-PD, which arises by alternative polyadenylation, lacks DRBM3 and has a unique C-terminus (PMID:19644447, PMID:19635780). X2J5X6 has the PB architecture (DRBM3 and the Dcr-1 binding C-terminus), and PB is the isoform that functions in miRNA biogenesis. The experiment is sound at gene level and is not disputed. Whether PF contributes to siRNA processing has not been tested, so the row cannot be accepted or rejected for this specific product.
their biogenesis depends on a new isoform of the Drosophila TRBP homologue loquacious, which arises by alternative polyadenylation and is distinct from the one that functions during the biogenesis of miRNAs
IMP PMID:18463630 The Drosophila hairpin RNA pathway generates endogenous shor...
UNDECIDED
Summary: Valid for the loqs gene through Loqs-PD. Not established for this PB-like isoform F accession.
Reason: Okamura et al. show that hairpin-derived endo-siRNAs depend on loqs. Isoform resolution (PMID:19635780, PMID:19644447) assigns endo-siRNA production to Loqs-PD, distinct from the miRNA-pathway PB, which X2J5X6 matches. The gene-level IMP is not disputed, but the row cannot be confirmed for this product.
IMP PMID:19644447 Endo-siRNAs depend on a new isoform of loquacious and target...
UNDECIDED
Summary: This paper itself identifies the endo-siRNA isoform as distinct from the miRNA isoform that X2J5X6 resembles.
Reason: Hartig et al. show that endo-siRNA biogenesis depends on a new loqs isoform from alternative polyadenylation (PD) and distinguish it from the miRNA isoform. X2J5X6 is PB-like, keeping DRBM3 and the PB C-terminus. The gene-level conclusion holds, but this product is not shown to take part.
their biogenesis depends on a new isoform of the Drosophila TRBP homologue loquacious, which arises by alternative polyadenylation and is distinct from the one that functions during the biogenesis of miRNAs
GO:0030718 germ-line stem cell population maintenance
IMP PMID:17320391 The miRNA pathway intrinsically controls self-renewal of Dro...
KEEP AS NON CORE
Summary: loqs-null germline stem cells are lost, and the defect is rescued by Loqs-PB. This is a downstream requirement of miRNA production.
Reason: The knockout and PB-specific rescue (PMID:17320391, with PMID:15918770 reporting the same GSC maintenance defect) show that the PB-type product is needed intrinsically in GSCs. The process term is well supported at gene level, but it reflects the regulatory output of miRNAs rather than a step Loqs itself performs. Retained as non-core.
IDA PMID:17666393 Functional anatomy of the Drosophila microRNA-generating enz...
ACCEPT
Summary: Core process. Loqs-PB binds the pre-miRNA and Dcr-1 and enhances Dcr-1 affinity for its substrate, contributing directly to the processing step.
Reason: Loqs does part of the work here. It is not the nuclease, but it supplies the dsRNA-binding and substrate-positioning contribution to cleavage (domain dissection and in vitro processing, PMID:17666393). X2J5X6 is PB except for Lys384, so transfer to this accession rests on sequence identity.
IMP PMID:15918770 Normal microRNA maturation and germ-line stem cell maintenan...
ACCEPT
Summary: Core process. The loqs mutant accumulates pre-miRNAs, and Loqs associates with Dcr-1.
Reason: Förstemann et al. show that normal Dcr-1 processing of pre-miRNAs needs Loqs. The IMP is consistent with the biochemical evidence for direct participation.
IMP PMID:19644447 Endo-siRNAs depend on a new isoform of loquacious and target...
ACCEPT
Summary: Core process. Hartig et al. separate the miRNA-pathway isoform from the endo-siRNA isoform PD; the miRNA function is the PB-type function that X2J5X6 shares.
Reason: Consistent with the other pre-miRNA processing evidence. The paper attributes miRNA biogenesis to the non-PD isoform.
their biogenesis depends on a new isoform of the Drosophila TRBP homologue loquacious, which arises by alternative polyadenylation and is distinct from the one that functions during the biogenesis of miRNAs
Summary: Independently supported by loqs female sterility and GSC loss, but it is a downstream requirement of miRNA production.
Reason: The ARBA row is not evidence in itself. Primary loss-of-function data (sterility and GSC loss rescued by PB) support a requirement in oogenesis. As with the GSC rows, this is an organismal consequence rather than the core activity.
Summary: Loqs-PB co-purifies with Dcr-1 in a functional pre-miRNA processing complex that also loads AGO1 (the miRLC). Membership holds even though Loqs is not required for miRISC assembly.
Reason: GO:0070578 describes the Dicer/dsRBP/Argonaute assembly that processes the precursor and loads RISC. Immunopurified Dcr-1/Loqs complexes process pre-miRNA (PMID:15918769), and the curated UniProt PB entry records Loqs-PB in the miRLC with Dcr-1 and AGO1. Liu et al. 2007 (PMID:17928574) found that loqs-null flies still assemble miRISC, so Loqs is a member of the complex but not essential for the loading step. The complex term is kept and no RISC-assembly process is asserted. The mammalian DICER1/TARBP2/AGO2 composition is not transferred (TARBP2 is in a different PANTHER subfamily, SF1).
It is uncertain if Dcr-1 and Loqs facilitate miRNA loading onto the miRISC complexes.
GO:0070920 regulation of regulatory ncRNA processing
IDA PMID:21419681 Phosphate and R2D2 restrict the substrate specificity of Dic...
UNDECIDED
Summary: The biochemistry behind this row is Loqs-PD lowering the KM of Dicer-2 for long dsRNA. That is a PD-specific activity, not shown for this PB-like isoform.
Reason: In Cenik et al. (full text), recombinant Loqs-PD lowers the KM of Dcr-2 for long dsRNA about 10-fold without changing kcat, and it does not inhibit Dcr-2 cleavage of pre-miRNA. The only PB statement in the paper argues against PB restricting Dcr-1 to the miRNA pathway. The regulatory effect is real for the gene but was measured with PD, which lacks DRBM3. X2J5X6 is PB-like, so the row is left undecided for this accession rather than removed.
Endo-siRNA production depends on a specific Loqs isoform, Loqs-PD, which is distinct from the one, Loqs-PB, required for the production of microRNAs.
Core Functions
Binds the double-stranded stem of pre-miRNA hairpins through DRBM2 (with DRBM1), supplying substrate recognition for Dcr-1 cleavage of pre-miRNAs into mature miRNAs in the cytoplasm.
All three DRBMs, the pre-miRNA-binding residues 308..309 and the Dcr-1-binding C-terminus are intact.
Binds Dicer-1 through DRBM3 and its preceding linker and increases Dcr-1 affinity for pre-miRNA and miRNA output, acting as an activating cofactor of the RNase III enzyme within the Dcr-1/Loqs-PB (miRNA RISC-loading) complex.
Q: Is the PF isoform (X2J5X6), which lacks PB Lys384 in the Dcr-1-binding linker before DRBM3, expressed at the protein level, and does it bind and stimulate Dcr-1 as well as PB?
Q: Can PB-type Loqs isoforms (PB/PF) contribute to Dcr-2-dependent siRNA production in any tissue, or is that role exclusive to Loqs-PD?
Suggested Experiments
Experiment: Compare recombinant Loqs-PB and Loqs-PF for Dcr-1 binding (pull-down or ITC) and stimulation of pre-let-7 processing, and test PF transgene rescue of loqs-null germline stem-cell loss.
External Prediction Reviews
These computational predictions are reviewed separately from the GOA annotation set used for this review. The assessments below are from this project and do not constitute official GO annotations or endorsement by GO/UniProt. They are not included in the existing annotation review above.
Broad regulation of gene expression is supported by conserved miRNA-processing biology, but is less precise than the existing pre-miRNA-processing annotations.
Review rationale: Loqs-PF retains all three domains of the experimentally characterized PB miRNA-processing cofactor, differing by one linker-region lysine. Primary experiments demonstrate Loqs-dependent regulation of miRNA-target expression. Existing annotations include the more informative pre-miRNA-processing function, as well as the broad predicted term itself. LSP therefore reflects a correct but less precise description; ARBA agreement is not treated as validation. The mammalian pathway wording in the separate narrative prediction does not invalidate this broad GO claim.
Supporting Evidence:
PMID:15918769: "These results support a model in which Loquacious mediates miRNA biogenesis and, thereby, the expression of genes regulated by miRNAs."
loquacious (loqs) encodes a double-stranded RNA binding protein with multiple splicing isoforms. loqs-PB and loqs-PA associate with the product of Dcr-1 to facilitate processing of precursor miRNA into mature miRNA, whereas loqs-PD interacts with the product of Dcr-2 to enhance processing of dsRNA into siRNA.