The HOXB-AS3 peptide is a 53-amino-acid microprotein translated from a small open reading frame in HOXB-AS3, a transcript annotated as a long non-coding RNA that is antisense to HOXB5 and HOXB6 in the HOXB cluster on chromosome 17q21.32. The open reading frame is conserved among primates. The peptide is predicted to be entirely disordered, has no transmembrane segment and no known homologs outside its own Pfam family. It has been detected endogenously by antibody and mass spectrometry. In colon cancer cells it binds the arginine residues of the RGG motif of the splicing regulator hnRNP A1 and competitively blocks hnRNP A1 binding to the intronic sequences flanking pyruvate kinase M (PKM) exon 9. This antagonizes hnRNP A1-dependent alternative splicing of PKM, lowers production of the PKM2 isoform and restrains aerobic glycolysis. In these cells the peptide, not the RNA, suppresses tumour growth, and low peptide levels are associated with poor prognosis. The peptide is also reported to reduce hnRNP A1-dependent processing of miR-18a. A separate study in oral squamous carcinoma cell lines found that the peptide co-immunoprecipitates with IGF2BP2 and sustains c-Myc levels and proliferation. Most other published work on HOXB-AS3 concerns the RNA rather than the peptide.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005515 protein binding | IPI PMID:28985503 A Peptide Encoded by a Putative lncRNA HOXB-AS3 Suppresses C... | MODIFY | Summary: Interaction with hnRNP A1 (HNRNPA1, P09651). The peptide binds the arginine residues of the hnRNP A1 RGG motif and in doing so blocks hnRNP A1 binding to its RNA targets flanking PKM exon 9. This is the central mechanistic finding for the peptide. Reason: Bare protein binding is uninformative. The interaction is described as competitive inhibition of an activity of hnRNP A1 (its binding to PKM pre-mRNA), so the informative MF is GO:0140678 molecular function inhibitor activity. hnRNP A1 is an RNA-binding splicing regulator, not an enzyme, transporter or transcription factor, so none of the more specific inhibitor children fit. Only the abstract of the Mol Cell paper is available; the mechanism is stated there explicitly. Proposed replacements: molecular function inhibitor activity Supporting Evidence: PMID:28985503 the HOXB-AS3 peptide competitively binds to the ariginine residues in RGG motif of hnRNP A1 and antagonizes the hnRNP A1-mediated regulation of pyruvate kinase M (PKM) splicing by blocking the binding of the ariginine residues in RGG motif of hnRNP A1 to the sequences flanking PKM exon 9 |
| GO:0005515 protein binding | IPI PMID:34457052 A micro-peptide encoded by HOXB-AS3 promotes the proliferati... | REMOVE | Summary: Interaction with IGF2BP2 (Q9Y6M1), supported by a single co-immunoprecipitation of overexpressed Flag-tagged HOXB-AS3 peptide from 293T cells. Despite the title's "directly binding", no direct binding assay, binding-site mapping or binding-deficient mutant was reported. Reason: Bare protein binding carries no functional information. The data do not support a more informative MF: there is one co-IP of an overexpressed tagged peptide, and nothing shows that the peptide changes IGF2BP2 activity, since IGF2BP2 levels were unchanged and c-Myc mRNA stability was never measured. The interaction is recorded in the description. Supporting Evidence: PMID:34457052 Next Flag-tagged HOXB-AS3 was transfected into the 293T cell line and was then subjected to a Co-IP assay. IGF2BP2 was detected and the results indicated that HOXB-AS3 binds with IGF2BP2 as a whole complex |
| GO:0043484 regulation of RNA splicing | IMP PMID:28985503 A Peptide Encoded by a Putative lncRNA HOXB-AS3 Suppresses C... | MODIFY | Summary: The peptide antagonizes hnRNP A1-mediated PKM splicing. hnRNP A1 binding next to exon 9 favours the exon 10-containing PKM2 isoform; by blocking that binding the peptide shifts splicing away from PKM2. Reason: Correct in substance but too general. The process affected is a mutually exclusive alternative exon choice in a nuclear pre-mRNA (PKM exon 9 versus exon 10), carried out by the spliceosome under the control of hnRNP A1. GO:0000381 regulation of alternative mRNA splicing, via spliceosome describes it exactly. A "negative regulation" term is not appropriate because the peptide changes which isoform is made rather than reducing splicing overall. This is the best-supported process for the peptide and is its core function. Proposed replacements: regulation of alternative mRNA splicing, via spliceosome Supporting Evidence: PMID:28985503 antagonizes the hnRNP A1-mediated regulation of pyruvate kinase M (PKM) splicing by blocking the binding of the ariginine residues in RGG motif of hnRNP A1 to the sequences flanking PKM exon 9, ensuring the formation of lower PKM2 |
| GO:0050821 protein stabilization | IMP PMID:34457052 A micro-peptide encoded by HOXB-AS3 promotes the proliferati... | REMOVE | Summary: Based on the OSCC study, where knockdown of HOXB-AS3 lowered c-Myc protein and re-expression of the ORF restored it. The authors attribute this to IGF2BP2-dependent stabilization of c-Myc mRNA, not of c-Myc protein. Reason: The full text contains no protein-stability experiment: no cycloheximide chase, no half-life measurement, no proteasome inhibition. The authors do not claim that the peptide stabilizes c-Myc protein. Their model is mRNA stabilization through the m6A reader IGF2BP2, and they did not measure mRNA decay either. The measured result is a change in c-Myc protein level after knockdown or re-expression, which could arise at any step of expression. Protein stabilization therefore misstates the paper, and the evidence is too thin to support a replacement such as GO:0043488 regulation of mRNA stability. Supporting Evidence: PMID:34457052 IGF2BP2 is a well-studied m6A reader stabilizing c-Myc mRNA (14); therefore, the protein expression level of c-Myc was analyzed in cells transfected with sh-HOXB-AS3-1 and −2 PMID:34457052 We found that HOXB-AS3 overexpression promoted the proliferation of OSCC by stabilizing c-myc mRNA by directly binding to IGF2BP2. |
| GO:2000632 negative regulation of pre-miRNA processing | IDA PMID:28985503 A Peptide Encoded by a Putative lncRNA HOXB-AS3 Suppresses C... | UNDECIDED | Summary: UniProt reports that the peptide suppresses hnRNP A1-mediated processing of miR-18a. This result is not in the abstract, and the full text of the Mol Cell paper was not available for this review. Reason: The underlying biology is plausible: the peptide binds the hnRNP A1 RGG motif, and hnRNP A1 is an auxiliary factor for miR-18a biogenesis. However, hnRNP A1 acts on the primary transcript (pri-miR-18a) before Drosha cleavage, not on the pre-miRNA Dicer step. If the paper measured the pri- to pre-miR-18a conversion, GO:2000635 negative regulation of primary miRNA processing would be the accurate term. Without the full text I cannot tell which step was assayed, so the curator's choice is neither confirmed nor overruled. In any case this is a secondary consequence of hnRNP A1 inhibition, not a core function. Supporting Evidence: PMID:17558416 Here we show that hnRNP A1 binds specifically to the primary RNA sequence pri-miR-18a before Drosha processing. |
Loading supporting content…
Download this section (compressed HTML)Q: In PMID:28985503, which step of miR-18a biogenesis did the peptide inhibit: Drosha cleavage of pri-miR-18a, which is where hnRNP A1 acts, or Dicer processing of pre-miR-18a? The answer decides whether the GOA row should be GO:2000635 (primary miRNA processing) rather than GO:2000632 (pre-miRNA processing).
Q: Where in the cell is the endogenous HOXB-AS3 peptide located? Antagonizing hnRNP A1 on PKM pre-mRNA implies the nucleus, but no cellular component annotation exists and the available abstract does not report localization.
Q: Is the "HOXB-AS3-32aa" peptide described in COPD bronchial epithelial cells (PMID:40417409) a separate ORF from the 53-aa C0HLZ6 chain? If so, does it exist endogenously? Its expression was measured only by qRT-PCR.
Q: How can the same peptide be tumour-suppressive in colon cancer (low levels associated with poor prognosis) and pro-proliferative in oral squamous carcinoma? Do the two studies measure the same endogenous product with equivalent reagents?
Experiment: Peptide-specific loss of function that leaves the RNA intact, such as a start-codon point mutation at the endogenous locus. Then measure PKM exon 9/10 usage, hnRNP A1 CLIP occupancy at PKM introns, and pri-, pre- and mature miR-18a levels, separating the peptide's effects from those of the lncRNA.
Experiment: Test binding of recombinant peptide to IGF2BP2 directly, for example by ITC or pull-down with purified proteins, and measure c-Myc mRNA decay after actinomycin D in peptide-depleted and rescued OSCC cells. This would establish whether the peptide regulates mRNA stability at all.
Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)