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Hox gene clusters encode transcription factors that drive regional specialization during animal development: e.g. the Hox factor Ubx is expressed in the insect metathoracic (T3) wing appendages and differentiates them from T2 mesothoracic identities. Hox transcriptional regulation requires silencing activities that prevent spurious activation and regulatory crosstalks in the wrong tissues, but this has seldom been studied in insects other than Drosophila, which shows a derived Hox dislocation into two genomic clusters that disjoined Antennapedia (Antp) and Ultrabithorax (Ubx). Here we investigated how Ubx is restricted to the hindwing in butterflies, amidst a contiguous Hox cluster. By analysing Hi-C and ATAC-seq data in the butterfly Junonia coenia, we show that a Topologically Associated Domain (TAD) maintains a hindwing-enriched profile of chromatin opening around Ubx. This TAD is bordered by a Boundary Element (BE) that separates it from a region of joined wing activity around the Antp locus. CRISPR mutational perturbation of this BE releases ectopic Ubx expression in forewings, inducing homeotic clones with hindwing identities. Further mutational interrogation of two non-coding RNA encoding regions and one putative cis-regulatory module within the Ubx TAD cause rare homeotic transformations in both directions, indicating the presence of both activating and repressing chromatin features. We also describe a series of spontaneous forewing homeotic phenotypes obtained in Heliconius butterflies, and discuss their possible mutational basis. By leveraging the extensive wing specialization found in butterflies, our initial exploration of Ubx regulation demonstrates the existence of silencing and insulating sequences that prevent its spurious expression in forewings.
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Ultrabithorax Is a Micromanager of Hindwing Identity in Butterflies and Moths
The forewings and hindwings of butterflies and moths (Lepidoptera) are differentiated from each other, with segment-specific morphologies and color patterns that mediate a wide range of functions in flight, signaling, and protection. The Hox gene Ultrabithorax ( Ubx ) is a master selector gene that differentiates metathoracic from mesothoracic identities across winged insects, and previous work has shown this role extends to at least some of the color patterns from the butterfly hindwing. Here we used CRISPR targeted mutagenesis to generate Ubx loss-of-function somatic mutations in two nymphalid butterflies ( Junonia coenia , Vanessa cardui ) and a pyralid moth ( Plodia interpunctella ). The resulting mosaic clones yielded hindwing-to-forewing transformations, showing Ubx is necessary for specifying many aspects of hindwing-specific identities, including scale morphologies, color patterns, and wing venation and structure. These homeotic phenotypes showed cell-autonomous, sharp transitions between mutant and non-mutant scales, except for clones that encroached into the border ocelli (eyespots) and resulted in composite and non-autonomous effects on eyespot ring determination. In the pyralid moth, homeotic clones converted the folding and depigmented hindwing into rigid and pigmented composites, affected the wing-coupling frenulum, and induced ectopic scent-scales in male androconia. These data confirm Ubx is a master selector of lepidopteran hindwing identity and suggest it acts on many gene regulatory networks involved in wing development and patterning.
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- PAR ID:
- 10232701
- Date Published:
- Journal Name:
- Frontiers in Ecology and Evolution
- Volume:
- 9
- ISSN:
- 2296-701X
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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Hoxgene clusters encode transcription factors that drive regional specialization during animal development: for example the Hox factor Ubx is expressed in the insect metathoracic (T3) wing appendages and differentiates them from T2 mesothoracic identities.Hoxtranscriptional regulation requires silencing activities that prevent spurious activation and regulatory crosstalks in the wrong tissues, but this has seldom been studied in insects other thanDrosophila, which shows a derivedHoxdislocation into two genomic clusters that disjoinedAntennapedia(Antp) andUltrabithorax(Ubx). Here, we investigated howUbxis restricted to the hindwing in butterflies, amidst a contiguousHoxcluster. By analysing Hi-C and ATAC-seq data in the butterflyJunonia coenia, we show that a Topologically Associated Domain (TAD) maintains a hindwing-enriched profile of chromatin opening aroundUbx. This TAD is bordered by a Boundary Element (BE) that separates it from a region of joined wing activity around theAntplocus. CRISPR mutational perturbation of this BE releases ectopicUbxexpression in forewings, inducing homeotic clones with hindwing identities. Further mutational interrogation of two non-coding RNA encoding regions and one putativecis-regulatory module within theUbxTAD cause rare homeotic transformations in both directions, indicating the presence of both activating and repressing chromatin features. We also describe a series of spontaneous forewing homeotic phenotypes obtained inHeliconiusbutterflies, and discuss their possible mutational basis. By leveraging the extensive wing specialization found in butterflies, our initial exploration ofUbxregulation demonstrates the existence of silencing and insulating sequences that prevent its spurious expression in forewings.more » « less
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- Free Publicly Accessible Full Text
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Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.3389/fevo.2021.643661
