Abstract
In most tetrapod vertebrates, limb skeletal progenitors condense with postaxial dominance. Posterior elements (such as ulna and fibula) appear prior to their anterior counterparts (radius and tibia), followed by digit-appearance order with continuing postaxial polarity. The only exceptions are urodele amphibians (salamanders), whose limb elements develop with preaxial polarity and who are also notable for their unique ability to regenerate complete limbs as adults. The mechanistic basis for this preaxial dominance has remained an enigma and has even been proposed to relate to the acquisition of novel genes involved in regeneration. However, recent fossil evidence suggests that preaxial polarity represents an ancestral rather than derived state. Here, we report that 5′Hoxd (Hoxd11-d13) gene deletion in mouse is atavistic and uncovers an underlying preaxial polarity in mammalian limb formation. We demonstrate this shift from postaxial to preaxial dominance in mouse results from excess Gli3 repressor (Gli3R) activity due to the loss of 5′Hoxd-Gli3 antagonism and is associated with cell-cycle changes promoting precocious cell-cycle exit in the anterior limb bud. We further show that Gli3 knockdown in axolotl results in a shift to postaxial dominant limb skeleton formation, as well as expanded paddle-shaped limb-bud morphology and ensuing polydactyly. Evolutionary changes in Gli3R activity level, which also played a key role in the fin-to-limb transition, appear to be fundamental to the shift from preaxial to postaxial polarity in formation of the tetrapod limb skeleton.
| Original language | English |
|---|---|
| Pages (from-to) | 4923-4934.e5 |
| Journal | Current Biology |
| Volume | 31 |
| Issue number | 22 |
| DOIs | |
| State | Published - Nov 22 2021 |
Bibliographical note
Funding Information:We thank Denis Duboule for generously providing the 5′HoxdDel(RXII) (d11-d13 deleted) mutant line, Deneen Wellik for the Hoxd11 mutant line, Steve Vokes for both RosaGrem1 and RosaGli3R transgenic lines, Trevor Williams for the AP2Cre transgenic line, Alan Godwin for help in generating the DSRed2 cassette used to create the Hoxd13 targeting construct, Matt Anderson for advice on HCR protocols, Rich Hwang for advice on tadpole analysis, and members of the CDBL for critical discussion. This research was supported by the Center for Cancer Research (S.M. Intramural Research Program) and the Frederick National Laboratory (W.F.H. and V.M. OMAL Contract no. 75N91019D00024), NCI, NIH; by the Office of Infrastructure Programs, NIH (S.R.V. and M.K. P40-OD019794); and by grants from OHSU Medical Research Foundation and Shriners Hospitals for Children (H.S.S.). S.M. A.T. and B.-L.H. designed the project. S.M. wrote the original paper draft, and S.M. B.-L.H. J.Z. and S.R.V. contributed to text editing and revisions. A.T. B.-L.H. J.Z. M.K. N.W.A.H.B. and S.R.V. performed the experiments; Y.S. and Y.-B.S. assisted with Xenopus; and M.K. and N.W.A.H.B. assisted with axolotl larval collection and processing for analyses. S.R.V. performed axolotl microinjections. W.F.H. and V.M. provided technical advice and assistance with live-imaging data collection and analysis. H.S.S. and B.T. generated key genetically engineered mice used for analyses. The authors declare no competing interests.
Funding Information:
We thank Denis Duboule for generously providing the 5′HoxdDel(RXII) (d11-d13 deleted) mutant line, Deneen Wellik for the Hoxd11 mutant line, Steve Vokes for both RosaGrem1 and RosaGli3R transgenic lines, Trevor Williams for the AP2Cre transgenic line, Alan Godwin for help in generating the DSRed2 cassette used to create the Hoxd13 targeting construct, Matt Anderson for advice on HCR protocols, Rich Hwang for advice on tadpole analysis, and members of the CDBL for critical discussion. This research was supported by the Center for Cancer Research (S.M., Intramural Research Program) and the Frederick National Laboratory (W.F.H. and V.M., OMAL Contract no. 75N91019D00024 ), NCI, NIH ; by the Office of Infrastructure Programs , NIH (S.R.V. and M.K., P40-OD019794 ); and by grants from OHSU Medical Research Foundation and Shriners Hospitals for Children (H.S.S.).
Publisher Copyright:
© 2021
Keywords
- 5'Hoxd genes
- Gli3
- axolotl Gli3 CRISPR knockdown
- limb axis formation polarity
- postaxial limb polarity in mutant axolotl
- preaxial limb polarity in mutant mice
- vertebrate limb development
ASJC Scopus subject areas
- Neuroscience (all)
- Biochemistry, Genetics and Molecular Biology (all)
- Agricultural and Biological Sciences (all)