Retinoic acid breakdown is required for proximodistal positional identity during axolotl limb regeneration

Timothy J. Duerr; Melissa Miller; Sage Kumar; Dareen Bakr; Jackson R. Griffiths; Aditya K. Gautham; Danielle Douglas; S. Randal Voss; James R. Monaghan

doi:10.1038/s41467-025-59497-5

Retinoic acid breakdown is required for proximodistal positional identity during axolotl limb regeneration

Timothy J. Duerr, Melissa Miller, Sage Kumar, Dareen Bakr, Jackson R. Griffiths, Aditya K. Gautham, Danielle Douglas, S. Randal Voss, James R. Monaghan

Research output: Contribution to journal › Article › peer-review

3 Scopus citations

Abstract

Regenerating limbs retain their proximodistal (PD) positional identity following amputation. This positional identity is genetically encoded by PD patterning genes that instruct blastema cells to regenerate the appropriate PD limb segment. Retinoic acid (RA) is known to specify proximal limb identity, but how RA signaling levels are established in the blastema is unknown. Here, we show that RA breakdown via CYP26B1 is essential for determining RA signaling levels within blastemas. CYP26B1 inhibition molecularly reprograms distal blastemas into a more proximal identity, phenocopying the effects of administering excess RA. We identify Shox as an RA-responsive gene that is differentially expressed between proximally and distally amputated limbs. Ablation of Shox results in shortened limbs with proximal skeletal elements that fail to initiate endochondral ossification. These results suggest that PD positional identity is determined by RA degradation and RA-responsive genes that regulate PD skeletal element formation during limb regeneration.

Original language	English
Article number	4798
Journal	Nature Communications
Volume	16
Issue number	1
DOIs	https://doi.org/10.1038/s41467-025-59497-5
State	Published - Dec 2025

Bibliographical note

Publisher Copyright:
© The Author(s) 2025.

Funding

The authors thank Guoxin Rong for his imaging expertise and assistance with microscopy. Additionally, we thank Prayag Murawala for providing transgenic animals and the Ambystoma Genetic Stock Center for non-transgenic animals. We thank Malcolm Maden for his critical analysis of the manuscript. We finally thank the Institute for Chemical Imaging of Living Systems at Northeastern University for consultation and imaging support. The work from this paper was funded by NIH grant R01HD099174 and by NSF grants 1558017 and 1656429, all obtained by J.R.M. Non-transgenic animals were obtained from the Ambystoma Genetic Stock Center funded through NIH grant P40-OD019794 obtained by S.R.V.

Funders	Funder number
National Institutes of Health (NIH)	R01HD099174
National Science Foundation Arctic Social Science Program	1558017, 1656429, P40-OD019794

ASJC Scopus subject areas

General Chemistry
General Biochemistry, Genetics and Molecular Biology
General
General Physics and Astronomy

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title = "Retinoic acid breakdown is required for proximodistal positional identity during axolotl limb regeneration",

abstract = "Regenerating limbs retain their proximodistal (PD) positional identity following amputation. This positional identity is genetically encoded by PD patterning genes that instruct blastema cells to regenerate the appropriate PD limb segment. Retinoic acid (RA) is known to specify proximal limb identity, but how RA signaling levels are established in the blastema is unknown. Here, we show that RA breakdown via CYP26B1 is essential for determining RA signaling levels within blastemas. CYP26B1 inhibition molecularly reprograms distal blastemas into a more proximal identity, phenocopying the effects of administering excess RA. We identify Shox as an RA-responsive gene that is differentially expressed between proximally and distally amputated limbs. Ablation of Shox results in shortened limbs with proximal skeletal elements that fail to initiate endochondral ossification. These results suggest that PD positional identity is determined by RA degradation and RA-responsive genes that regulate PD skeletal element formation during limb regeneration.",

author = "Duerr, \{Timothy J.\} and Melissa Miller and Sage Kumar and Dareen Bakr and Griffiths, \{Jackson R.\} and Gautham, \{Aditya K.\} and Danielle Douglas and \{Randal Voss\}, S. and Monaghan, \{James R.\}",

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AU - Duerr, Timothy J.

AU - Miller, Melissa

AU - Kumar, Sage

AU - Bakr, Dareen

AU - Griffiths, Jackson R.

AU - Gautham, Aditya K.

AU - Douglas, Danielle

AU - Randal Voss, S.

AU - Monaghan, James R.

PY - 2025/12

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N2 - Regenerating limbs retain their proximodistal (PD) positional identity following amputation. This positional identity is genetically encoded by PD patterning genes that instruct blastema cells to regenerate the appropriate PD limb segment. Retinoic acid (RA) is known to specify proximal limb identity, but how RA signaling levels are established in the blastema is unknown. Here, we show that RA breakdown via CYP26B1 is essential for determining RA signaling levels within blastemas. CYP26B1 inhibition molecularly reprograms distal blastemas into a more proximal identity, phenocopying the effects of administering excess RA. We identify Shox as an RA-responsive gene that is differentially expressed between proximally and distally amputated limbs. Ablation of Shox results in shortened limbs with proximal skeletal elements that fail to initiate endochondral ossification. These results suggest that PD positional identity is determined by RA degradation and RA-responsive genes that regulate PD skeletal element formation during limb regeneration.

AB - Regenerating limbs retain their proximodistal (PD) positional identity following amputation. This positional identity is genetically encoded by PD patterning genes that instruct blastema cells to regenerate the appropriate PD limb segment. Retinoic acid (RA) is known to specify proximal limb identity, but how RA signaling levels are established in the blastema is unknown. Here, we show that RA breakdown via CYP26B1 is essential for determining RA signaling levels within blastemas. CYP26B1 inhibition molecularly reprograms distal blastemas into a more proximal identity, phenocopying the effects of administering excess RA. We identify Shox as an RA-responsive gene that is differentially expressed between proximally and distally amputated limbs. Ablation of Shox results in shortened limbs with proximal skeletal elements that fail to initiate endochondral ossification. These results suggest that PD positional identity is determined by RA degradation and RA-responsive genes that regulate PD skeletal element formation during limb regeneration.

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Retinoic acid breakdown is required for proximodistal positional identity during axolotl limb regeneration

Abstract

Bibliographical note

Funding

ASJC Scopus subject areas

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