Retinoic Acid Receptors and the Control of Positional Information in the Regenerating Axolotl Limb

Abstract

We know little about the control of positional information (PI) during axolotl limb regeneration, which ensures that the limb regenerates exactly what was amputated, and the work reported here investigates this phenomenon. Retinoic acid administration changes the PI in a proximal direction so that a complete limb can be regenerated from a hand. Rather than identifying all the genes altered by RA treatment of the limb, we have eliminated many off-target effects by using retinoic acid receptor selective agonists. We firstly identify the receptor involved in this respecification process as RARα and secondly, identify the genes involved by RNA sequencing of the RARα-treated blastemal mesenchyme. We find 1177 upregulated genes and 1403 downregulated genes, which could be identified using the axolotl genome. These include several genes known to be involved in retinoic acid metabolism and in patterning. Since positional information is thought to be a property of the cell surface of blastemal cells when we examine our dataset with an emphasis on this aspect, we find the top canonical pathway is integrin signaling. In the extracellular matrix compartment, we find a MMP and several collagens are upregulated; several cell membrane genes and secretory factors are also upregulated. This provides data for future testing of the function of these candidates in the control of PI during limb regeneration.

Integration failure of regenerated limb tissue is associated with incongruencies in positional information in the Mexican axolotl

“…Apart from HoxC5 and Chrdl1, none of the genes that we analyzed had significant differences in expression between the week 0 (pre-RA treatment) and week 1 (1 week post RA) samples. It was surprising that we did not detect a decrease in FGF8 expression since a significant loss was previously detected by microarray in amputation blastemas treated with a retinoic acid receptor (RAR) agonist (Polvadore and Maden, 2021), and a bona fide repressive retinoic acid response element (RARE) has been identified upstream of the FGF8 promoter in multiple vertebrate species (Kumar and Duester, 2014). We also identified multiple predicted RARE consensus sequences, including one that appears to align spatially with conserved inhibitory FGF8 RARE in vertebrates, as well as in all the other axolotl genes that we analyzed (Supplementary Table S1).…”

Retinoic Acid and Retinoid X Receptors

“…The article by Polvadore and Maden [ 19 ], for example, explores the roles of RAR during limb regeneration in the axolotl Ambystoma mexicanum , a salamander native to Mexico. Using a differential transcriptomic screening approach based on pharmacological treatments with synthetic RAR paralog-specific agonists, they identify RARα as the dominant RAR to control positional identity in the regenerating limb and define a list of candidates acting downstream of RARα during this process.…”