GLI transcriptional repression is inert prior to Hedgehog pathway activation

Abstract

The Hedgehog (HH) pathway regulates a spectrum of developmental processes through the transcriptional mediation of GLI proteins. GLI repressors control tissue patterning by preventing sub-threshold activation of HH target genes, presumably even before HH induction, while lack of GLI repression activates most targets. Despite GLI repression being central to HH regulation, it is unknown when it first becomes established in HH-responsive tissues. Here, we investigate whether GLI3 prevents precocious gene expression during limb development. Contrary to current dogma, we find that GLI3 is inert prior to HH signaling. While GLI3 binds to most targets, loss of Gli3 does not increase target gene expression, enhancer acetylation or accessibility, as it does post-HH signaling. Furthermore, GLI repression is established independently of HH signaling, but after its onset. Collectively, these surprising results challenge current GLI pre-patterning models and demonstrate that GLI repression is not a default state for the HH pathway.

Sonic hedgehog is not a limb morphogen but acts as a trigger to specify all digits in mice

“…No activity was detected in the remaining 50%, presumably because recombination occurred prior to Shh expression onset in those embryos. Notably, transient Shh activity is also first detectable in controls at 29 somites (completely absent in all embryos at 28 somites; this study Figures 1C, 3, S1A, and S2;in agreement with Zhu et al, 2008;Lex et al, 2022). Within 2 h, by the 30-somite stage (+1 somite) when Shh activity (Ptch1, Gli1 expression) is detected in 100% of control (Shh +/À ) sibling embryos, Shh activity had ceased in all Shh-CKO embryos (Figures 1C and S1A) and subsequently remained absent.…”

“…We considered the possibility that either persistent low-level, residual pathway activity or a long-range response during the transient Shh expression pulse had escaped detection but think this unlikely for the following reasons. (1) Low-level Shh activity persistence, owing to either incomplete Shh recombination or some compensatory feedback leading to ectopic Hh ligand activation, should result in the continued activation of direct targets Gli1 and Ptch1, which are detectable at much lower levels than is Shh RNA (Lewis et al, 2001;Lex et al, 2022;Zhu et al, 2008). Furthermore, Shh-expressing cells proliferate in the early limb bud to give rise to d4/d5, and activation of direct target reporters should be increasingly apparent over time (E10.5-11.5).…”

“…Stable alterations in target gene expression following transient Shh exposure could involve several, non-mutually exclusive mechanisms, including chromatin and/or DNA modifications and relay mechanisms incorporating lock-on circuitry (Alon, 2007). Recent work suggests that Shh targets are poised for expression as soon as Shh activity initiates and that Gli3-mediated repression commences only after this point (Lex et al, 2020;Lex et al, 2022). A transient burst of Shh activity could trigger a self-reinforcing bi-stable switch (Alon, 2007) if activating factors that reinforce target expression also block the introduction of repressive marks by Gli3R post-Shh onset.…”

Wumei Pill Ameliorates AOM/DSS-Induced Colitis-Associated Colon Cancer through Inhibition of Inflammation and Oxidative Stress by Regulating S-Adenosylhomocysteine Hydrolase- (AHCY-) Mediated Hedgehog Signaling in Mice

“…In addition, a study found that the engagement of Hh ligands to Patched can be negatively regulated by endogenous antagonist of Hh ligands, i.e., hedgehog-interacting protein (Hhip). Besides, the translocation of Gli 1 from the cytoplasm to the nucleus is concomitant with the activation of Hh signaling [ 65 , 66 ]. Hh signaling has been shown to be involved in the AOM/DSS-induced CAC, and suppression of Hh signaling contributed to the amelioration of CAC [ 67 ].…”

Sonic Hedgehog is not a limb morphogen but acts as a trigger to specify all digits

“…A low-level persistence of Shh pathway activation, owing to either incomplete recombination to delete Shh during the period of tamoxifen activity or to some compensatory feedback leading to ectopic Shh activation or activation of another Hh ligand should all result in the continued activation of direct targets Gli1 and Ptch1 , which are detectable at much lower levels than is Shh RNA 5, 12, 42 . Furthermore, because Shh-expressing cells proliferate in the early limb bud (later giving rise to posterior digits), the activation of direct target reporters should become increasingly apparent over time (E10.5-11.5).…”