NRF2 negatively regulates primary ciliogenesis and hedgehog signaling

Abstract

Primary cilia are lost during cancer development, but the mechanism regulating cilia degeneration is not determined. While transcription factor nuclear factor-erythroid 2-like 2 (NRF2) protects cells from oxidative, proteotoxic, and metabolic stress in normal cells, hyperactivation of NRF2 is oncogenic, although the detailed molecular mechanisms by which uncontrolled NRF2 activation promotes cancer progression remain unclear. Here, we report that NRF2 suppresses hedgehog (Hh) signaling through Patched 1 (PTCH1) and primary ciliogenesis via p62/sequestosome 1 (SQSTM1). PTCH1, a negative regulator of Hh signaling, is an NRF2 target gene, and as such, hyperactivation of NRF2 impairs Hh signaling. NRF2 also suppresses primary cilia formation through p62-dependent inclusion body formation and blockage of Bardet-Biedl syndrome 4 (BBS4) entrance into cilia. Simultaneous ablation of PTCH1 and p62 completely abolishes NRF2-mediated inhibition of both primary ciliogenesis and Hh signaling. Our findings reveal a previously unidentified role of NRF2 in controlling a cellular organelle, the primary cilium, and its associated Hh signaling pathway and also uncover a mechanism by which NRF2 hyperactivation promotes tumor progression via primary cilia degeneration and aberrant Hh signaling. A better understanding of the crosstalk between NRF2 and primary cilia/Hh signaling could not only open new avenues for cancer therapeutic discovery but could also have significant implications regarding pathologies other than cancer, including developmental disorders, in which improper primary ciliogenesis and Hh signaling play a major role.

NRF2 and Primary Cilia: An Emerging Partnership

“…Moreover, we identified high-confidence ARE sequences in the promoters of Gpr161 , Gli2 , Gli3 and Ift27 , with the putative AREs in Gli2 and Gli3 being perfect matches with the ARE consensus sequence [ 58 ]. We also found candidate AREs in Ptch1 , the Hh receptor and target gene that has recently been confirmed as a bona fide direct NRF2 target [ 58 , 59 ]. Further evidence that Gpr161 and Ift27 (and the ciliogenic Ift74 ) are NRF2 targets comes from their promoters being identified in a ChIP-seq experiment aiming to identify NRF2-binding sites [ 104 ].…”

“…The NRF2-dependent Hh pathway components include both positive ( Smo , Gli2 , Ift27 ) and negative ( Gpr161 , Gli3, Sufu , Ptch1 ) regulators of the Hh pathway, such that the net effects of their downregulation on Hh responsiveness are not immediately obvious [ 12 , 58 ]. For example, one consideration is that Ptch1 is not only a Hh/GLI target gene but also, as shown recently, an NRF2 target gene; thus, the reduction we observed in SAG-treated NRF2-null cells may be explained, at least partly, by lack of direct NRF2 activation [ 58 , 59 ]. This may also be the reason why we found, as mentioned above, a downward trend in unstimulated Ptch1 levels in NRF2-null cells.…”

“…Furthermore, as we also showed, primary cilia-NRF2 connections are bidirectional, as NRF2 affects ciliogenesis and Hh signaling, both of which it regulates transcriptionally [ 58 , 59 ]. Interestingly, though, while we found a positive effect of NRF2 on cilia and Hh signaling [ 58 , 59 ], a very recent study found a negative connection instead [ 58 , 59 ]. In the remainder of this paper, we will review the reciprocal functional interactions between primary cilia and NRF2, discussing their possible pathophysiological meanings and proposing possible explanations where the evidence appears contradictory.…”

“…However, there is so far no direct evidence indicating that Gli1 is an NRF2 target gene. Moreover, the observed NRF2-dependent changes in Ptch1 expression are weak compared to those in other NRF2 target genes, and therefore lack of direct NRF2 activation may explain only a small fraction of the strong reduction in SAG-induced Ptch1 levels observed in NRF2-null cells [ 58 , 59 ]. Consistently, we detected no increase in Gli1 or Ptch1 mRNAs in WT MEFs upon NRF2 stabilization with dimethyl fumarate, an electrophile activator of NRF2 [ 58 , 105 , 106 , 107 ].…”

“…However, recent work indicates that NRF2 can also function as a negative regulator of ciliogenesis and Hh pathway activity [ 59 ]. It is therefore of interest to review these data and discuss whether and how NRF2 may act both positively and negatively in these processes.…”