Retinas from mice with a targeted disruption of the gene encoding forkhead transcription factor Foxn3 contained additional displaced amacrine interneurons and retinal astrocytes in the inner plexiform and ganglion cell layers, as well as ectopic primary cilia on bipolar and amacrine interneurons. Foxn3 is a transcriptional repressor and numerous genes linked to cilia structure or assembly were upregulated in embryonic retinas with disrupted Foxn3. CUT&RUN analysis revealed that many upregulated retinal genes were bound by the Foxn3 and Rfx3 proteins. A short hydrophobic motif (LXXLXWL) shared by Foxn3, Foxn4, and Foxj1 was required for association with Rfx3 and for full transcriptional repression by Foxn3, as well as for full transcriptional activation by Foxj1 or Foxn4. AlphaFold 3 predicted interaction between the hydrophobic motif and the Rfx3 dimerization domain. Mutations in Rfx3 at the predicted interaction site disrupted association of Rfx3 with Foxn3, Foxn4 or Foxj1. These results reveal a new layer of transcriptional regulation of genes required for cilia, with Foxn3 functioning as a repressor of cilia genes and limiting primary cilia formation in the developing retina.
Zhang et al. (Mon,) studied this question.