Microproteins Simba1 and Simba2 Activate Wingless Signaling during the Reactivation of Neural Stem Cells in Drosophila
The ability of neural stem cells (NSC) to switch between quiescent and proliferative states is fundamental for adult neurogenesis and regeneration. Microproteins or short open reading frame (sORF)-encoded peptides (SEPs), are highly abundant yet largely understudied, and their role in brain development remains unclear. Here, we demonstrate that two microproteins, Simba1 and Simba2 encoded by highly conserved sORFs CG15715 and CG18081, respectively, govern the reactivation of Drosophila quiescent NSCs. Both Simba1 and Simba2 function in NSCs and Blood-Brain-Barrier (BBB) glial cells to promote NSC reactivation. Mechanistically, Simba1 and Simba2 act as transcription factors activating the WNT/Wingless signalling pathway during NSC reactivation. We uncovered a critical role of Wg signalling molecules in promoting NSC reactivation and the translocation of Wingless from BBB glia to NSCs. Moreover, ZNF706, the human ortholog of Simba1/2, is required for WNT signaling activation in human STF3A cells and may also target the WNT signaling pathway in human cancer cells. Our findings reveal a novel role for microproteins in regulating NSC reactivation through Wg signalling. The conserved function of Simba1/2/ZNF706 in activating WNT/Wg signalling in Drosophila and humans suggest that this new regulatory paradigm may be applicable to broader cellular processes and disease conditions. Overall design: Examination of gene regions targeted by Simba1 or Simba2 in NSCs. Two batches of experiments were conducted. In each batch, one control (UAS-Dam) and two gene-of-interest (UAS-Dam-Simba1 and UAS-Dam-Simba2) were included. UAS contructs were induced under NSC-specific driver, grh-Gal4. Dam-Simba1/2 then bound to DNA and methylated adenines (m6A) at the nearby GATC motifs. By examining the DNA methlyation status through next generation sequencing, Simba1/2 target regions could be identified.