P-bodies are sites of rapid RNA decay during the neural crest epithelial—mesenchymal transition

Abstract

The epithelial-mesenchymal transition (EMT) drives cellular movements during development to create specialized tissues and structures in metazoans, using mechanisms often coopted during metastasis. Neural crest cells are a multipotent stem cell population that undergo a developmentally regulated EMT and are prone to metastasis in the adult, providing an excellent model to study cell state changes and mechanisms underlying EMT. A hallmark of neural crest EMT during avian development is temporally restricted expression followed by rapid down-regulation of the Wnt antagonist Draxin. Using live RNA imaging, here we demonstrate that rapid clearance of Draxin transcripts is mediated post-transcriptionally via localization to processing bodies (P-bodies), small cytoplasmic granules which are established sites of RNA processing. Contrasting with recent work in immortalized cell lines suggesting that P-bodies are sites of storage rather than degradation, we show that targeted decay of Draxin occurs within P-bodies during neural crest migration. Furthermore, P-body disruption via DDX6 knockdown inhibits not only endogenous Draxin down-regulation but also neural crest EMT in vivo. Together, our data highlight a novel and important role for P-bodies in an intact organismal context−controlling a developmental EMT program via post-transcriptional target degradation.

Biomolecular condensates amplify mRNA decapping by biasing enzyme conformation

“…Although the molecular composition and interactions within P-bodies is well studied, their biological function is controversial. Several studies suggest P-bodies are sites of decay because colocalized mRNA are cleared over time 16 , 18 – 20 . Alternatively, P-bodies may function as sites of mRNA storage due to the absence of decay intermediates in sequencing and live cell imaging data 15 , 21 , 22 .…”

RNA-binding protein Elavl1/HuR is required for maintenance of cranial neural crest specification

“…MOs were co-electroporated with pCIG ( Megason and McMahon, 2002 ) or pCI-H2B-RFP ( Betancur et al, 2010 ) to increase electroporation efficiency and to visualize successfully electroporated cells. The Draxin -FLAG overexpression construct ( Draxin OE; Hutchins and Bronner, 2018 ; Hutchins and Bronner, 2019 ) and the MCP-GFP construct Hutchins et al, 2020 have been previously described. The MS2- Draxin 3′-UTR construct used in this study was generated by exchanging the mTurq2 coding region for a Luciferase coding region in a previously described MS2- Draxin 3′-UTR construct ( Hutchins et al, 2020 ).…”

RNA-binding protein Elavl1/HuR is required for maintenance of cranial neural crest specification

“…MOs were co-electroporated with pCIG (Megason and McMahon, 2002) or pCI-H2B-RFP (Betancur et al, 2010) to increase electroporation efficiency and to visualize successfully electroporated cells. The Draxin-FLAG overexpression construct (Draxin OE) (Hutchins and Bronner, 2018, 2019) and the MCP-GFP construct (Hutchins et al, 2020) have been previously described. The MS2- Draxin 3’UTR construct used in this study was generated by exchanging the mTurq2 coding region for a Luciferase coding region in a previously described MS2- Draxin 3’UTR construct (Hutchins et al, 2020).…”

“…The Draxin-FLAG overexpression construct (Draxin OE) (Hutchins and Bronner, 2018, 2019) and the MCP-GFP construct (Hutchins et al, 2020) have been previously described. The MS2- Draxin 3’UTR construct used in this study was generated by exchanging the mTurq2 coding region for a Luciferase coding region in a previously described MS2- Draxin 3’UTR construct (Hutchins et al, 2020). Electroporations were performed on HH4 gastrula stage chicken embryos as described previously (Hutchins and Bronner, 2018, 2019).…”