Cilia are sensory organelles protruding from cell surfaces. Release of Extracellular Vesicles (EVs) from cilia was previously observed in mammals, Chlamydomonas, and in male C. elegans. Using the EV marker TSP-6 (an ortholog of mammalian CD9) and other ciliary receptors, we show that EVs are formed from ciliated sensory neurons in C. elegans hermaphrodites. Release of EVs is observed from two ciliary locations: the cilia tip and/or Periciliary Membrane Compartment (PCMC). Outward budding of EVs from the cilia tip leads to their release into the environment. EVs budding from the PCMC are concomitantly phagocytosed by the associated glial cells. To maintain cilia composition, a tight regulation of cargo import and removal is achieved by the action of Intra-Flagellar Transport (IFT). Unbalanced IFT due to cargo overexpression or mutations in the IFT machinery leads to local accumulation of ciliary proteins. Disposal of excess ciliary proteins via EVs reduces their local accumulation and exports them to the environment and/or to the glia associated to these ciliated neurons. We suggest that EV budding from cilia subcompartments acts as a safeguard mechanism to remove deleterious excess of ciliary material.
All data generated or analysed during this study are included in the manuscript and supporting files. The manuscript is a microscopy study all datapoints are represented in figures and figure supplement. Supplementary table contain all material used in this work.
- Patrick FM Laurent
- Adria Razzauti
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Douglas Portman, University of Rochester, United States
© 2021, Razzauti & Laurent
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
The phagocytic receptor CED-1 mediates apoptotic cell recognition by phagocytic cells, enabling cell corpse clearance in Caenorhabditis elegans. Whether appropriate levels of CED-1 are maintained for executing the engulfment function remains unknown. Here, we identified the C. elegans E3 ubiquitin ligase tripartite motif containing-21 (TRIM-21) as a component of the CED-1 pathway for apoptotic cell clearance. When the NPXY motif of CED-1 was bound to the adaptor protein CED-6 or the YXXL motif of CED-1 was phosphorylated by tyrosine kinase SRC-1 and subsequently bound to the adaptor protein NCK-1 containing the SH2 domain, TRIM-21 functioned in conjunction with UBC-21 to catalyze K48-linked poly-ubiquitination on CED-1, targeting it for proteasomal degradation. In the absence of TRIM-21, CED-1 accumulated post-translationally and drove cell corpse degradation defects, as evidenced by direct binding to VHA-10. These findings reveal a unique mechanism for the maintenance of appropriate levels of CED-1 to regulate apoptotic cell clearance.
Key protein adapters couple translation to mRNA decay on specific classes of problematic mRNAs in eukaryotes. Slow decoding on non-optimal codons leads to codon-optimality-mediated decay (COMD) and prolonged arrest at stall sites leads to no-go decay (NGD). The identities of the decay factors underlying these processes and the mechanisms by which they respond to translational distress remain open areas of investigation. We use carefully designed reporter mRNAs to perform genetic screens and functional assays in Saccharomyces cerevisiae. We characterize the roles of Hel2, Syh1, and Smy2 in coordinating translational repression and mRNA decay on NGD reporter mRNAs, finding that Syh1 and, to a lesser extent its paralog Smy2, act in a distinct pathway from Hel2. This Syh1/Smy2-mediated pathway acts as a redundant, compensatory pathway to elicit NGD when Hel2-dependent NGD is impaired. Importantly, we observe that these NGD factors are not involved in the degradation of mRNAs enriched in non-optimal codons. Further, we establish that a key factor previously implicated in COMD, Not5, contributes modestly to the degradation of an NGD-targeted mRNA. Finally, we use ribosome profiling to reveal distinct ribosomal states associated with each reporter mRNA that readily rationalize the contributions of NGD and COMD factors to degradation of these reporters. Taken together, these results provide new insight into the role of Syh1 and Smy2 in NGD and into the ribosomal states that correlate with the activation of distinct pathways targeting mRNAs for degradation in yeast.