Programmed elimination of cells by caspase-independent cell extrusion in C. elegans

Abstract

The elimination of unnecessary or defective cells from metazoans occurs during normal development and tissue homeostasis, as well as in response to infection or cellular damage1. Although many cells are removed through caspase-mediated apoptosis followed by phagocytosis by engulfing cells2, other mechanisms of cell elimination occur3, including the extrusion of cells from epithelia through a poorly understood, possibly caspase-independent, process4. Here we identify a mechanism of cell extrusion that is caspase independent and that can eliminate a subset of the Caenorhabditis elegans cells programmed to die during embryonic development. In wild-type animals, these cells die soon after their generation through caspase-mediated apoptosis. However, in mutants lacking all four C. elegans caspase genes, these cells are eliminated by being extruded from the developing embryo into the extra-embryonic space of the egg. The shed cells show apoptosis-like cytological and morphological characteristics, indicating that apoptosis can occur in the absence of caspases in C. elegans. We describe a kinase pathway required for cell extrusion involving PAR-4, STRD-1 and MOP-25.1/-25.2, the C. elegans homologues of the mammalian tumour-suppressor kinase LKB1 and its binding partners STRADα and MO25α. The AMPK-related kinase PIG-1, a possible target of the PAR-4–STRD-1–MOP-25 kinase complex, is also required for cell shedding. PIG-1 promotes shed-cell detachment by preventing the cell-surface expression of cell-adhesion molecules. Our findings reveal a mechanism for apoptotic cell elimination that is fundamentally distinct from that of canonical programmed cell death.

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Figure 1: Cells with apoptotic morphology are shed from C. elegans embryos lacking caspase activity.
Figure 2: Cells that are shed from ced-3 -mutant embryos are normally fated to die early during wild-type embryogenesis.
Figure 3: The LKB1 homologue PAR-4 and the AMPK-related kinase PIG-1 are required for cell shedding from ced-3 embryos.
Figure 4: Shed cells lack cell-adhesion molecules that are inappropriately expressed in pig-1 mutants, possibly because of impaired endocytosis.

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Acknowledgements

We thank Z. Zhou, T. Hirose and S. Nakano for reporter constructs; B. Castor, E. Murphy and R. Droste for determining DNA sequences; S. Dennis for screening of the deletion library; N. An for strain management; and, J. Yuan, S. Nakano, N. Paquin, C. Engert and A. Saffer for discussions. The Caenorhabditis Genetic Center, which is funded by the National Institutes of Health National Center for Research Resources (NCRR), provided many strains. S. Mitani provided grp-1(tm1956). D.P.D. was supported by post-doctoral fellowships from the Damon Runyon Cancer Research Foundation and from the Charles A. King Trust. H.R.H. is the David H. Koch Professor of Biology at the Massachusetts Institute of Technology and an Investigator at the Howard Hughes Medical Institute.

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D.P.D. and H.R.H. designed the experiments, analysed the data and wrote the manuscript. D.P.D. and V.H. performed the experiments.

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Correspondence to H. Robert Horvitz.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Tables 1 -11, Supplementary Figures 1-8, legends for Supplementary Movies 1-3 (see separate files for Supplementary Movies), Supplementary References and the RNAi Construct Sequences. (PDF 9773 kb)

Supplementary Movie 1

This file contains a time-lapse movie of a developing ced-3(n3692) embryo (see Supplementary Information file for full legend). (MOV 5090 kb)

Supplementary Movie 2

This file contains a time-lapse movie of a developing pig-1(gm344) embryo (see Supplementary Information file for full legend). (MOV 2086 kb)

Supplementary Movie 3

This file contains a time-lapse movie of a developing pig-1(gm344) ced-3(n3692) embryo (see Supplementary Information file for full legend). (MOV 3778 kb)

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Denning, D., Hatch, V. & Horvitz, H. Programmed elimination of cells by caspase-independent cell extrusion in C. elegans. Nature 488, 226–230 (2012). https://doi.org/10.1038/nature11240

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