Candidate Adaptor Protein CED-6 Promotes the Engulfment of Apoptotic Cells in C. elegans

Autophagosomes are double-membrane intracellular vesicles that degrade protein aggregates, intracellular organelles, and other cellular components. During the development of the nematode Caenorhabditis elegans, many somatic and germ cells undergo apoptosis. These cells are engulfed and degraded by their neighboring cells. We discovered a novel role of autophagosomes in facilitating the degradation of apoptotic cells using a real-time imaging technique. Specifically, the double-membrane autophagosomes in engulfing cells are recruited to the surfaces of phagosomes containing apoptotic cells and subsequently fuse to phagosomes, allowing the inner vesicle to enter the phagosomal lumen. Mutants defective in the production of autophagosomes display significant defects in the degradation of apoptotic cells, demonstrating the importance of autophagosomes to this process. The signaling pathway led by the phagocytic receptor CED-1, the adaptor protein CED-6, and the large GTPase dynamin (DYN-1) promotes the recruitment of autophagosomes to phagosomes. Moreover, the subsequent fusion of autophagosomes with phagosomes requires the functions of the small GTPase RAB-7 and the HOPS complex components. Further observations suggest that autophagosomes provide apoptotic cell-degradation activities in addition to and in parallel of lysosomes. Our findings reveal that, unlike the single-membrane, LC3-associated phagocytosis (LAP) vesicles reported for mammalian phagocytes, the canonical double-membrane autophagosomes facilitate the clearance of C. elegans apoptotic cells. These findings add autophagosomes to the collection of intracellular organelles that contribute to phagosome maturation, identify novel crosstalk between the autophagy and phagosome maturation pathways, and discover the upstream signaling molecules that initiate this crosstalk.

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Faculty Opinions recommendation of C. elegans PAT-4/ILK functions as an adaptor protein within integrin adhesion complexes.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1006346.96519 ◽

2002 ◽

Author(s):

Christopher Turner

Keyword(s):

Adaptor Protein ◽

C Elegans

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C. elegans Dynamin Mediates the Signaling of Phagocytic Receptor CED-1 for the Engulfment and Degradation of Apoptotic Cells

Developmental Cell ◽

10.1016/j.devcel.2006.04.007 ◽

2006 ◽

Vol 10 (6) ◽

pp. 743-757 ◽

Cited By ~ 102

Author(s):

Xiaomeng Yu ◽

Sampeter Odera ◽

Chin-Hua Chuang ◽

Nan Lu ◽

Zheng Zhou

Keyword(s):

Apoptotic Cells ◽

C Elegans

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Autophagosomes fuse to phagosomes and play important roles in the degradation of apoptotic cells in Caenorhabditis elegans

10.1101/2021.07.16.452694 ◽

2021 ◽

Author(s):

Omar Pena-Ramos ◽

Lucia Chiao ◽

Xianghua Liu ◽

Tianyou Yao ◽

Henry He ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Small Gtpase ◽

Apoptotic Cells ◽

Phagosome Maturation ◽

Double Membrane ◽

C Elegans ◽

Intracellular Organelles ◽

Intracellular Vesicles ◽

Cellular Components

Autophagosomes are double-membrane intracellular vesicles that degrade protein aggregates, intracellular organelles, and other cellular components. In the nematode Caenorhabditis elegans, 113 somatic cells undergo apoptosis during embryogenesis and are engulfed and degraded by their neighboring cells. We discovered a novel role of autophagosomes in facilitating the degradation of apoptotic cells in C. elegans embryos using a real-time imaging technique. Specifically, double-membrane autophagosomes in engulfing cells are recruited to the surfaces of phagosomes containing apoptotic cells and subsequently fuse to phagosomes, allowing the inner membrane to enter the phagosomal lumen. Mutants defective in the production of autophagosomes display significant delays in the degradation of apoptotic cells, demonstrating the important contribution of autophagosomes to this process. The signaling pathway led by the phagocytic receptor CED-1, CED-1s adaptor CED-6, and the large GTPase dynamin (DYN-1) promote the recruitment of autophagosomes to phagosomes. Moreover, the subsequent fusion of autophagosomes with phagosomes requires the functions of the small GTPase RAB-7 and the HOPS complex. Our findings reveal that, unlike the single-membrane, LC3- associated phagocytosis (LAP) vesicles reported for mammalian phagocytes, canonical autophagosomes function in the clearance of C. elegans apoptotic cells. These findings add autophagosomes to the collection of intracellular organelles that contribute to phagosome maturation, identify novel crosstalk between the autophagy and phagosome maturation pathways, and discover the upstream factors that initiate this crosstalk.

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C. elegans mitochondrial factor WAH-1 promotes phosphatidylserine externalization in apoptotic cells through phospholipid scramblase SCRM-1

Nature Cell Biology ◽

10.1038/ncb1574 ◽

2007 ◽

Vol 9 (5) ◽

pp. 541-549 ◽

Cited By ~ 85

Author(s):

Xiaochen Wang ◽

Jin Wang ◽

Keiko Gengyo-Ando ◽

Lichuan Gu ◽

Chun-Ling Sun ◽

...

Keyword(s):

Apoptotic Cells ◽

C Elegans ◽

Phospholipid Scramblase ◽

Phosphatidylserine Externalization

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SLI-1 Cbl Inhibits the Engulfment of Apoptotic Cells in C. elegans through a Ligase-Independent Function

PLoS Genetics ◽

10.1371/journal.pgen.1003115 ◽

2012 ◽

Vol 8 (12) ◽

pp. e1003115 ◽

Cited By ~ 6

Author(s):

Courtney Anderson ◽

Shan Zhou ◽

Emma Sawin ◽

H. Robert Horvitz ◽

Michael E. Hurwitz

Keyword(s):

Apoptotic Cells ◽

Independent Function ◽

C Elegans

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Src-like Adaptor Protein Promotes the CD103+ dendritic Cell Homeostasis in the Lung

Blood ◽

10.1182/blood.v128.22.1339.1339 ◽

2016 ◽

Vol 128 (22) ◽

pp. 1339-1339

Author(s):

Namit Sharma ◽

Pan Zhongda ◽

Tracy Lauren Smith ◽

Savar Kaul ◽

Emilie Ernoult ◽

...

Keyword(s):

Bone Marrow ◽

Immune Response ◽

Immune Cell ◽

Conflicts Of Interest ◽

Adaptor Protein ◽

Cell Receptor ◽

Apoptotic Cells ◽

Direct Role

Abstract Dendritic cells (DCs) along with mast cells function as sentinels for the innate immune system and perform as antigen presenting cells (APCs) to mount an adaptive immune response against invading pathogen. FLT3 receptor tyrosine kinase signaling has been shown to regulate the homeostatic mechanisms of subsets of DCs particularly, CD103+DCs compared to CD11b+DCs. CD103+DCs are regarded as APCs with superior capabilities to mount an effective immune response, thus understanding their homeostasis mechanism(s)/function is of paramount importance to devise effective therapeutics including DC vaccines. The Src-like adapter protein (SLAP) has been shown to dampen the signaling downstream of receptor tyrosine kinases including FLT3, cKit, and immune cell receptors including T cell receptor, B cell receptor, and Granulocyte-monocyte colony stimulating factor receptor via by recruiting c-Cbl, an ubiquitin ligase. Here, we report that SLAP deficient mice (KO) have reduced numbers of CD103+DC in lung while equal numbers in liver and kidney compared to control mice. To further confirm reduced CD103+DC in the lung, efferocytosis assays that are dependent upon CD+103 DC in lung epithelium to cleanse the apoptotic cells were performed. Flow cytometric quantification of CD103+DCs that uptake fluorescently labeled apoptotic cells administered via intranasal route and migrate to mediastinal lymph nodes confirmed reduced number of CD103+DCs in SLAP KO mice. Further analysis of DC progenitor populations showed reduced pre-DC progenitor in the lung in SLAP KO mice while bone marrow compartment showed equal progenitor populations including pre-DC and common dendritic progenitors suggesting the role of SLAP in localized FLT3 signaling in the lung. Consistently, DCs in lymphoid compartment including spleen, thymus, inguinal and popliteal lymph node did not show any defects. Upon further dissecting the cellular mechanism, SLAP KO DCs showed increased apoptosis while having similar proliferation potential in vivo at steady state.Bone marrow progenitors from SLAP KO mice failed to generate mature DCs in the presence of FLT3 ligand in vitrodue to enhanced apoptosis at early time points. Also, submaximal inhibition of FLT3 with an inhibitor, quizartinib partially rescues the apoptotic phenotype of SLAP KO bone marrow progenitors suggesting a cell-intrinsic role of SLAP in the survival of DCs. Biochemical analysis revealed that SLAP is directly recruited to the juxta-membrane residues of the FLT3 receptor in an inducible manner suggesting a direct role of SLAP in the regulation of FLT3 signaling. Phosphoflow analysis of DCs generated in the combined presence of GMCSF and FLT3 ligands showed that SLAP promotes the signaling to SHP2 while perturbs signaling to the mTOR pathway. Together these results suggest that SLAP is a critical regulator of CD103+DCs homeostasis in selective peripheral organs including the lung. Disclosures No relevant conflicts of interest to declare.

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Acentrosomal spindle assembly and stability in C. elegans oocytes requires a kinesin-12 non-motor microtubule interaction domain

10.1101/2021.06.17.448874 ◽

2021 ◽

Author(s):

Ian Daniel Wolff ◽

Jeremy Alden Hollis ◽

Sarah Marie Wignall

Keyword(s):

Adaptor Protein ◽

Direct Interaction ◽

Spindle Assembly ◽

Meiotic Spindle ◽

Interaction Domain ◽

Microtubule Binding ◽

C Elegans ◽

Insight Into

During the meiotic divisions in oocytes, microtubules are sorted and organized by motor proteins to generate a bipolar spindle in the absence of centrosomes. In most organisms, kinesin-5 family members crosslink and slide microtubules to generate outward force that promotes acentrosomal spindle bipolarity. However, the mechanistic basis for how other kinesin families act on acentrosomal spindles has not been explored. We investigated this question in C. elegans oocytes, where kinesin-5 is not required to generate outward force. Instead, the kinesin-12 family motor KLP-18 performs this function. KLP-18 acts with adaptor protein MESP-1 (meiotic spindle 1) to sort microtubule minus ends to the periphery of a microtubule array, where they coalesce into spindle poles. If either of these proteins is depleted, outward sorting of microtubules is lost and minus ends converge to form a monoaster. Here we use a combination of in vitro biochemical assays and in vivo mutant analysis to provide insight into the mechanism by which these proteins collaborate to promote acentrosomal spindle assembly. We identify a microtubule binding site on the C-terminal stalk of KLP-18 and demonstrate that a direct interaction between the KLP-18 stalk and MESP-1 activates non-motor microtubule binding. We also provide evidence that this C-terminal domain is required for KLP-18 activity during spindle assembly and show that KLP-18 is continuously required to maintain spindle bipolarity. This study thus provides new insight into the construction and maintenance of the oocyte acentrosomal spindle as well as into kinesin-12 mechanism and regulation.

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