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Phagocytic receptor CED-1 initiates a signaling pathway for degrading engulfed apoptotic cells.

Yu X, Lu N, Zhou Z - PLoS Biol. (2008)

Bottom Line: We found that mutations in each pathway not only delay or block engulfment, but also delay the degradation of engulfed apoptotic cells.We propose that RAB-7 functions as a downstream effector of the CED-1 pathway to mediate phagolysosome formation.Our work suggests that phagocytic receptors, which were thought to act specifically in initiating engulfment, also control phagosome maturation through the sequential activation of multiple effectors such as dynamin, PI(3)P, and Rab GTPases.

View Article: PubMed Central - PubMed

Affiliation: Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, Texas, United States of America.

ABSTRACT
Apoptotic cells in animals are engulfed by phagocytic cells and subsequently degraded inside phagosomes. To study the mechanisms controlling the degradation of apoptotic cells, we developed time-lapse imaging protocols in developing Caenorhabditis elegans embryos and established the temporal order of multiple events during engulfment and phagosome maturation. These include sequential enrichment on phagocytic membranes of phagocytic receptor cell death abnormal 1 (CED-1), large GTPase dynamin (DYN-1), phosphatidylinositol 3-phosphate (PI(3)P), and the small GTPase RAB-7, as well as the incorporation of endosomes and lysosomes to phagosomes. Two parallel genetic pathways are known to control the engulfment of apoptotic cells in C. elegans. We found that mutations in each pathway not only delay or block engulfment, but also delay the degradation of engulfed apoptotic cells. One of the pathways, composed of CED-1, the adaptor protein CED-6, and DYN-1, controls the rate of enrichment of PI(3)P and RAB-7 on phagosomal surfaces and the formation of phagolysosomes. We further identified an essential role of RAB-7 in promoting the recruitment and fusion of lysosomes to phagosomes. We propose that RAB-7 functions as a downstream effector of the CED-1 pathway to mediate phagolysosome formation. Our work suggests that phagocytic receptors, which were thought to act specifically in initiating engulfment, also control phagosome maturation through the sequential activation of multiple effectors such as dynamin, PI(3)P, and Rab GTPases.

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PI(3)P and RAB-7 Are Sequentially Enriched on the Surface of Maturing Phagosomes Containing Apoptotic Cells(A) Diagram of phagocytosis and the subsequent degradation of a cell corpse.(B) The two partially redundant and parallel pathways that control cell-corpse removal in C. elegans. Names in parentheses indicate mammalian homologs.(C and D) Time-lapse images of wild-type embryos coexpressing Pced-1ced-1::gfp / Pced-12xFYVE::mrfp1 (C) and Pced-12xFYVE::mrfp1 / Pced-1gfp::rab-7 (D). Recording started at approximately 320 min after first cleavage. Anterior is to the top. Ventral faces readers. Scale bars indicate 10 μm. (C) Cell corpse C3 (white arrow) is engulfed by ABplaapppp (white arrowhead). Yellow arrows and arrowheads indicate C1 and C2 and their corresponding engulfing cells. “0 min” represents when CED-1::GFP is first detected on the budding pseudopods. (D) Time-lapse images of signals around C3 (arrows) during its degradation. “0 min” represents when ABplaapppp fully internalized C3. Arrows indicate C1, C2, and C3 in (a), of which the corresponding engulfing cells are outlined and indicated by arrowheads. Insets in (e) and (l) show an amplified view of the region surrounding C3; arrowheads indicate tubular structures. The yellow arrowhead in (d) depicts the nucleus of ABpraapppa, which has engulfed C2. (o) The relative reporter signal intensity measured from (a) to (n) is plotted over time.(E) Summary of the temporal order of each reporter detected on the surfaces of phagocytic cups and phagosomes. Data represent means obtained from time-lapse recording of multiple C1, C2, and C3 cell corpses (see text for actual numbers). “0 min” represents when budding pseudopods are first detected by CED-1::GFP. 2xFYVE, HGRS-1, RAB-7, and CTNS-1 reporters localize to phagosome surface until the cell corpse is fully degraded. The conversion of pale to deep pink colors of HGRS-1::mRFP1 and CTNS-1::mRFP1 indicates that starting from an initial weak signal, the signal intensity gradually increases.
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pbio-0060061-g001: PI(3)P and RAB-7 Are Sequentially Enriched on the Surface of Maturing Phagosomes Containing Apoptotic Cells(A) Diagram of phagocytosis and the subsequent degradation of a cell corpse.(B) The two partially redundant and parallel pathways that control cell-corpse removal in C. elegans. Names in parentheses indicate mammalian homologs.(C and D) Time-lapse images of wild-type embryos coexpressing Pced-1ced-1::gfp / Pced-12xFYVE::mrfp1 (C) and Pced-12xFYVE::mrfp1 / Pced-1gfp::rab-7 (D). Recording started at approximately 320 min after first cleavage. Anterior is to the top. Ventral faces readers. Scale bars indicate 10 μm. (C) Cell corpse C3 (white arrow) is engulfed by ABplaapppp (white arrowhead). Yellow arrows and arrowheads indicate C1 and C2 and their corresponding engulfing cells. “0 min” represents when CED-1::GFP is first detected on the budding pseudopods. (D) Time-lapse images of signals around C3 (arrows) during its degradation. “0 min” represents when ABplaapppp fully internalized C3. Arrows indicate C1, C2, and C3 in (a), of which the corresponding engulfing cells are outlined and indicated by arrowheads. Insets in (e) and (l) show an amplified view of the region surrounding C3; arrowheads indicate tubular structures. The yellow arrowhead in (d) depicts the nucleus of ABpraapppa, which has engulfed C2. (o) The relative reporter signal intensity measured from (a) to (n) is plotted over time.(E) Summary of the temporal order of each reporter detected on the surfaces of phagocytic cups and phagosomes. Data represent means obtained from time-lapse recording of multiple C1, C2, and C3 cell corpses (see text for actual numbers). “0 min” represents when budding pseudopods are first detected by CED-1::GFP. 2xFYVE, HGRS-1, RAB-7, and CTNS-1 reporters localize to phagosome surface until the cell corpse is fully degraded. The conversion of pale to deep pink colors of HGRS-1::mRFP1 and CTNS-1::mRFP1 indicates that starting from an initial weak signal, the signal intensity gradually increases.

Mentions: An engulfing cell recognizes an apoptotic cell through phagocytic receptor(s) and extends thin pseudopods around it to generate a phagocytic cup. The fusion of growing pseudopods and scission of a vacuole containing the apoptotic cell from the plasmalemma generate a phagosome inside the host cell (Figure 1A) [1]. The swift engulfment (phagocytosis) and subsequent degradation of apoptotic cells inside phagosomes eliminate dying cells before they release any potentially harmful contents, and actively prevent tissue injury, inflammatory responses, and autoimmune diseases [2]. Inefficient degradation of the components of engulfed apoptotic cells, such as nuclear DNA, similar to inefficient engulfment, results in severe inflammatory and autoimmune responses [3,4].


Phagocytic receptor CED-1 initiates a signaling pathway for degrading engulfed apoptotic cells.

Yu X, Lu N, Zhou Z - PLoS Biol. (2008)

PI(3)P and RAB-7 Are Sequentially Enriched on the Surface of Maturing Phagosomes Containing Apoptotic Cells(A) Diagram of phagocytosis and the subsequent degradation of a cell corpse.(B) The two partially redundant and parallel pathways that control cell-corpse removal in C. elegans. Names in parentheses indicate mammalian homologs.(C and D) Time-lapse images of wild-type embryos coexpressing Pced-1ced-1::gfp / Pced-12xFYVE::mrfp1 (C) and Pced-12xFYVE::mrfp1 / Pced-1gfp::rab-7 (D). Recording started at approximately 320 min after first cleavage. Anterior is to the top. Ventral faces readers. Scale bars indicate 10 μm. (C) Cell corpse C3 (white arrow) is engulfed by ABplaapppp (white arrowhead). Yellow arrows and arrowheads indicate C1 and C2 and their corresponding engulfing cells. “0 min” represents when CED-1::GFP is first detected on the budding pseudopods. (D) Time-lapse images of signals around C3 (arrows) during its degradation. “0 min” represents when ABplaapppp fully internalized C3. Arrows indicate C1, C2, and C3 in (a), of which the corresponding engulfing cells are outlined and indicated by arrowheads. Insets in (e) and (l) show an amplified view of the region surrounding C3; arrowheads indicate tubular structures. The yellow arrowhead in (d) depicts the nucleus of ABpraapppa, which has engulfed C2. (o) The relative reporter signal intensity measured from (a) to (n) is plotted over time.(E) Summary of the temporal order of each reporter detected on the surfaces of phagocytic cups and phagosomes. Data represent means obtained from time-lapse recording of multiple C1, C2, and C3 cell corpses (see text for actual numbers). “0 min” represents when budding pseudopods are first detected by CED-1::GFP. 2xFYVE, HGRS-1, RAB-7, and CTNS-1 reporters localize to phagosome surface until the cell corpse is fully degraded. The conversion of pale to deep pink colors of HGRS-1::mRFP1 and CTNS-1::mRFP1 indicates that starting from an initial weak signal, the signal intensity gradually increases.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2267821&req=5

pbio-0060061-g001: PI(3)P and RAB-7 Are Sequentially Enriched on the Surface of Maturing Phagosomes Containing Apoptotic Cells(A) Diagram of phagocytosis and the subsequent degradation of a cell corpse.(B) The two partially redundant and parallel pathways that control cell-corpse removal in C. elegans. Names in parentheses indicate mammalian homologs.(C and D) Time-lapse images of wild-type embryos coexpressing Pced-1ced-1::gfp / Pced-12xFYVE::mrfp1 (C) and Pced-12xFYVE::mrfp1 / Pced-1gfp::rab-7 (D). Recording started at approximately 320 min after first cleavage. Anterior is to the top. Ventral faces readers. Scale bars indicate 10 μm. (C) Cell corpse C3 (white arrow) is engulfed by ABplaapppp (white arrowhead). Yellow arrows and arrowheads indicate C1 and C2 and their corresponding engulfing cells. “0 min” represents when CED-1::GFP is first detected on the budding pseudopods. (D) Time-lapse images of signals around C3 (arrows) during its degradation. “0 min” represents when ABplaapppp fully internalized C3. Arrows indicate C1, C2, and C3 in (a), of which the corresponding engulfing cells are outlined and indicated by arrowheads. Insets in (e) and (l) show an amplified view of the region surrounding C3; arrowheads indicate tubular structures. The yellow arrowhead in (d) depicts the nucleus of ABpraapppa, which has engulfed C2. (o) The relative reporter signal intensity measured from (a) to (n) is plotted over time.(E) Summary of the temporal order of each reporter detected on the surfaces of phagocytic cups and phagosomes. Data represent means obtained from time-lapse recording of multiple C1, C2, and C3 cell corpses (see text for actual numbers). “0 min” represents when budding pseudopods are first detected by CED-1::GFP. 2xFYVE, HGRS-1, RAB-7, and CTNS-1 reporters localize to phagosome surface until the cell corpse is fully degraded. The conversion of pale to deep pink colors of HGRS-1::mRFP1 and CTNS-1::mRFP1 indicates that starting from an initial weak signal, the signal intensity gradually increases.
Mentions: An engulfing cell recognizes an apoptotic cell through phagocytic receptor(s) and extends thin pseudopods around it to generate a phagocytic cup. The fusion of growing pseudopods and scission of a vacuole containing the apoptotic cell from the plasmalemma generate a phagosome inside the host cell (Figure 1A) [1]. The swift engulfment (phagocytosis) and subsequent degradation of apoptotic cells inside phagosomes eliminate dying cells before they release any potentially harmful contents, and actively prevent tissue injury, inflammatory responses, and autoimmune diseases [2]. Inefficient degradation of the components of engulfed apoptotic cells, such as nuclear DNA, similar to inefficient engulfment, results in severe inflammatory and autoimmune responses [3,4].

Bottom Line: We found that mutations in each pathway not only delay or block engulfment, but also delay the degradation of engulfed apoptotic cells.We propose that RAB-7 functions as a downstream effector of the CED-1 pathway to mediate phagolysosome formation.Our work suggests that phagocytic receptors, which were thought to act specifically in initiating engulfment, also control phagosome maturation through the sequential activation of multiple effectors such as dynamin, PI(3)P, and Rab GTPases.

View Article: PubMed Central - PubMed

Affiliation: Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, Texas, United States of America.

ABSTRACT
Apoptotic cells in animals are engulfed by phagocytic cells and subsequently degraded inside phagosomes. To study the mechanisms controlling the degradation of apoptotic cells, we developed time-lapse imaging protocols in developing Caenorhabditis elegans embryos and established the temporal order of multiple events during engulfment and phagosome maturation. These include sequential enrichment on phagocytic membranes of phagocytic receptor cell death abnormal 1 (CED-1), large GTPase dynamin (DYN-1), phosphatidylinositol 3-phosphate (PI(3)P), and the small GTPase RAB-7, as well as the incorporation of endosomes and lysosomes to phagosomes. Two parallel genetic pathways are known to control the engulfment of apoptotic cells in C. elegans. We found that mutations in each pathway not only delay or block engulfment, but also delay the degradation of engulfed apoptotic cells. One of the pathways, composed of CED-1, the adaptor protein CED-6, and DYN-1, controls the rate of enrichment of PI(3)P and RAB-7 on phagosomal surfaces and the formation of phagolysosomes. We further identified an essential role of RAB-7 in promoting the recruitment and fusion of lysosomes to phagosomes. We propose that RAB-7 functions as a downstream effector of the CED-1 pathway to mediate phagolysosome formation. Our work suggests that phagocytic receptors, which were thought to act specifically in initiating engulfment, also control phagosome maturation through the sequential activation of multiple effectors such as dynamin, PI(3)P, and Rab GTPases.

Show MeSH
Related in: MedlinePlus