- Email: [email protected]
Phagocytosis in C. elegans: CED-1 reveals its secrets
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News & Comment
TRENDS in Cell Biology Vol.11 No.4 April 2001
AP-3: what color’s your coat? Signal-mediated delivery of proteins in several intracellular trafficking pathways requires the adaptor complexes AP-1, AP-2 and AP-3. Adaptors are heterotetrameric protein complexes that serve in cargo sorting through binding to di-leucine or tyrosinebased signals present in the cytoplasmic tails of cargo proteins. Adaptors team up with cytoplasmic coat proteins to effect inclusion of cargo into transport vesicles. AP-1 associates with vesicles budding from the trans-side of the Golgi complex, and AP-2 with endocytic vesicles budding from the plasma membrane. Both AP-1 and AP-2 cooperate with clathrin, a complex of three heavy chains and three light chains, which polymerizes to form the scaffold of the coat. The AP-3 adaptor mediates selective transport to lysosomes and lysosome-related organelles and has important biological roles in organisms as diverse as humans, flies, mice and yeast. In yeast, AP-3 function is required for the ALP (alkaline phosphatase) pathway to the vacuole. Mutations in AP-3 subunits in mice result in coat color defects and bleeding disorders and, in Drosophila, result in defects in eye pigmentation. In humans, mutations in the AP-3 β3A subunit cause an inherited disorder, Hermansky–Pudlak syndrome (HPS), in which patients show deficiencies in skin and eye pigmentation and a complete lack of dense granules in platelets, resulting in impaired blood clotting.
An ongoing controversy involves the identity of the coat protein(s) the AP-3 adaptor utilizes to effect vesicle formation in vivo. Drake et al. demonstrated in vitro that AP-3 and clathrin, together with ARF-GTP, can nucleate clathrin-coated buds and vesicles from synthetic liposomes1. Furthermore, clathrin can associate with AP-3 in vitro through a ‘clathrin box’ motif present in the mammalian β3 subunit. These observations are consistent with the notion that AP-3 and clathrin team up to mediate transport in this pathway. Intriguingly, however, there is strong evidence from yeast that AP-3 does not functionally associate with clathrin in this organism2,3. For instance, the phenotypes of clathrin and AP-3 mutants are dissimilar, and clathrin does not copurify with AP-3-coated vesicles. Instead, Rehling et al. demonstrated that mutants in the yeast VPS41 gene, one of ~45 genes involved in Golgi-to-vacuole transport, display ALP-specific sorting defects, and that Vps41p can bind to the yeast AP-3 δ subunit in vitro2. This suggests that AP-3 cooperates with Vps41p, rather than clathrin, in the formation of ALP pathway transport vesicles in yeast. Darsow et al.3 have now characterized novel vps41 alleles that came from a screen for additional components of the ALP pathway. These mutants show severe ALP sorting defects and defects in ALP vesicle formation but, unlike the vps41 null-allele, display a normal vacuolar morphology.
This is important because Vps41p is also a component of the docking and fusion machinery at the vacuole, the C-Vps–HOPS complex. Thus, Vps41p might have two clearly separable functions. Darsow et al. further show that a conserved N-terminal domain on Vps41p mediates its binding to the AP-3 δ subunit, whereas a C-terminal region on Vps41p, containing a clathrin heavy chain repeat, mediates homooligomerization of Vps41p3. The latter is a feature expected of a coat protein. Clearly, AP-3 has two good candidates for ‘partners’ to choose from. A definitive answer to the ‘AP-3 coat question’ will require characterization of mammalian orthologs of Vps41p, followed by a careful study of the colocalization and interactions of AP-3 with Vps41p and/or clathrin. 1 Drake, M. et al. (2000) The assembly of AP-3 adaptor complex-containing clathrin-coated vesicles on synthetic liposomes. Mol. Biol. Cell 11, 3723–3736 2 Rehling, P. et al. (1999) Formation of AP-3 transport intermediates requires Vps41p function. Nat. Cell Biol. 1, 346–353 3 Darsow, T. et al. (2001) Vps41p function in the alkaline phosphatase pathway requires homo-oligomerization and interaction with AP-3 through two distinct domains. Mol. Biol. Cell 12, 37–51
Rainer Duden [email protected]
Phagocytosis in C. elegans : CED-1 reveals its secrets During Caenorhabditis elegans development, exactly 131 somatic cells undergo apoptosis, their corpses being engulfed by neighbouring cells. This phenomenon provides a powerful tool to study phagocytosis as mutants that are unable to engulf cell corpses (ced mutants) are readily identifiable. So far, six separate genes required for engulfment have been identified and assigned to two, partially redundant, signal pathways; ced-2, ced-5 and ced-10 in one, ced-1, ced-6 and ced-7 in the other. With five of the six genes cloned, the identification of CED-1 by Zhou et al.1 completes the list – at least for now. CED-1 is a transmembrane receptor that binds to cell corpses through its extracellular domain. Intriguingly, this
binding requires the function of CED-7, a protein previously shown to resemble an ATP-binding cassette (ABC) transporter and encoded by the only ced gene that functions both in dying and in engulfing cells. The function of CED-7 remains unknown, although the authors suggest that it might present a cell-corpse ligand to CED-1. However, the intracellular domain of CED-1, which shows no overall homology to other proteins but contains an NPXY and a YXXL motif, is perhaps its most fascinating feature. Both motifs are necessary for the function of CED-1 and are partially redundant with each other. NPXY motifs bind to phosphotyrosinebinding (PTB) domains. As CED-6 contains a PTB, and is in the same functional group
as CED-1, it is possible that CED-6 relays signals from CED-1, although the authors have so far failed to detect binding between the two proteins. YXXL motifs are phosphorylation sites for tyrosine kinases and are found in mammalian Fcγ-receptors, which mediate engulfment of IgG-coated particles. CED-1 is a fascinating evolutionary link – a single molecule that combines motifs found in separate proteins in higher organisms. Now the question is: how does it work? 1 Zhou, Z. et al. (2001) CED-1 is a transmembrane receptor that mediates cell corpse engulfment in C. elegans. Cell 104, 43–56
Robin C. May [email protected]
http://tcb.trends.com 0962-8924/01/$ – see front matter © 2001 Elsevier Science Ltd. All rights reserved.
News & Comment
TRENDS in Cell Biology Vol.11 No.4 April 2001
AP-3: what color’s your coat? Signal-mediated delivery of proteins in several intracellular trafficking pathways requires the adaptor complexes AP-1, AP-2 and AP-3. Adaptors are heterotetrameric protein complexes that serve in cargo sorting through binding to di-leucine or tyrosinebased signals present in the cytoplasmic tails of cargo proteins. Adaptors team up with cytoplasmic coat proteins to effect inclusion of cargo into transport vesicles. AP-1 associates with vesicles budding from the trans-side of the Golgi complex, and AP-2 with endocytic vesicles budding from the plasma membrane. Both AP-1 and AP-2 cooperate with clathrin, a complex of three heavy chains and three light chains, which polymerizes to form the scaffold of the coat. The AP-3 adaptor mediates selective transport to lysosomes and lysosome-related organelles and has important biological roles in organisms as diverse as humans, flies, mice and yeast. In yeast, AP-3 function is required for the ALP (alkaline phosphatase) pathway to the vacuole. Mutations in AP-3 subunits in mice result in coat color defects and bleeding disorders and, in Drosophila, result in defects in eye pigmentation. In humans, mutations in the AP-3 β3A subunit cause an inherited disorder, Hermansky–Pudlak syndrome (HPS), in which patients show deficiencies in skin and eye pigmentation and a complete lack of dense granules in platelets, resulting in impaired blood clotting.
An ongoing controversy involves the identity of the coat protein(s) the AP-3 adaptor utilizes to effect vesicle formation in vivo. Drake et al. demonstrated in vitro that AP-3 and clathrin, together with ARF-GTP, can nucleate clathrin-coated buds and vesicles from synthetic liposomes1. Furthermore, clathrin can associate with AP-3 in vitro through a ‘clathrin box’ motif present in the mammalian β3 subunit. These observations are consistent with the notion that AP-3 and clathrin team up to mediate transport in this pathway. Intriguingly, however, there is strong evidence from yeast that AP-3 does not functionally associate with clathrin in this organism2,3. For instance, the phenotypes of clathrin and AP-3 mutants are dissimilar, and clathrin does not copurify with AP-3-coated vesicles. Instead, Rehling et al. demonstrated that mutants in the yeast VPS41 gene, one of ~45 genes involved in Golgi-to-vacuole transport, display ALP-specific sorting defects, and that Vps41p can bind to the yeast AP-3 δ subunit in vitro2. This suggests that AP-3 cooperates with Vps41p, rather than clathrin, in the formation of ALP pathway transport vesicles in yeast. Darsow et al.3 have now characterized novel vps41 alleles that came from a screen for additional components of the ALP pathway. These mutants show severe ALP sorting defects and defects in ALP vesicle formation but, unlike the vps41 null-allele, display a normal vacuolar morphology.
This is important because Vps41p is also a component of the docking and fusion machinery at the vacuole, the C-Vps–HOPS complex. Thus, Vps41p might have two clearly separable functions. Darsow et al. further show that a conserved N-terminal domain on Vps41p mediates its binding to the AP-3 δ subunit, whereas a C-terminal region on Vps41p, containing a clathrin heavy chain repeat, mediates homooligomerization of Vps41p3. The latter is a feature expected of a coat protein. Clearly, AP-3 has two good candidates for ‘partners’ to choose from. A definitive answer to the ‘AP-3 coat question’ will require characterization of mammalian orthologs of Vps41p, followed by a careful study of the colocalization and interactions of AP-3 with Vps41p and/or clathrin. 1 Drake, M. et al. (2000) The assembly of AP-3 adaptor complex-containing clathrin-coated vesicles on synthetic liposomes. Mol. Biol. Cell 11, 3723–3736 2 Rehling, P. et al. (1999) Formation of AP-3 transport intermediates requires Vps41p function. Nat. Cell Biol. 1, 346–353 3 Darsow, T. et al. (2001) Vps41p function in the alkaline phosphatase pathway requires homo-oligomerization and interaction with AP-3 through two distinct domains. Mol. Biol. Cell 12, 37–51
Rainer Duden [email protected]
Phagocytosis in C. elegans : CED-1 reveals its secrets During Caenorhabditis elegans development, exactly 131 somatic cells undergo apoptosis, their corpses being engulfed by neighbouring cells. This phenomenon provides a powerful tool to study phagocytosis as mutants that are unable to engulf cell corpses (ced mutants) are readily identifiable. So far, six separate genes required for engulfment have been identified and assigned to two, partially redundant, signal pathways; ced-2, ced-5 and ced-10 in one, ced-1, ced-6 and ced-7 in the other. With five of the six genes cloned, the identification of CED-1 by Zhou et al.1 completes the list – at least for now. CED-1 is a transmembrane receptor that binds to cell corpses through its extracellular domain. Intriguingly, this
binding requires the function of CED-7, a protein previously shown to resemble an ATP-binding cassette (ABC) transporter and encoded by the only ced gene that functions both in dying and in engulfing cells. The function of CED-7 remains unknown, although the authors suggest that it might present a cell-corpse ligand to CED-1. However, the intracellular domain of CED-1, which shows no overall homology to other proteins but contains an NPXY and a YXXL motif, is perhaps its most fascinating feature. Both motifs are necessary for the function of CED-1 and are partially redundant with each other. NPXY motifs bind to phosphotyrosinebinding (PTB) domains. As CED-6 contains a PTB, and is in the same functional group
as CED-1, it is possible that CED-6 relays signals from CED-1, although the authors have so far failed to detect binding between the two proteins. YXXL motifs are phosphorylation sites for tyrosine kinases and are found in mammalian Fcγ-receptors, which mediate engulfment of IgG-coated particles. CED-1 is a fascinating evolutionary link – a single molecule that combines motifs found in separate proteins in higher organisms. Now the question is: how does it work? 1 Zhou, Z. et al. (2001) CED-1 is a transmembrane receptor that mediates cell corpse engulfment in C. elegans. Cell 104, 43–56
Robin C. May [email protected]
http://tcb.trends.com 0962-8924/01/$ – see front matter © 2001 Elsevier Science Ltd. All rights reserved.