scholarly journals The Disabled 1 Phosphotyrosine-Binding Domain Binds to the Internalization Signals of Transmembrane Glycoproteins and to Phospholipids

1999 ◽  
Vol 19 (7) ◽  
pp. 5179-5188 ◽  
Author(s):  
Brian W. Howell ◽  
Lorene M. Lanier ◽  
Ronald Frank ◽  
Frank B. Gertler ◽  
Jonathan A. Cooper
Keyword(s):  
Hippocampal Neurons ◽  
System Development ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Nervous System Development ◽  
Amino Terminal ◽  
Density Lipoprotein Receptor ◽  
The Disabled ◽  
Phosphorylated Peptide ◽  
Ptb Domain

ABSTRACT Disabled gene products are important for nervous system development in drosophila and mammals. In mice, the Dab1 protein is thought to function downstream of the extracellular protein Reln during neuronal positioning. The structures of Dab proteins suggest that they mediate protein-protein or protein-membrane docking functions. Here we show that the amino-terminal phosphotyrosine-binding (PTB) domain of Dab1 binds to the transmembrane glycoproteins of the amyloid precursor protein (APP) and low-density lipoprotein receptor families and the cytoplasmic signaling protein Ship. Dab1 associates with the APP cytoplasmic domain in transfected cells and is coexpressed with APP in hippocampal neurons. Screening of a set of altered peptide sequences showed that the sequence GYXNPXY present in APP family members is an optimal binding sequence, with approximately 0.5 μM affinity. Unlike other PTB domains, the Dab1 PTB does not bind to tyrosine-phosphorylated peptide ligands. The PTB domain also binds specifically to phospholipid bilayers containing phosphatidylinositol 4P (PtdIns4P) or PtdIns4,5P2 in a manner that does not interfere with protein binding. We propose that the PTB domain permits Dab1 to bind specifically to transmembrane proteins containing an NPXY internalization signal.

scholarly journals The apoptotic engulfment protein Ced-6 participates in clathrin-mediated yolk uptake in Drosophila egg chambers

2012 ◽  
Vol 23 (9) ◽  
pp. 1742-1764 ◽  
Author(s):  
Anupma Jha ◽  
Simon C. Watkins ◽  
Linton M. Traub
Keyword(s):  
Ovarian Follicle ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Biochemical Properties ◽  
Ovarian Follicle Development ◽  
Density Lipoprotein Receptor ◽  
Egg Chambers ◽  
Clathrin Adaptor ◽  
Ptb Domain

Clathrin-mediated endocytosis and phagocytosis are both selective surface internalization processes but have little known mechanistic similarity or interdependence. Here we show that the phosphotyrosine-binding (PTB) domain protein Ced-6, a well-established phagocytosis component that operates as a transducer of so-called “eat-me” signals during engulfment of apoptotic cells and microorganisms, is expressed in the female Drosophila germline and that Ced-6 expression correlates with ovarian follicle development. Ced-6 exhibits all the known biochemical properties of a clathrin-associated sorting protein, yet ced-6–null flies are semifertile despite massive accumulation of soluble yolk precursors in the hemolymph. This is because redundant sorting signals within the cytosolic domain of the Drosophila vitellogenin receptor Yolkless, a low density lipoprotein receptor superfamily member, occur; a functional atypical dileucine signal binds to the endocytic AP-2 clathrin adaptor directly. Nonetheless, the Ced-6 PTB domain specifically recognizes the noncanonical Yolkless FXNPXA sorting sequence and in HeLa cells promotes the rapid, clathrin-dependent uptake of a Yolkless chimera lacking the distal dileucine signal. Ced-6 thus operates in vivo as a clathrin adaptor. Because the human Ced-6 orthologue GULP similarly binds to clathrin machinery, localizes to cell surface clathrin-coated structures, and is enriched in placental clathrin-coated vesicles, new possibilities for Ced-6/Gulp operation during phagocytosis must be considered.

scholarly journals Initiating protease with modular domains interacts with β-glucan recognition protein to trigger innate immune response in insects

2015 ◽  
Vol 112 (45) ◽  
pp. 13856-13861 ◽  
Author(s):  
Daisuke Takahashi ◽  
Brandon L. Garcia ◽  
Michael R. Kanost
Keyword(s):  
Immune Responses ◽  
Molecular Basis ◽  
Molecular Mechanisms ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Dependent Manner ◽  
Calcium Dependent ◽  
Amino Terminal ◽  
Density Lipoprotein Receptor ◽  
Recognition Protein

The autoactivation of an initiating serine protease upon binding of pattern recognition proteins to pathogen surfaces is a crucial step in eliciting insect immune responses such as the activation of Toll and prophenoloxidase pathways. However, the molecular mechanisms responsible for autoactivation of the initiating protease remains poorly understood. Here, we investigated the molecular basis for the autoactivation of hemolymph protease 14 (HP14), an initiating protease in hemolymph of Manduca sexta, upon the binding of β-1,3-glucan by its recognition protein, βGRP2. Biochemical analysis using HP14 zymogen (proHP14), βGRP2, and the recombinant proteins as truncated forms showed that the amino-terminal modular low-density lipoprotein receptor class A (LA) domains within HP14 are required for proHP14 autoactivation that is stimulated by its interaction with βGRP2. Consistent with this result, recombinant LA domains inhibit the activation of proHP14 and prophenoloxidase, likely by competing with the interaction between βGRP2 and LA domains within proHP14. Using surface plasmon resonance, we demonstrated that immobilized LA domains directly interact with βGRP2 in a calcium-dependent manner and that high-affinity interaction requires the C-terminal glucanase-like domain of βGRP2. Importantly, the affinity of LA domains for βGRP2 increases nearly 100-fold in the presence of β-1,3-glucan. Taken together, these results present the first experimental evidence to our knowledge that LA domains of an insect modular protease and glucanase-like domains of a βGRP mediate their interaction, and that this binding is essential for the protease autoactivation. Thus, our study provides important insight into the molecular basis underlying the initiation of protease cascade in insect immune responses.

scholarly journals Polarized Traffic of LRP1 Involves AP1B and SNX17 Operating on Y-dependent Sorting Motifs in Different Pathways

2009 ◽  
Vol 20 (1) ◽  
pp. 481-497 ◽  
Author(s):  
Maribel Donoso ◽  
Jorge Cancino ◽  
Jiyeon Lee ◽  
Peter van Kerkhof ◽  
Claudio Retamal ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Low Density Lipoprotein ◽  
Basolateral Membrane ◽  
Density Lipoprotein ◽  
Recycling Endosome ◽  
Sorting Nexin ◽  
Sorting Signals ◽  
Density Lipoprotein Receptor ◽  
Npxy Motif ◽  
Lipoprotein Receptor Related Protein

Low-density lipoprotein receptor–related protein 1 (LRP1) is an endocytic recycling receptor with two cytoplasmic tyrosine-based basolateral sorting signals. Here we show that during biosynthetic trafficking LRP1 uses AP1B adaptor complex to move from a post-TGN recycling endosome (RE) to the basolateral membrane. Then it recycles basolaterally from the basolateral sorting endosome (BSE) involving recognition by sorting nexin 17 (SNX17). In the biosynthetic pathway, Y29 but not N26 from a proximal NPXY directs LRP1 basolateral sorting from the TGN. A N26A mutant revealed that this NPXY motif recognized by SNX17 is required for the receptor's exit from BSE. An endocytic Y63ATL66 motif also functions in basolateral recycling, in concert with an additional endocytic motif (LL86,87), by preventing LRP1 entry into the transcytotic apical pathway. All this sorting information operates similarly in hippocampal neurons to mediate LRP1 somatodendritic distribution regardless of the absence of AP1B in neurons. LRP1 basolateral distribution results then from spatially and temporally segregation steps mediated by recognition of distinct tyrosine-based motifs. We also demonstrate a novel function of SNX17 in basolateral/somatodendritic recycling from a different compartment than AP1B endosomes.

The Role of the Low-Density Lipoprotein Receptor-Related Protein (LRP) in the Plasma Clearance and Liver Uptake of Recombinant Single-chain Urokinase-type Plasminogen Activator in Rats

1997 ◽  
Vol 77 (04) ◽  
pp. 710-717 ◽  
Author(s):  
Marieke E van der Kaaden ◽  
Dingeman C Rijken ◽  
J Kar Kruijt ◽  
Theo J C van Berkel ◽  
Johan Kuiper
Keyword(s):  
Plasminogen Activator ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Single Chain ◽  
Liver Uptake ◽  
Type Plasminogen Activator ◽  
Parenchymal Liver ◽  
Urokinase Type Plasminogen Activator ◽  
Density Lipoprotein Receptor

SummaryUrokinase-type plasminogen activator (u-PA) is used as a thrombolytic agent in the treatment of acute myocardial infarction. In vitro, recombinant single-chain u-PA (rscu-PA) expressed in E.coli is recognized by the Low-Density Lipoprotein Receptor-related Protein (LRP) on rat parenchymal liver cells. In this study we investigated the role of LRP in the liver uptake and plasma clearance of rscu-PA in rats. A preinjection of the LRP inhibitor GST-RAP reduced the maximal liver uptake of 125I-rscu-PA at 5 min after injection from 50 to 30% of the injected dose and decreased the clearance of rscu-PA from 2.37 ml/min to 1.58 ml/min. Parenchymal, Kupffer and endothelial cells were responsible for 40, 50 and 10% of the liver uptake, respectively. The reduction in liver uptake of rscu-PA by the preinjection of GST-RAP was caused by a 91 % and 62% reduction in the uptake by parenchymal and Kupffer cells, respectively. In order to investigate the part of rscu-PA that accounted for the interaction with LRP, experiments were performed with a mutant of rscu-PA lacking residues 11-135 (= deltal25- rscu-PA). Deletion of residues 11-135 resulted in a 80% reduction in liver uptake and a 2.4 times slower clearance (0.97 ml/min). The parenchymal, Kupffer and endothelial cells were responsible for respectively 60, 33 and 7% of the liver uptake of 125I-deltal25-rscu-PA. Preinjection of GST-RAP completely reduced the liver uptake of delta 125-rscu-PA and reduced its clearance to 0.79 ml/min. Treatment of isolated Kupffer cells with PI-PLC reduced the binding of rscu-PA by 40%, suggesting the involvement of the urokinase-type Plasminogen Activator Receptor (u-PAR) in the recognition of rscu-PA. Our results demonstrate that in vivo LRP is responsible for more than 90% of the parenchymal liver cell mediated uptake of rscu-PA and for 60% of the Kupffer cell interaction. It is also suggested that u-PAR is involved in the Kupffer cell recognition of rscu-PA.

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