scholarly journals The Extracellular Linker of pro-Neuregulin-α2c Is Required for Efficient Sorting and Juxtacrine Function

2007 ◽  
Vol 18 (2) ◽  
pp. 380-393 ◽  
Author(s):  
Juan C. Montero ◽  
Ruth Rodríguez-Barrueco ◽  
Laura Yuste ◽  
Pedro P. Juanes ◽  
Joana Borges ◽  
...  
Keyword(s):  
Biological Activity ◽  
Plasma Membrane ◽  
Cell Surface ◽  
Signal Sequence ◽  
Transmembrane Proteins ◽  
Biologically Active ◽  
Erbb Receptors ◽  
Animal Physiology ◽  
Juxtamembrane Region ◽  
In Trans

The neuregulins (NRGs) play important roles in animal physiology, and their disregulation has been linked to diseases such as cancer or schizophrenia. The NRGs may be produced as transmembrane proteins (proNRGs), even though they lack an N-terminal signal sequence. This raises the question of how NRGs are sorted to the plasma membrane. It is also unclear whether in their transmembrane state, the NRGs are biologically active. During studies aimed at solving these questions, we found that deletion of the extracellular juxtamembrane region termed the linker, decreased cell surface exposure of the mutant proNRGΔLinker, and caused its entrapment at the cis-Golgi. We also found that cell surface–exposed transmembrane NRG forms retain biological activity. Thus, a mutant whose cleavage is impaired but is correctly sorted to the plasma membrane activated ErbB receptors in trans and also stimulated proliferation. Because the linker is implicated in surface sorting and the regulation of the cleavage of transmembrane NRGs, our data indicate that this region exerts multiple important roles in the physiology of NRGs.

scholarly journals Transautocrine Signaling by Membrane Neuregulins Requires Cell Surface Targeting, Which Is Controlled by Multiple Domains

2011 ◽  
Vol 286 (27) ◽  
pp. 24350-24363 ◽  
Author(s):  
Juan Carlos Montero ◽  
Ruth Rodríguez-Barrueco ◽  
Atanasio Pandiella
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Signal Sequence ◽  
Biologically Active ◽  
Physical Contact ◽  
Hydrophobic Region ◽  
Activating Receptors ◽  
Egf Family ◽  
Multiple Domains ◽  
Membrane Anchoring

The neuregulins (NRGs) play important roles in animal development and homeostasis, and their deregulation has been linked to diseases such as cancer and schizophrenia. The NRGs belong to the epidermal growth factor (EGF) family of transmembrane growth factors. Although NRGs may be synthesized as transmembrane proteins (the pro-NRGs), some of them lack an N-terminal signal sequence, raising the question of how these pro-NRGs are directed to the plasma membrane. Here we have explored the domains of pro-NRGs that are required for their membrane anchoring, cell surface exposure, and biological activity. We show that an internal hydrophobic region acts as a membrane-anchoring domain, but other regions of pro-NRG are required for proper sorting to the plasma membrane. Using mutants that are located in different subcellular compartments, we show that only plasma membrane-exposed pro-NRG is biologically active. At this location, the pro-NRGs may act as transautocrine molecules (i.e. as membrane factors able to activate receptors present in cells that are in physical contact with the pro-NRG-producing cells (in trans) or capable of activating receptors present in the pro-NRG-producing cells (in cis)).

scholarly journals Biologically active LIL proteins built with minimal chemical diversity

2015 ◽  
Vol 112 (34) ◽  
pp. E4717-E4725 ◽  
Author(s):  
Erin N. Heim ◽  
Jez L. Marston ◽  
Ross S. Federman ◽  
Anne P. B. Edwards ◽  
Alexander G. Karabadzhak ◽  
...  
Keyword(s):  
Amino Acids ◽  
Biological Activity ◽  
Transmembrane Domain ◽  
Transmembrane Proteins ◽  
Chemical Diversity ◽  
Biologically Active ◽  
Cellular Context ◽  
Hydrophobic Amino Acids ◽  
Β Receptor ◽  
Specific Sequences

We have constructed 26-amino acid transmembrane proteins that specifically transform cells but consist of only two different amino acids. Most proteins are long polymers of amino acids with 20 or more chemically distinct side-chains. The artificial transmembrane proteins reported here are the simplest known proteins with specific biological activity, consisting solely of an initiating methionine followed by specific sequences of leucines and isoleucines, two hydrophobic amino acids that differ only by the position of a methyl group. We designate these proteins containing leucine (L) and isoleucine (I) as LIL proteins. These proteins functionally interact with the transmembrane domain of the platelet-derived growth factor β-receptor and specifically activate the receptor to transform cells. Complete mutagenesis of these proteins identified individual amino acids required for activity, and a protein consisting solely of leucines, except for a single isoleucine at a particular position, transformed cells. These surprisingly simple proteins define the minimal chemical diversity sufficient to construct proteins with specific biological activity and change our view of what can constitute an active protein in a cellular context.

scholarly journals Direct stimulation of cells expressing receptors for macrophage colony- stimulating factor (CSF-1) by a plasma membrane-bound precursor of human CSF-1

Blood ◽  
1990 ◽  
Vol 76 (7) ◽  
pp. 1308-1314 ◽  
Author(s):  
J Stein ◽  
GV Borzillo ◽  
CW Rettenmier
Keyword(s):  
Plasma Membrane ◽  
Growth Factor ◽  
Cell Surface ◽  
Mononuclear Phagocytes ◽  
Biologically Active ◽  
Colony Stimulating Factor ◽  
Membrane Bound ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Secreted forms of macrophage colony-stimulating factor (M-CSF or CSF-1) are generated by proteolytic cleavage of membrane-bound glycoprotein precursors. Alternatively spliced transcripts of the human CSF-1 gene encode at least two different transmembrane precursors that are differentially processed in mammalian expression systems. The larger precursor rapidly undergoes proteolysis to yield the secreted growth factor and does not give rise to forms of CSF-1 detected on the cell surface. By contrast, the smaller human CSF-1 precursor is stably expressed on the plasma membrane where it is inefficiently cleaved to release a soluble molecule. To determine whether the smaller precursor is biologically active on the cell surface, mouse NIH-3T3 fibroblasts expressing the different forms of human CSF-1 were killed by chemical fixation and tested for their ability to support the proliferation of cells that require this growth factor. Only fixed cells expressing human CSF-1 precursors on their surface stimulated the growth in vitro of a murine macrophage cell line or normal mouse bone marrow-derived mononuclear phagocytes. The ability of these nonviable fibroblasts to induce the proliferation of CSF-1-dependent cells was not mediated by release of soluble growth factor, required direct contact with the target cells, and was blocked by neutralizing antiserum to CSF-1. These results demonstrate that the cell surface form of the human CSF-1 precursor is biologically active and indicate that plasma membrane- bound growth factors can functionally interact with receptor-bearing targets by direct cell-cell contact.

The biophysics of ligand–receptor interactions

1980 ◽  
Vol 13 (2) ◽  
pp. 201-230 ◽  
Author(s):  
Charles DeLisi
Keyword(s):  
Biological Activity ◽  
Plasma Membrane ◽  
Cell Surface ◽  
Background Noise ◽  
Information Transfer ◽  
Receptor Interactions ◽  
A Cell ◽  
Cell Surface Glycoproteins ◽  
High Degree ◽  
To Receive

For the cells of an organism to act in the coordinated fashion necessary for complex functioning, they must be able to receive and transmit information. Information transfer is mediated by molecules released by the cells and may be local, as in the case of neurotransmitters, or long range, as in the case of hormones. It is apparent, however, that irrespective of the range of interaction, a cell must be able to distinguish, with a high degree of precision, the signals relevant to it from an enormous flow of background noise.Molecular recognition at the cell surface is mediated by receptors: cell surface glycoproteins that usually form an integral part of the plasma membrane (see, for example, Cuatrecasas & Greaves, 1978). Typically, receptors bind the ligands they are designed to recognize with affinities of the order of 108 M-1, and they translate that interaction into a sequence of signals that ultimately lead to biological activity.

scholarly journals Using selenomethionyl derivatives to assign sequence in low-resolution structures of the AP2 clathrin adaptor

2016 ◽  
Vol 72 (3) ◽  
pp. 336-345 ◽  
Author(s):  
Bernard T. Kelly ◽  
Stephen C. Graham ◽  
David J. Owen
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Binding Site ◽  
Point Mutations ◽  
Transmembrane Proteins ◽  
Low Resolution ◽  
Transport Vesicles ◽  
Transport Vesicle ◽  
Clathrin Adaptor ◽  
Sequence Assignment

Selenomethionine incorporation is a powerful technique for assigning sequence to regions of electron density at low resolution. Genetic introduction of methionine point mutations and the subsequent preparation and crystallization of selenomethionyl derivatives permits unambiguous sequence assignment by enabling the placement of the anomalous scatterers (Se atoms) thus introduced. Here, the use of this approach in the assignment of sequence in a part of the AP2 clathrin adaptor complex that is responsible for clathrin binding is described. AP2 plays a pivotal role in clathrin-mediated endocytosis, a tightly regulated process in which cell-surface transmembrane proteins are internalized from the plasma membrane by incorporation into lipid-enclosed transport vesicles. AP2 binds cargo destined for internalization and recruits clathrin, a large trimeric protein that helps to deform the membrane to produce the transport vesicle. By selenomethionine labelling of point mutants, it was shown that the clathrin-binding site is buried within a deep cleft of the AP2 complex. A membrane-stimulated conformational change in AP2 releases the clathrin-binding site from autoinhibition, thereby linking clathrin recruitment to membrane localization.

scholarly journals A Short N-Terminal Peptide Motif on Flavivirus Nonstructural Protein NS1 Modulates Cellular Targeting and Immune Recognition

2010 ◽  
Vol 84 (18) ◽  
pp. 9516-9532 ◽  
Author(s):  
Soonjeon Youn ◽  
Hyelim Cho ◽  
Daved H. Fremont ◽  
Michael S. Diamond
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Viral Replication ◽  
Signal Sequence ◽  
Nonstructural Protein ◽  
Immune Recognition ◽  
Peptide Motif ◽  
Membrane Expression ◽  
Cellular Targeting ◽  
Plasma Membrane Expression

ABSTRACT Flavivirus NS1 is a versatile nonstructural glycoprotein, with intracellular NS1 functioning as an essential cofactor for viral replication and cell surface and secreted NS1 antagonizing complement activation. Even though NS1 has multiple functions that contribute to virulence, the genetic determinants that regulate the spatial distribution of NS1 in cells among different flaviviruses remain uncharacterized. Here, by creating a panel of West Nile virus-dengue virus (WNV-DENV) NS1 chimeras and site-specific mutants, we identified a novel, short peptide motif immediately C-terminal to the signal sequence cleavage position that regulates its transit time through the endoplasmic reticulum and differentially directs NS1 for secretion or plasma membrane expression. Exchange of two amino acids within this motif reciprocally changed the cellular targeting pattern of DENV or WNV NS1. For WNV, this substitution also modulated infectivity and antibody-induced phagocytosis of infected cells. Analysis of a mutant lacking all three conserved N-linked glycosylation sites revealed an independent requirement of N-linked glycans for secretion but not for plasma membrane expression of WNV NS1. Collectively, our experiments define the requirements for cellular targeting of NS1, with implications for the protective host responses, immune antagonism, and association with the host cell sorting machinery. These studies also suggest a link between the effects of NS1 on viral replication and the levels of secreted or cell surface NS1.

scholarly journals H-ras but Not K-ras Traffics to the Plasma Membrane through the Exocytic Pathway

2000 ◽  
Vol 20 (7) ◽  
pp. 2475-2487 ◽  
Author(s):  
Ann Apolloni ◽  
Ian A. Prior ◽  
Margaret Lindsay ◽  
Robert G. Parton ◽  
John F. Hancock
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Brefeldin A ◽  
Biologically Active ◽  
Membrane Targeting ◽  
Ras Proteins ◽  
Golgi Region ◽  
Inner Surface ◽  
Exocytic Pathway ◽  
Caax Motif

ABSTRACT Ras proteins must be localized to the inner surface of the plasma membrane to be biologically active. The motifs that effect Ras plasma membrane targeting consist of a C-terminal CAAX motif plus a second signal comprising palmitoylation of adjacent cysteine residues or the presence of a polybasic domain. In this study, we examined how Ras proteins access the cell surface after processing of the CAAX motif is completed in the endoplasmic reticulum (ER). We show that palmitoylated CAAX proteins, in addition to being localized at the plasma membrane, are found throughout the exocytic pathway and accumulate in the Golgi region when cells are incubated at 15°C. In contrast, polybasic CAAX proteins are found only at the cell surface and not in the exocytic pathway. CAAX proteins which lack a second signal for plasma membrane targeting accumulate in the ER and Golgi. Brefeldin A (BFA) significantly inhibits the plasma membrane accumulation of newly synthesized, palmitoylated CAAX proteins without inhibiting their palmitoylation. BFA has no effect on the trafficking of polybasic CAAX proteins. We conclude that H-ras and K-ras traffic to the cell surface through different routes and that the polybasic domain is a sorting signal diverting K-Ras out of the classical exocytic pathway proximal to the Golgi. Farnesylated Ras proteins that lack a polybasic domain reach the Golgi but require palmitoylation in order to traffic further to the cell surface. These data also indicate that a Ras palmitoyltransferase is present in an early compartment of the exocytic pathway.

scholarly journals Direct stimulation of cells expressing receptors for macrophage colony- stimulating factor (CSF-1) by a plasma membrane-bound precursor of human CSF-1

Blood ◽  
1990 ◽  
Vol 76 (7) ◽  
pp. 1308-1314 ◽  
Author(s):  
J Stein ◽  
GV Borzillo ◽  
CW Rettenmier
Keyword(s):  
Plasma Membrane ◽  
Growth Factor ◽  
Cell Surface ◽  
Mononuclear Phagocytes ◽  
Biologically Active ◽  
Colony Stimulating Factor ◽  
Membrane Bound ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

Abstract Secreted forms of macrophage colony-stimulating factor (M-CSF or CSF-1) are generated by proteolytic cleavage of membrane-bound glycoprotein precursors. Alternatively spliced transcripts of the human CSF-1 gene encode at least two different transmembrane precursors that are differentially processed in mammalian expression systems. The larger precursor rapidly undergoes proteolysis to yield the secreted growth factor and does not give rise to forms of CSF-1 detected on the cell surface. By contrast, the smaller human CSF-1 precursor is stably expressed on the plasma membrane where it is inefficiently cleaved to release a soluble molecule. To determine whether the smaller precursor is biologically active on the cell surface, mouse NIH-3T3 fibroblasts expressing the different forms of human CSF-1 were killed by chemical fixation and tested for their ability to support the proliferation of cells that require this growth factor. Only fixed cells expressing human CSF-1 precursors on their surface stimulated the growth in vitro of a murine macrophage cell line or normal mouse bone marrow-derived mononuclear phagocytes. The ability of these nonviable fibroblasts to induce the proliferation of CSF-1-dependent cells was not mediated by release of soluble growth factor, required direct contact with the target cells, and was blocked by neutralizing antiserum to CSF-1. These results demonstrate that the cell surface form of the human CSF-1 precursor is biologically active and indicate that plasma membrane- bound growth factors can functionally interact with receptor-bearing targets by direct cell-cell contact.

scholarly journals Presenilin-1 affects trafficking and processing of βAPP and is targeted in a complex with nicastrin to the plasma membrane

2002 ◽  
Vol 158 (3) ◽  
pp. 551-561 ◽  
Author(s):  
Christoph Kaether ◽  
Sven Lammich ◽  
Dieter Edbauer ◽  
Michaela Ertl ◽  
Jens Rietdorf ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Secretory Pathway ◽  
Amyloid Β ◽  
Biologically Active ◽  
Dual Function ◽  
Amyloid Β Peptide ◽  
Secretase Activity ◽  
Β Amyloid ◽  
Aβ Production

Amyloid β-peptide (Aβ) is generated by the consecutive cleavages of β- and γ-secretase. The intramembraneous γ-secretase cleavage critically depends on the activity of presenilins (PS1 and PS2). Although there is evidence that PSs are aspartyl proteases with γ-secretase activity, it remains controversial whether their subcellular localization overlaps with the cellular sites of Aβ production. We now demonstrate that biologically active GFP-tagged PS1 as well as endogenous PS1 are targeted to the plasma membrane (PM) of living cells. On the way to the PM, PS1 binds to nicastrin (Nct), an essential component of the γ-secretase complex. This complex is targeted through the secretory pathway where PS1-bound Nct becomes endoglycosidase H resistant. Moreover, surface-biotinylated Nct can be coimmunoprecipitated with PS1 antibodies, demonstrating that this complex is located to cellular sites with γ-secretase activity. Inactivating PS1 or PS2 function by mutagenesis of one of the critical aspartate residues or by γ-secretase inhibitors results in delayed reinternalization of the β-amyloid precursor protein and its accumulation at the cell surface. Our data suggest that PS is targeted as a biologically active complex with Nct through the secretory pathway to the cell surface and suggest a dual function of PS in γ-secretase processing and in trafficking.

scholarly journals Quality control at the plasma membrane: One mechanism does not fit all

2014 ◽  
Vol 205 (1) ◽  
pp. 11-20 ◽  
Author(s):  
Markus Babst
Keyword(s):  
Quality Control ◽  
Plasma Membrane ◽  
Control System ◽  
Cell Surface ◽  
Transmembrane Proteins ◽  
Surface Proteins ◽  
Quality Control System ◽  
Internal Quality ◽  
Control Mechanisms ◽  
Cell Integrity

The plasma membrane quality control system of eukaryotic cells is able to recognize and degrade damaged cell surface proteins. Recent studies have identified two mechanisms involved in the recognition of unfolded transmembrane proteins. One system uses chaperones to detect unfolded cytoplasmic domains of transmembrane proteins, whereas the second mechanism relies on an internal quality control system of the protein, which can trigger degradation when the protein deviates from the folded state. Both quality control mechanisms are key to prevent proteotoxic effects at the cell surface and to ensure cell integrity.

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