scholarly journals Targeting CCR5 trafficking to inhibit HIV-1 infection

2019 ◽  
Vol 5 (10) ◽  
pp. eaax0821 ◽  
Author(s):  
Gaelle Boncompain ◽  
Floriane Herit ◽  
Sarah Tessier ◽  
Aurianne Lescure ◽  
Elaine Del Nery ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Molecular Level ◽  
High Content Screening ◽  
Differential Protein ◽  
Early Secretory Pathway ◽  
A Cell ◽  
Hiv Entry ◽  
Hiv 1

Using a cell-based assay monitoring differential protein transport in the secretory pathway coupled to high-content screening, we have identified three molecules that specifically reduce the delivery of the major co-receptor for HIV-1, CCR5, to the plasma membrane. They have no effect on the closely related receptors CCR1 and CXCR4. These molecules are also potent in primary macrophages as they markedly decrease HIV entry. At the molecular level, two of these molecules inhibit the critical palmitoylation of CCR5 and thereby block CCR5 in the early secretory pathway. Our results open a clear therapeutics avenue based on trafficking control and demonstrate that preventing HIV infection can be performed at the level of its receptor delivery.

Mammalian GPI-anchored proteins require p24 proteins for their efficient transport from the ER to the plasma membrane

2007 ◽  
Vol 409 (2) ◽  
pp. 555-562 ◽  
Author(s):  
Satoshi Takida ◽  
Yusuke Maeda ◽  
Taroh Kinoshita
Keyword(s):  
Plasma Membrane ◽  
Membrane Trafficking ◽  
Secretory Pathway ◽  
Direct Evidence ◽  
Temperature Sensitive ◽  
Early Secretory Pathway ◽  
Cargo Proteins ◽  
Transport Vesicle ◽  
A Cell ◽  
P24 Proteins

The GPI (glycosylphosphatidylinositol) moiety is attached to newly synthesized proteins in the lumen of the ER (endoplasmic reticulum). The modified proteins are then directed to the PM (plasma membrane). Less well understood is how nascent mammalian GPI-anchored proteins are targeted from the ER to the PM. In the present study, we investigated mechanisms underlying membrane trafficking of the GPI-anchored proteins, focusing on the early secretory pathway. We first established a cell line that stably expresses inducible temperature-sensitive GPI-fused proteins as a reporter and examined roles of transport-vesicle constituents called p24 proteins in the traffic of the GPI-anchored proteins. We selectively suppressed one of the p24 proteins, namely p23, employing RNAi (RNA interference) techniques. The suppression resulted in pronounced delays of PM expression of the GPI-fused reporter proteins. Furthermore, maturation of DAF (decay-accelerating factor), one of the GPI-anchored proteins in mammals, was slowed by the suppression of p23, indicating delayed trafficking of DAF from the ER to the Golgi. Trafficking of non-GPI-linked cargo proteins was barely affected by p23 knockdown. This is the first to demonstrate direct evidence for the transport of mammalian GPI-anchored proteins being mediated by p24 proteins.

scholarly journals A Cell-Specific Transgenic Approach in Xenopus Reveals the Importance of a Functional p24 System for a Secretory Cell

2004 ◽  
Vol 15 (3) ◽  
pp. 1244-1253 ◽  
Author(s):  
Gerrit Bouw ◽  
Rick Van Huizen ◽  
Eric J.R. Jansen ◽  
Gerard J.M. Martens
Keyword(s):  
Secretory Pathway ◽  
Protein Transport ◽  
Transgenic Animal ◽  
Secretory Protein ◽  
Physiological Effect ◽  
Early Secretory Pathway ◽  
Golgi Transport ◽  
Cargo Transport ◽  
Transport Vesicles ◽  
A Cell

The p24α, -β, -γ, and -δ proteins are major multimeric constituents of cycling endoplasmic reticulum-Golgi transport vesicles and are thought to be involved in protein transport through the early secretory pathway. In this study, we targeted transgene overexpression of p24δ2 specifically to the Xenopus intermediate pituitary melanotrope cell that is involved in background adaptation of the animal and produces high levels of its major secretory cargo proopiomelanocortin (POMC). The transgene product effectively displaced the endogenous p24 proteins, resulting in a melanotrope cell p24 system that consisted predominantly of the transgene p24δ2 protein. Despite the severely distorted p24 machinery, the subcellular structures as well as the level of POMC synthesis were normal in these cells. However, the number and pigment content of skin melanophores were reduced, impairing the ability of the transgenic animal to fully adapt to a black background. This physiological effect was likely caused by the affected profile of POMC-derived peptides observed in the transgenic melanotrope cells. Together, our results suggest that in the early secretory pathway an intact p24 system is essential for efficient secretory cargo transport or for supplying cargo carriers with the correct protein machinery to allow proper secretory protein processing.

scholarly journals A stress assembly that confers cell viability by preserving ERES components during amino-acid starvation

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Margarita Zacharogianni ◽  
Angelica Aguilera-Gomez ◽  
Tineke Veenendaal ◽  
Jan Smout ◽  
Catherine Rabouille
Keyword(s):  
Amino Acid ◽  
Cell Viability ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Liquid Droplet ◽  
Protein Translation ◽  
Amino Acid Starvation ◽  
Early Secretory Pathway ◽  
Er Exit Sites ◽  
Nutritional Restriction

Nutritional restriction leads to protein translation attenuation that results in the storage and degradation of free mRNAs in cytoplasmic assemblies. In this study, we show in Drosophila S2 cells that amino-acid starvation also leads to the inhibition of another major anabolic pathway, the protein transport through the secretory pathway, and to the formation of a novel reversible non-membrane bound stress assembly, the Sec body that incorporates components of the ER exit sites. Sec body formation does not depend on membrane traffic in the early secretory pathway, yet requires both Sec23 and Sec24AB. Sec bodies have liquid droplet-like properties, and they act as a protective reservoir for ERES components to rebuild a functional secretory pathway after re-addition of amino-acids acting as a part of a survival mechanism. Taken together, we propose that the formation of these structures is a novel stress response mechanism to provide cell viability during and after nutrient stress.

scholarly journals Casein Kinase I Regulates Membrane Binding by ARF GAP1

2002 ◽  
Vol 13 (8) ◽  
pp. 2559-2570 ◽  
Author(s):  
Sidney Yu ◽  
Michael G. Roth
Keyword(s):  
Amino Acid ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Membrane Binding ◽  
Casein Kinase ◽  
Protein Domain ◽  
Amino Terminal ◽  
Early Secretory Pathway

ARF GAP1, a 415-amino acid GTPase activating protein (GAP) for ADP-ribosylation factor (ARF) contains an amino-terminal 115-amino acid catalytic domain and no other recognizable features. Amino acids 203–334 of ARF GAP1 were sufficient to target a GFP-fusion protein to Golgi membranes in vivo. When overexpressed in COS-1 cells, this protein domain inhibited protein transport between the ER and Golgi and, in vitro, competed with the full-length ARF GAP1 for binding to membranes. Membrane binding by ARF GAP1 in vitro was increased by a factor in cytosol and this increase was inhibited by IC261, an inhibitor selective for casein kinase Iδ (CKIδ), or when cytosol was treated with antibody to CKIδ. The noncatalytic domain of ARF GAP1 was phosphorylated both in vivo and in vitro by CKI. IC261 blocked membrane binding by ARF GAP1 in vivo and inhibited protein transport in the early secretory pathway. Overexpression of a catalytically inactive CKIδ also inhibited the binding of ARF GAP1 to membranes and interfered with protein transport. Thus, a CKI isoform is required for protein traffic through the early secretory pathway and can modulate the amount of ARF GAP1 that can bind to membranes.

Receptor-mediated protein transport in the early secretory pathway

2007 ◽  
Vol 32 (8) ◽  
pp. 381-388 ◽  
Author(s):  
Andrea C. Baines ◽  
Bin Zhang
Keyword(s):  
Secretory Pathway ◽  
Protein Transport ◽  
Early Secretory Pathway

scholarly journals Sphingomyelin Synthase 2, but Not Sphingomyelin Synthase 1, Is Involved in HIV-1 Envelope-mediated Membrane Fusion

2014 ◽  
Vol 289 (44) ◽  
pp. 30842-30856 ◽  
Author(s):  
Yasuhiro Hayashi ◽  
Yoko Nemoto-Sasaki ◽  
Takashi Tanikawa ◽  
Saori Oka ◽  
Kiyoto Tsuchiya ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Membrane Fusion ◽  
Actin Polymerization ◽  
Plasma Membranes ◽  
Viral Envelope ◽  
Target Cells ◽  
Nonreceptor Tyrosine Kinase ◽  
Sphingomyelin Synthase ◽  
A Cell ◽  
Hiv 1

Membrane fusion between the viral envelope and plasma membranes of target cells has previously been correlated with HIV-1 infection. Lipids in the plasma membrane, including sphingomyelin, may be crucially involved in HIV-1 infection; however, the role of lipid-metabolic enzymes in membrane fusion remains unclear. In this study, we examined the roles of sphingomyelin synthase (SMS) in HIV-1 Env-mediated membrane fusion using a cell-cell fusion assay with HIV-1 mimetics and their target cells. We employed reconstituted cells as target cells that stably express Sms1 or Sms2 in Sms-deficient cells. Fusion susceptibility was ∼5-fold higher in Sms2-expressing cells (not in Sms1-expressing cells) than in Sms-deficient cells. The enhancement of fusion susceptibility observed in Sms2-expressing cells was reversed and reduced by Sms2 knockdown. We also found that catalytically nonactive Sms2 promoted membrane fusion susceptibility. Moreover, SMS2 co-localized and was constitutively associated with the HIV receptor·co-receptor complex in the plasma membrane. In addition, HIV-1 Env treatment resulted in a transient increase in nonreceptor tyrosine kinase (Pyk2) phosphorylation in Sms2-expressing and catalytically nonactive Sms2-expressing cells. We observed that F-actin polymerization in the region of membrane fusion was more prominent in Sms2-expressing cells than Sms-deficient cells. Taken together, our research provides insight into a novel function of SMS2 which is the regulation of HIV-1 Env-mediated membrane fusion via actin rearrangement.

Biosynthetic protein transport through the early secretory pathway

1998 ◽  
Vol 109 (5-6) ◽  
pp. 477-486 ◽  
Author(s):  
W. Nickel ◽  
Felix T. Wieland
Keyword(s):  
Secretory Pathway ◽  
Protein Transport ◽  
Early Secretory Pathway

scholarly journals Chs6p-dependent Anterograde Transport of Chs3p from the Chitosome to the Plasma Membrane inSaccharomyces cerevisiae

1998 ◽  
Vol 9 (6) ◽  
pp. 1565-1576 ◽  
Author(s):  
Michael Ziman ◽  
John S. Chuang ◽  
Michael Tsung ◽  
Susan Hamamoto ◽  
Randy Schekman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Cell Surface ◽  
Cell Separation ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Subcellular Fractionation ◽  
Chitin Synthase ◽  
Anterograde Transport ◽  
Membrane Compartment

Chitin synthase III (CSIII), an enzyme required to form a chitin ring in the nascent division septum of Saccharomyces cerevisiae, may be transported to the cell surface in a regulated manner. Chs3p, the catalytic subunit of CSIII, requires the product of CHS6 to be transported to or activated at the cell surface. We find that chs6Δ strains have morphological abnormalities similar to those of chs3mutants. Subcellular fractionation and indirect immunofluorescence indicate that Chs3p distribution is altered in chs6mutant cells. Order-of-function experiments usingend4–1 (endocytosis-defective) and chs6mutants indicate that Chs6p is required for anterograde transport of Chs3p from an internal endosome-like membrane compartment, the chitosome, to the plasma membrane. As a result, chs6strains accumulate Chs3p in chitosomes. Chs1p, a distinct chitin synthase that acts during or after cell separation, is transported normally in chs6 mutants, suggesting that Chs1p and Chs3p are independently packaged during protein transport through the late secretory pathway.

scholarly journals Visualizing Wnt secretion from Endoplasmic Reticulum to Filopodia

2018 ◽  
Author(s):  
Naushad Moti ◽  
Jia Yu ◽  
Gaelle Boncompain ◽  
Franck Perez ◽  
David M Virshup
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Secretory Pathway ◽  
Signaling Cascades ◽  
Downstream Signaling ◽  
Cell Compartments ◽  
A Cell ◽  
Cortical Regions ◽  
Rate Limiting ◽  
Wnt Signal

AbstractWnts are a family of secreted palmitoleated glycoproteins that play a key role in cell to cell communications during development and regulate stem cell compartments in adults. Wnt receptors, downstream signaling cascades and target pathways have been extensively studied while less is known about how Wnts are secreted and move from producing cells to receiving cells. We used the synchronization system called Retention Using Selective Hook (RUSH) to study Wnt trafficking from endoplasmic reticulum to Golgi and then to plasma membrane and filopodia in real time. Consistent with prior studies, inhibition of porcupine (PORCN) or knockout of Wntless (WLS) blocked Wnt exit from the ER. Indeed, WLS was rate-limiting for Wnt ER exit. Wnt-containing vesicles paused at sub-cortical regions of the plasma membrane before exiting the cell. Wnt-containing vesicles were transported to adjacent cells associated with filopodia. Increasing the number of filopodia by expression of LGR5 in the producing cell increased the ability of a cell to send a Wnt signal. The RUSH system is a powerful tool to provide new insights into the Wnt secretory pathway.

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