S-Acylation of the cellulose synthase complex is essential for its plasma membrane localization

ABSTRACT Palmitoylation is a reversible, posttranslational modification that helps target proteins to cellular membranes. The alphavirus small membrane proteins 6K and TF have been reported to be palmitoylated and to positively regulate budding. 6K and TF are isoforms that are identical in their N termini but unique in their C termini due to a −1 ribosomal frameshift during translation. In this study, we used cysteine (Cys) mutants to test differential palmitoylation of the Sindbis virus 6K and TF proteins. We modularly mutated the five Cys residues in the identical N termini of 6K and TF, the four additional Cys residues in TF's unique C terminus, or all nine Cys residues in TF. Using these mutants, we determined that TF palmitoylation occurs primarily in the N terminus. In contrast, 6K is not palmitoylated, even on these shared residues. In the C-terminal Cys mutant, TF protein levels increase both in the cell and in the released virion compared to the wild type. In viruses with the N-terminal Cys residues mutated, TF is much less efficiently localized to the plasma membrane, and it is not incorporated into the virion. The three Cys mutants have minor defects in cell culture growth but a high incidence of abnormal particle morphologies compared to the wild-type virus as determined by transmission electron microscopy. We propose a model where the C terminus of TF modulates the palmitoylation of TF at the N terminus, and palmitoylated TF is preferentially trafficked to the plasma membrane for virus budding. IMPORTANCE Alphaviruses are a reemerging viral cause of arthritogenic disease. Recently, the small 6K and TF proteins of alphaviruses were shown to contribute to virulence in vivo. Nevertheless, a clear understanding of the molecular mechanisms by which either protein acts to promote virus infection is missing. The TF protein is a component of budded virions, and optimal levels of TF correlate positively with wild-type-like particle morphology. In this study, we show that the palmitoylation of TF regulates its localization to the plasma membrane, which is the site of alphavirus budding. Mutants in which TF is not palmitoylated display drastically reduced plasma membrane localization, which effectively prevents TF from participating in budding or being incorporated into virus particles. Investigation of the regulation of TF will aid current efforts in the alphavirus field searching for approaches to mitigate alphaviral disease in humans.

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The secondary structure of the TGR5 membrane-proximal C-terminus determines plasma membrane localization and responsiveness towards extracellular ligands

European Journal of Medical Research ◽

10.1186/2047-783x-19-s1-s14 ◽

2014 ◽

Vol 19 (Suppl 1) ◽

pp. S14

Author(s):

Christoph Gertzen ◽

Lina Spomer ◽

Birte Schmitz ◽

Dieter Häussinger ◽

Verena Keitel ◽

...

Keyword(s):

Plasma Membrane ◽

Secondary Structure ◽

Membrane Localization ◽

Plasma Membrane Localization ◽

C Terminus

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Nicotine Induces Progressive Properties of Lung Adenocarcinoma A549 Cells by Inhibiting Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Expression and Plasma Membrane Localization

Technology in Cancer Research & Treatment ◽

10.1177/1533033818809984 ◽

2018 ◽

Vol 17 ◽

pp. 153303381880998 ◽

Cited By ~ 6

Author(s):

Hui Li ◽

Ningxia Ma ◽

Jing Wang ◽

Ying Wang ◽

Chao Yuan ◽

...

Keyword(s):

Cystic Fibrosis ◽

Plasma Membrane ◽

Lung Adenocarcinoma ◽

A549 Cells ◽

Membrane Localization ◽

Transmembrane Conductance Regulator ◽

Transmembrane Conductance ◽

Plasma Membrane Localization ◽

Lung Adenocarcinoma A549 ◽

Cystic Fibrosis Transmembrane

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A novel RCE1 isoform is required for H-Ras plasma membrane localization and is regulated by USP17

Biochemical Journal ◽

10.1042/bj20131213 ◽

2013 ◽

Vol 457 (2) ◽

pp. 289-300 ◽

Cited By ~ 10

Author(s):

Jakub Jaworski ◽

Ureshnie Govender ◽

Cheryl McFarlane ◽

Michelle de la Vega ◽

Michelle K. Greene ◽

...

Keyword(s):

Plasma Membrane ◽

Membrane Localization ◽

Plasma Membrane Localization

We have identified a novel RCE1 isoform which is required for proper H-Ras processing and plasma membrane localization. In addition, we have shown that USP17 can regulate this novel isoform and thus RCE1 activity by deubiquitinating Lys43.

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Role of Ezrin/Radixin/Moesin in the Surface Localization of Programmed Cell Death Ligand-1 in Human Colon Adenocarcinoma LS180 Cells

Pharmaceuticals ◽

10.3390/ph14090864 ◽

2021 ◽

Vol 14 (9) ◽

pp. 864

Author(s):

Takuro Kobori ◽

Chihiro Tanaka ◽

Mayuka Tameishi ◽

Yoko Urashima ◽

Takuya Ito ◽

...

Keyword(s):

Cell Death ◽

Plasma Membrane ◽

Programmed Cell Death ◽

Cell Surface ◽

Actin Cytoskeleton ◽

Mrna Level ◽

Scaffold Proteins ◽

Membrane Localization ◽

Erm Proteins ◽

Plasma Membrane Localization

Programmed cell death ligand-1 (PD-L1), an immune checkpoint protein highly expressed on the cell surface in various cancer cell types, binds to programmed cell death-1 (PD-1), leading to T-cell dysfunction and tumor survival. Despite clinical successes of PD-1/PD-L1 blockade therapies, patients with colorectal cancer (CRC) receive little benefit because most cases respond poorly. Because high PD-L1 expression is associated with immune evasion and poor prognosis in CRC patients, identifying potential modulators for the plasma membrane localization of PD-L1 may represent a novel therapeutic strategy for enhancing the efficacy of PD-1/PD-L1 blockade therapies. Here, we investigated whether PD-L1 expression in human colorectal adenocarcinoma cells (LS180) is affected by ezrin/radixin/moesin (ERM), functioning as scaffold proteins that crosslink plasma membrane proteins with the actin cytoskeleton. We observed colocalization of PD-L1 with all three ERM proteins in the plasma membrane and detected interactions involving PD-L1, the three ERM proteins, and the actin cytoskeleton. Furthermore, gene silencing of ezrin and radixin, but not of moesin, substantially decreased the expression of PD-L1 on the cell surface without affecting its mRNA level. Thus, in LS180 cells, ezrin and radixin may function as scaffold proteins mediating the plasma membrane localization of PD-L1, possibly by post-translational modification.

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