Cytoplasmic targeting signals mediate delivery of phospholemman to the plasma membrane

2006 ◽  
Vol 290 (5) ◽  
pp. C1275-C1286 ◽  
Author(s):  
Kristan L. Lansbery ◽  
Lauren C. Burcea ◽  
Margaretta L. Mendenhall ◽  
Robert W. Mercer
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Mdck Cells ◽  
Protein Family ◽  
Site Directed Mutagenesis ◽  
Membrane Localization ◽  
Cardiac Sarcolemma ◽  
Plasma Membrane Localization ◽  
The Family ◽  
Corticosteroid Hormone

The FXYD protein family consists of several small, single-span membrane proteins that exhibit a high degree of homology. The best-known members of the family include the γ-subunit of the Na+-K+-ATPase and phospholemman (PLM), a phosphoprotein of cardiac sarcolemma. Other members of the family include corticosteroid hormone-induced factor (CHIF), mammary tumor protein of 8 kDa (Mat-8), and related to ion channels (RIC). The exact physiological roles of the FXYD proteins remain unknown. To better characterize the function of the members of the FXYD protein family, we expressed several members of the family in Madin-Darby canine kidney (MDCK) cells. All of the FXYD proteins, with the exception of PLM, were primarily found in the basolateral plasma membrane. Surprisingly, PLM, a previously characterized plasma membrane protein, was found to colocalize with the endoplasmic reticulum marker protein disulfide isomerase. Treatment of MDCK cells expressing PLM with an agonist of PKC caused some of the PLM to be redistributed to the plasma membrane. Site-directed mutagenesis of residues within the cytoplasmic domain of PLM indicated that a negative charge at Ser69 is necessary to shift the localization of PLM to the plasma membrane. In addition, other regions of PLM necessary for either its endoplasmic reticulum or plasma membrane localization have been elucidated. In contrast to PLM, the plasma membrane localization of CHIF and RIC was not altered by mutation of potential cytoplasmic phosphorylation sites. Overall, these results suggest that phosphorylation of specific residues of PLM may direct PLM from an intracellular compartment to the plasma membrane.

scholarly journals A Dibasic Motif in the Tail of a Class XIV Apicomplexan Myosin Is an Essential Determinant of Plasma Membrane Localization

2000 ◽  
Vol 11 (4) ◽  
pp. 1385-1400 ◽  
Author(s):  
Christine Hettmann ◽  
Angelika Herm ◽  
Ariane Geiter ◽  
Bernd Frank ◽  
Eva Schwarz ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Actin Cytoskeleton ◽  
Fluorescent Protein ◽  
Gliding Motility ◽  
Site Directed Mutagenesis ◽  
Membrane Localization ◽  
Membrane Association ◽  
Dependent Manner ◽  
Plasma Membrane Localization ◽  
Arginine Residues

Obligate intracellular parasites of the phylum Apicomplexa exhibit gliding motility, a unique form of substrate-dependent locomotion essential for host cell invasion and shown to involve the parasite actin cytoskeleton and myosin motor(s). Toxoplasma gondii has been shown to express three class XIV myosins, TgM-A, -B, and -C. We identified an additional such myosin, TgM-D, and completed the sequences of a related Plasmodium falciparum myosin, PfM-A. Despite divergent structural features, TgM-A purified from parasites bound actin in an ATP-dependent manner. Isoform-specific antibodies revealed that TgM-A and recombinant mycTgM-A were localized right beneath the plasma membrane, and subcellular fractionation indicated a tight membrane association. Recombinant TgM-D also had a peripheral although not as sharply defined localization. Truncation of their respective tail domains abolished peripheral localization and tight membrane association. Conversely, fusion of the tails to green fluorescent protein (GFP) was sufficient to confer plasma membrane localization and sedimentability. The peripheral localization of TgM-A and of the GFP-tail fusion did not depend on an intact F-actin cytoskeleton, and the GFP chimera did not localize to the plasma membrane of HeLa cells. Finally, we showed that the specific localization determinants were in the very C terminus of the TgM-A tail, and site-directed mutagenesis revealed two essential arginine residues. We discuss the evidence for a proteinaceous plasma membrane receptor and the implications for the invasion process.

scholarly journals Erf4p and Erf2p Form an Endoplasmic Reticulum-associated Complex Involved in the Plasma Membrane Localization of Yeast Ras Proteins

2002 ◽  
Vol 277 (51) ◽  
pp. 49352-49359 ◽  
Author(s):  
Lihong Zhao ◽  
Sandra Lobo ◽  
Xiangwen Dong ◽  
Addison D. Ault ◽  
Robert J. Deschenes
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Membrane Protein ◽  
Secretory Pathway ◽  
Integral Membrane Protein ◽  
Genetic Screen ◽  
Membrane Localization ◽  
Ras Proteins ◽  
Transport Pathway ◽  
Plasma Membrane Localization

Ras oncogene proteins are plasma membrane-associated signal transducers that are found in all eukaryotes. Posttranslational addition of lipid to a carboxyl-terminal CaaXbox (where “C” represents a cysteine, “a” is generally an aliphatic residue, andXcan be any amino acid) is required to target Ras proteins to the cytosolic surface of the plasma membrane. The pathway by which Ras translocates from the endoplasmic reticulum to the plasma membrane is currently not clear. We have performed a genetic screen to identify components of the Ras plasma membrane localization pathway. Mutations in two genes,ERF2andERF4/SHR5, have been shown to affect the palmitoylation and subcellular localization of Ras proteins. In this report, we show that Erf4p is localized on the endoplasmic reticulum as a peripheral membrane protein in a complex with Erf2p, an integral membrane protein that was identified from the same genetic screen. Erf2p has been shown to be required for the plasma membrane localization of GFP-Ras2p via a pathway distinct from the classical secretory pathway (X. Dong and R. J. Deschenes, manuscript in preparation). We show here that Erf4p, like Erf2p, is involved in the plasma membrane localization of Ras2p. Erf2p and Erf4p represent components of a previously uncharacterized subcellular transport pathway involved in the plasma membrane targeting of Ras proteins.

scholarly journals Lipid binding by the N-terminal motif mediates plasma membrane localization of Bordetella effector protein BteA

2021 ◽  
pp. 100607
Author(s):  
Ivana Malcova ◽  
Ladislav Bumba ◽  
Filip Uljanic ◽  
Darya Kuzmenko ◽  
Jana Nedomova ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Lipid Binding ◽  
Membrane Localization ◽  
Effector Protein ◽  
Plasma Membrane Localization

scholarly journals Palmitoylation of Sindbis Virus TF Protein Regulates Its Plasma Membrane Localization and Subsequent Incorporation into Virions

2016 ◽  
Vol 91 (3) ◽  
Author(s):  
Jolene Ramsey ◽  
Emily C. Renzi ◽  
Randy J. Arnold ◽  
Jonathan C. Trinidad ◽  
Suchetana Mukhopadhyay
Keyword(s):  
Plasma Membrane ◽  
Sindbis Virus ◽  
Molecular Mechanisms ◽  
Wild Type Virus ◽  
Membrane Localization ◽  
Clear Understanding ◽  
Wild Type ◽  
Plasma Membrane Localization ◽  
C Terminus ◽  
N Terminus

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.

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

2013 ◽  
Vol 457 (2) ◽  
pp. 289-300 ◽  
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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