Plasma Membrane
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2021 ◽  
Mohammad Haroon Qureshi ◽  
M. Talha Cinko ◽  
Halil Bayraktar ◽  
Cansu Akkaya ◽  
Altug Kamacioglu ◽  

Cell migration requires spatiotemporally coordinated activities of multicomponent structures including the actomyosin cortex, plasma membrane, adhesion complexes and the polarity proteins. How they function together to drive this complex dynamic process remains an outstanding question. Here, we show that a member of the protocadherin family, PCDH7 displays a polarized localization in migratory cells with a dynamic enrichment at the leading and rear edges. Perturbation of PCDH7 interferes with the migration of nontransformed retinal pigment epithelial cells and invasion of cancer cells. The overexpression of PCDH7 enhances the migration capability of cortical neurons in vivo. PCDH7 interacts with the myosin phosphatase subunits MYPT1 and PP1cβ and it enhances the phosphorylation of regulatory light chain and ERM at the leading and rear edges of migratory cells. The chemical inhibition of phosphatase activity recovers migration phenotypes of PCDH7 knockout cells. We propose that PCDH7 regulate phosphorylation thus activity of myosin and ERM at the polarized cortex by quenching myosin phosphatase that results in a higher persistence of migrating cells. Collectively, our study suggests a new mechanism for the spatial coordination of plasma membrane and the cortex during cell migration.

Biomolecules ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 1393
Mauro Marra ◽  
Lorenzo Camoni ◽  
Sabina Visconti ◽  
Anna Fiorillo ◽  
Antonio Evidente

Fusicoccin is the α glucoside of a carbotricyclic diterpene, produced by the fungus Phomopsis amygdali (previously classified as Fusicoccum amygdali), the causal agent of almond and peach canker disease. A great interest in this molecule started when it was discovered that it brought about an irreversible stomata opening of higher plants, thereby inducing the wilting of their leaves. Since then, several studies were carried out to elucidate its biological activity, biosynthesis, structure, structure-activity relationships and mode of action. After sixty years of research and more than 1800 published articles, FC is still the most studied phytotoxin and one of the few whose mechanism of action has been elucidated in detail. The ability of FC to stimulate several fundamental plant processes depends on its ability to activate the plasma membrane H+-ATPase, induced by eliciting the association of 14-3-3 proteins, a class of regulatory molecules widespread in eukaryotes. This discovery renewed interest in FC and prompted more recent studies aimed to ascertain the ability of the toxin to influence the interaction between 14-3-3 proteins and their numerous client proteins in animals, involved in the regulation of basic cellular processes and in the etiology of different diseases, including cancer. This review covers the different aspects of FC research partially treated in different previous reviews, starting from its discovery in 1964, with the aim to outline the extraordinary pathway which led this very uncommon diterpenoid to evolve from a phytotoxin into a tool in plant physiology and eventually into a 14-3-3-targeted drug.

Chad A. Sallaberry ◽  
Barbie J. Voss ◽  
Jaroslaw Majewski ◽  
Jacek Biernat ◽  
Eckhard Mandelkow ◽  

Tau misfolding and assembly is linked to a number of neurodegenerative diseases collectively described as tauopathies, including Alzheimer’s disease (AD) and Parkinson’s disease. Anionic cellular membranes, such as the cytosolic leaflet of the plasma membrane, are sites that concentrate and neutralize tau, primarily due to electrostatic interactions with tau’s microtubule binding repeat domain (RD). In addition to electrostatic interactions with lipids, tau also has interactions with membrane proteins, which are important for tau’s cellular functions. Tau also interacts with lipid tails to facilitate direct translocation across the membrane and can form stable protein-lipid complexes involved in cell-to-cell transport. Concentrated tau monomers at the membrane surface can form reversible condensates, change secondary structures, and induce oligomers, which may eventually undergo irreversible crosslinking and fibril formation. These β-sheet rich tau structures are capable of disrupting membrane organization and are toxic in cell-based assays. Given the evidence for relevant membrane-based tau assembly, we review the emerging hypothesis that polyanionic membranes may serve as a site for phase-separated tau condensation. Membrane-mediated phase separation may have important implications for regulating tau folding/misfolding, and may be a powerful mechanism to spatially direct tau for native membrane-mediated functions.

Yushi Uchida ◽  
Kumiko Torisu ◽  
Kenji Ueki ◽  
Kazuhiko Tsuruya ◽  
Toshiaki Nakano ◽  

Receptor-mediated albumin transport in proximal tubule epithelial cells (PTECs) is important to control proteinuria. Autophagy is an evolutionarily conserved degradation pathway and its role in intracellular trafficking through interaction with the endocytic pathway has recently been highlighted. Here, we determined whether autophagy regulates albumin transcytosis in PTECs and suppresses albumin-induced cytotoxicity using human proximal tubule (HK-2) cells. Neonatal Fc-receptor (FcRn), a receptor for albumin transcytosis, partially co-localized with autophagosomes. Recycling of FcRn was attenuated and FcRn accumulated in autophagy related 7 (ATG7) knockdown HK-2 cells. Co-localization of FcRn with RAB7-positive late endosomes and RAB11-positive recycling endosomes was reduced in ATG7 knockdown cells, resulting in decreased recycling of FcRn to the plasma membrane. In ATG7 knockdown cells, albumin transcytosis was significantly reduced and intracellular albumin accumulation was increased. Finally, release of KIM-1, a marker of tubule injury, from ATG7 knockdown cells was increased in response to excess albumin. In conclusion, suppression of autophagy in tubules impairs FcRn transport, thereby inhibiting albumin transcytosis. The resulting accumulation of albumin induces cytotoxicity in tubules.

eLife ◽  
2021 ◽  
Vol 10 ◽  
Jason D Vevea ◽  
Grant F Kusick ◽  
Kevin C Courtney ◽  
Erin Chen ◽  
Shigeki Watanabe ◽  

Synaptotagmin 7 (SYT7) has emerged as a key regulator of presynaptic function, but its localization and precise role in the synaptic vesicle cycle remain the subject of debate. Here, we used iGluSnFR to optically interrogate glutamate release, at the single-bouton level, in SYT7KO-dissociated mouse hippocampal neurons. We analyzed asynchronous release, paired-pulse facilitation, and synaptic vesicle replenishment and found that SYT7 contributes to each of these processes to different degrees. ‘Zap-and-freeze’ electron microscopy revealed that a loss of SYT7 diminishes docking of synaptic vesicles after a stimulus and inhibits the recovery of depleted synaptic vesicles after a stimulus train. SYT7 supports these functions from the axonal plasma membrane, where its localization and stability require both γ-secretase-mediated cleavage and palmitoylation. In summary, SYT7 is a peripheral membrane protein that controls multiple modes of synaptic vesicle (SV) exocytosis and plasticity, in part, through enhancing activity-dependent docking of SVs.

Roos Houtsma ◽  
Nisha K. van der Meer ◽  
Kees Meijer ◽  
Linde Morsink ◽  
Shanna M. Hogeling ◽  

Acute myeloid leukemia (AML) often presents as an oligoclonal disease whereby multiple genetically distinct subclones can co-exist within patients. Differences in signaling and drug sensitivity of such subclones complicates treatment and warrants tools to identify them and track disease progression. We previously identified over 50 AML-specific plasma membrane (PM) proteins and seven of these (CD82, CD97, FLT3, IL1RAP, TIM3, CD25 and CD123) were implemented in routine diagnostics in patients with AML (n=256) and MDS (n=33). We developed a pipeline termed CombiFlow in which expression data of multiple PM markers is merged, allowing a Principle Component-based analyses to identify distinctive marker expression profiles and to generate single cell tSNE landscapes to longitudinally track clonal evolution. Positivity for one or more of the markers after 2 courses of intensive chemotherapy predicted a shorter relapse-free survival supporting a role of these markers in measurable residual disease (MRD) detection. CombiFlow also allowed the tracking of clonal evolution in paired diagnosis and relapse samples (n=12). Extending the panel to 36 AML-specific markers further refined the CombiFlow pipeline. In conclusion, CombiFlow provides a valuable tool in the diagnosis, MRD detection, clonal tracking, and the understanding of clonal heterogeneity in AML.

2021 ◽  
Vol 22 (18) ◽  
pp. 10120
Alessia Gloria ◽  
Alberto Contri ◽  
Elena Mele ◽  
Silvia Fasano ◽  
Riccardo Pierantoni ◽  

Alongside the well-known central modulatory role, the Kisspeptin system, comprising Kiss1, its cleavage products (Kisspeptins), and Kisspeptin receptor (Kiss1R), was found to regulate gonadal functions in vertebrates; however, its functional role in the male gamete and its localization during maturation have been poorly understood. The present study analyzed Kisspeptin system in dog testis and spermatozoa recovered from different segments of the epididymis, with focus on Kiss1R on sperm surface alongside the maturation during epididymal transit, demonstrated by modification in sperm kinetic, morphology, and protamination. The proteins Kiss1 and Kiss1R were detected in dog testis. The receptor Kiss1R only was detected in total protein extracts from epididymis spermatozoa, whereas dot blot revealed Kiss1 immunoreactivity in the epidydimal fluid. An increase of the Kiss1R protein on sperm surface along the length of the epididymis, with spermatozoa in the tail showing plasma membrane integrity and Kiss1R protein (p < 0.05 vs. epididymis head and body) was observed by flow cytometry and further confirmed by epifluorescence microscopy and Western blot carried on sperm membrane preparations. In parallel, during the transit in the epididymis spermatozoa significantly modified their ability to move and the pattern of motility; a progressive increase in protaminization also occurred. In conclusion, Kisspeptin system was detected in dog testis and spermatozoa. Kiss1R trafficking toward plasma membrane along the length of the epididymis and Kiss1 in epididymal fluid suggested a new functional role of the Kisspeptin system in sperm maturation and storage.

Membranes ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 715
Agnieszka Chytła ◽  
Weronika Gajdzik-Nowak ◽  
Agnieszka Biernatowska ◽  
Aleksander F. Sikorski ◽  
Aleksander Czogalla

Our recent studies have pointed to an important role of the MAGUK family member, MPP1, as a crucial molecule interacting with flotillins and involved in the lateral organization of the erythroid plasma membrane. The palmitoylation of MPP1 seems to be an important element in this process; however, studies on the direct effect of palmitoylation on protein–protein or protein–membrane interactions in vitro are still challenging due to the difficulties in obtaining functional post-translationally modified recombinant proteins and the lack of comprehensive protocols for the purification of palmitoylated proteins. In this work, we present an optimized approach for the high-yield overexpression and purification of palmitoylated recombinant MPP1 protein in mammalian HEK-293F cells. The presented approach facilitates further studies on the molecular mechanism of lateral membrane organization and the functional impact of the palmitoylation of MPP1, which could also be carried out for other palmitoylated proteins.

2021 ◽  
Anudari Letian ◽  
Eyoel Yemanaberhan ◽  
Paola Cavaliere ◽  
Noah Dephoure ◽  
Nasser K Altorki ◽  

PD-L1 is a ligand for immune checkpoint receptor PD1. Anti-PD-L1 antibody is an effective therapy for a variety of solid tumors, although a durable response is only achieved in a subset of patients. For unknown reasons, EGFR-mutant tumors respond poorly to checkpoint blockade. Applying quantitative cell biological methods to study PD-L1 biology in lung cancer cells, we establish that growth factors acutely regulate PD-L1 trafficking between the plasma membrane and the interior of cells. Changes in plasma membrane PD-L1 levels will impact PD1 engagement on T cells, thereby influencing PD-L1's immune suppressive activity. To discover potential cell-intrinsic functions of PD-L1, we used APEX2 biotinylation to generate a high-resolution map of the PD-L1 proximal proteome. ESCRT pathway proteins were enriched in PD-L1's proximal proteome, and two ESCRT-dependent functions, turnover of mutant EGFR and biogenesis of extracellular vesicles, were affected by anti-PD-L1 treatment, suggesting a link between PD-L1 and ESCRT function. Proteins that control cytoskeletal dynamics were also enriched in the PD-L1 proteome, and anti-PD-L1 treatment reduced cell migration, identifying migration as a PD-L1 associated function. PD-L1 knockout mimics the effects of the antibody treatment, suggesting anti-PD-L1 antibody effects are loss of function(s). The effects of anti-PD-L1 on the ESCRT-dependent functions and cell migration were restricted to cells harboring oncogenic EGFR mutations. Wildtype and KRAS mutant cells lines were unaffected. Our study reveals new cell-intrinsic roles for PD-L1 in EGFR mutant cells, activities that might contribute to the resistance of EGFR mutant tumors to PD-L1 checkpoint blockade.

2021 ◽  
Vol 7 (38) ◽  
Veneta Gerganova ◽  
Iker Lamas ◽  
David M. Rutkowski ◽  
Aleksandar Vještica ◽  
Daniela Gallo Castro ◽  

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