Nutrient-Dependency of CendR Endocytosis Pathway is Uniquely Regulated by Mutant Kras and Targetable

ABSTRACT Entry of the budded virus form of baculoviruses into insect and mammalian cells is generally thought to occur through a low-pH-dependent endocytosis pathway, possibly through clathrin-coated pits. This insight is primarily based on (immuno)electron microscopy studies but requires biochemical support to exclude the use of other pathways. Here, we demonstrate using various inhibitors that functional entry of baculoviruses into insect and mammalian cells is primarily dependent on clathrin-mediated endocytosis. Our results further suggest that caveolae are somehow involved in baculovirus entry in mammalian cells. A caveolar endocytosis inhibitor, genistein, enhances baculovirus transduction in these cells considerably.

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Spatial regulation of clathrin-mediated endocytosis through position-dependent site maturation

10.1101/834523 ◽

2019 ◽

Author(s):

Ross TA Pedersen ◽

Julian E Hassinger ◽

Paul Marchando ◽

David G Drubin

Keyword(s):

Saccharomyces Cerevisiae ◽

Transition Point ◽

Quantitative Imaging ◽

Imaging Data ◽

Cortical Actin ◽

Initiation Phase ◽

Spatial Regulation ◽

Polarized Cells ◽

Endocytosis Pathway ◽

Dependent Site

AbstractDuring clathrin-mediated endocytosis (CME), over 50 different proteins assemble on the plasma membrane to reshape it into a cargo-laden vesicle. It has long been assumed that cargo triggers local CME site assembly in Saccharomyces cerevisiae based on the discovery that cortical actin patches clustered near exocytic sites are CME sites. Quantitative imaging data reported here lead to a radically different view of which CME steps are regulated and which steps are deterministic. We quantitatively and spatially describe progression through the CME pathway and pinpoint a cargo-sensitive regulatory transition point that governs progression from the initiation phase of CME to the internalization phase. Thus, site maturation, rather than site initiation, accounts for the previously observed polarized distribution of actin patches in this organism. While previous studies suggested that cargo ensures its own internalization by regulating either CME initiation rates or frequency of abortive events, our data instead identify maturation through a checkpoint in the pathway as the cargo-sensitive step.SummaryPedersen, Hassinger, et al. investigate steps of the clathrin-mediated endocytosis pathway that are subject to regulation. They report position-dependent differences in endocytic site maturation rates in polarized cells and suggest that cargo controls endocytic internalization through tuning site maturation rather than site initiation.

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Orthogonal genome-wide screens of bat cells identify MTHFD1 as a target of broad antiviral therapy

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2104759118 ◽

2021 ◽

Vol 118 (39) ◽

pp. e2104759118

Author(s):

Danielle E. Anderson ◽

Jin Cui ◽

Qian Ye ◽

Baoying Huang ◽

Ya Tan ◽

...

Keyword(s):

Antiviral Therapy ◽

Influenza A ◽

Secretory Pathway ◽

Viral Infections ◽

Innate Immune ◽

Genome Wide ◽

Genomic Studies ◽

Independent Screening ◽

Endocytosis Pathway ◽

Lower Expression

Bats are responsible for the zoonotic transmission of several major viral diseases, including those leading to the 2003 SARS outbreak and likely the ongoing COVID-19 pandemic. While comparative genomics studies have revealed characteristic adaptations of the bat innate immune system, functional genomic studies are urgently needed to provide a foundation for the molecular dissection of the viral tolerance in bats. Here we report the establishment of genome-wide RNA interference (RNAi) and CRISPR libraries for the screening of the model megabat, Pteropus alecto. We used the complementary RNAi and CRISPR libraries to interrogate P. alecto cells for infection with two different viruses: mumps virus and influenza A virus, respectively. Independent screening results converged on the endocytosis pathway and the protein secretory pathway as required for both viral infections. Additionally, we revealed a general dependence of the C1-tetrahydrofolate synthase gene, MTHFD1, for viral replication in bat cells and human cells. The MTHFD1 inhibitor, carolacton, potently blocked replication of several RNA viruses, including SARS-CoV-2. We also discovered that bats have lower expression levels of MTHFD1 than humans. Our studies provide a resource for systematic inquiry into the genetic underpinnings of bat biology and a potential target for developing broad-spectrum antiviral therapy.

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A myosin-7B–dependent endocytosis pathway mediates cellular entry of α-synuclein fibrils and polycation-bearing cargos

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1918617117 ◽

2020 ◽

Vol 117 (20) ◽

pp. 10865-10875 ◽

Cited By ~ 5

Author(s):

Qi Zhang ◽

Yue Xu ◽

Juhyung Lee ◽

Michal Jarnik ◽

Xufeng Wu ◽

...

Keyword(s):

Cell Surface ◽

Actin Filaments ◽

Membrane Dynamics ◽

Cell Entry ◽

Sulfate Proteoglycan ◽

Actin Network ◽

Coated Pits ◽

Endocytosis Pathway ◽

Underlying Mechanisms ◽

Pathological Event

Cell-to-cell transmission of misfolding-prone α-synuclein (α-Syn) has emerged as a key pathological event in Parkinson’s disease. This process is initiated when α-Syn–bearing fibrils enter cells via clathrin-mediated endocytosis, but the underlying mechanisms are unclear. Using a CRISPR-mediated knockout screen, we identify SLC35B2 and myosin-7B (MYO7B) as critical endocytosis regulators for α-Syn preformed fibrils (PFFs). We show that SLC35B2, as a key regulator of heparan sulfate proteoglycan (HSPG) biosynthesis, is essential for recruiting α-Syn PFFs to the cell surface because this process is mediated by interactions between negatively charged sugar moieties of HSPGs and clustered K-T-K motifs in α-Syn PFFs. By contrast, MYO7B regulates α-Syn PFF cell entry by maintaining a plasma membrane-associated actin network that controls membrane dynamics. Without MYO7B or actin filaments, many clathrin-coated pits fail to be severed from the membrane, causing accumulation of large clathrin-containing “scars” on the cell surface. Intriguingly, the requirement for MYO7B in endocytosis is restricted to α-Syn PFFs and other polycation-bearing cargos that enter cells via HSPGs. Thus, our study not only defines regulatory factors for α-Syn PFF endocytosis, but also reveals a previously unknown endocytosis mechanism for HSPG-binding cargos in general, which requires forces generated by MYO7B and actin filaments.

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Efficient Therapeutic Delivery by a Novel Cell-Penetrating Peptide Derived from Acinus

Cancers ◽

10.3390/cancers12071858 ◽

2020 ◽

Vol 12 (7) ◽

pp. 1858

Author(s):

Justine Habault ◽

Claire Fraser ◽

Ewa Pasquereau-Kotula ◽

Maëlys Born-Bony ◽

Anne Marie-Cardine ◽

...

Keyword(s):

Ex Vivo ◽

Clinical Settings ◽

Aspartic Acid Residue ◽

Circulating Cells ◽

C Terminus ◽

Cell Penetrating ◽

Endocytosis Pathway ◽

Cancerous Cells

In this study, we have identified a novel cell-penetrating sequence, termed hAP10, from the C-terminus of the human protein Acinus. hAP10 was able to efficiently enter various normal and cancerous cells, likely through an endocytosis pathway, and to deliver an EGFP cargo to the cell interior. Cell penetration of a peptide, hAP10DR, derived from hAP10 by mutation of an aspartic acid residue to an arginine was dramatically increased. Interestingly, a peptide containing a portion of the heptad leucine repeat region domain of the survival protein AAC-11 (residues 377–399) fused to either hAP10 or hAP10DR was able to induce tumor cells, but not normal cells, death both ex vivo on Sézary patients’ circulating cells and to inhibit tumor growth in vivo in a sub-cutaneous xenograft mouse model for the Sézary syndrome. Combined, our results indicate that hAP10 and hAP10DR may represent promising vehicles for the in vitro or in vivo delivery of bioactive cargos, with potential use in clinical settings.

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Peste des Petits Ruminants Virus Enters Caprine Endometrial Epithelial Cells via the Caveolae-Mediated Endocytosis Pathway

Frontiers in Microbiology ◽

10.3389/fmicb.2018.00210 ◽

2018 ◽

Vol 9 ◽

Cited By ~ 6

Author(s):

Bo Yang ◽

Xuefeng Qi ◽

Hui Guo ◽

Peilong Jia ◽

Shuying Chen ◽

...

Keyword(s):

Epithelial Cells ◽

Peste Des Petits Ruminants ◽

Endocytosis Pathway ◽

Endometrial Epithelial Cells

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Cell-to-Cell Spread of Borna Disease Virus Proceeds in the Absence of the Virus Primary Receptor and Furin-Mediated Processing of the Virus Surface Glycoprotein

Journal of Virology ◽

10.1128/jvi.02426-06 ◽

2007 ◽

Vol 81 (11) ◽

pp. 5968-5977 ◽

Cited By ~ 23

Author(s):

Roberto Clemente ◽

Juan C. de la Torre

Keyword(s):

Disease Virus ◽

Primary Infection ◽

Borna Disease Virus ◽

Virus Surface ◽

Receptor Mediated Endocytosis ◽

Extracellular Virus ◽

Primary Receptor ◽

Endocytosis Pathway ◽

Cell Spread ◽

Borna Disease

ABSTRACT Borna disease virus (BDV) is an enveloped virus with a nonsegmented negative-strand RNA genome whose organization is characteristic of Mononegavirales. BDV cell entry follows a receptor-mediated endocytosis pathway, which is initiated by the recognition of an as-yet-unidentified receptor at the cell surface by the virus glycoprotein G. BDV G is synthesized as a precursor (GPC) that is cleaved by the cellular protease furin to produce the mature glycoproteins GP1 and GP2, which have been implicated in receptor recognition and pH-dependent fusion events, respectively. BDV is highly neurotropic and its spread in cultured cells proceeds in the absence of detectable extracellular virus or syncytium formation. BDV spread has been proposed to be strictly dependent on the expression and correct processing of BDV G. Here we present evidence that cell-to-cell spread of BDV required neither the expression of cellular receptors involved in virus primary infection, nor the furin-mediated processing of BDV G. We also show that in furin-deficient cells, the release of BDV particles induced by the treatment of BDV-infected cells with hypertonic buffer was not significantly affected, while virion infectivity was dramatically impaired, correlating with the decreased incorporation of BDV G species into viral particles. These findings support the view that the propagation of BDV within the central nervous systems of infected hosts involves both a primary infection that follows a receptor-mediated endocytosis pathway and a subsequent cell-to-cell spread that is independent of the expression of the primary receptor and does not require the processing of BDV G into GP1 and GP2.

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