Oregano Oil and Its Principal Component, Carvacrol, Inhibit HIV-1 Fusion into Target Cells

ABSTRACT Oregano essential oil has long been known for its health-promoting benefits. Here, we report its activity against viral replication. Oregano oil was found to specifically inhibit lentiviruses, such as human and simian immunodeficiency viruses (HIV and SIV), irrespective of virus tropism, but not hepatitis C virus, adenovirus 5 (ADV5), Zika virus, and influenza (H1N1) virus. Oregano oil’s most abundant components, carvacrol and its isomer, thymol, were shown to block virus-target cell fusion while not perturbing other stages of the virus life cycle. We detected changes in virus particle density, suggesting that cholesterol depletion from the HIV-1 envelope membrane reduces virus entry. Furthermore, infection was rescued by adding exogenous cholesterol. The evolution of viral resistance to carvacrol supported this mechanism of action with the identification of mutations in the viral gp41 fusion protein that counteracted cholesterol depletion. In addition, resistance to carvacrol emerged later than typically observed for other clinically used drugs, strengthening its antiviral potential. Structure-activity relationship studies revealed key motifs of carvacrol and thymol required for HIV neutralization and identified previously unknown active analogs. Carvacrol was also shown to additively cooperate with antiretroviral therapy. In sum, oregano oil and improved carvacrol and thymol analogs could be considered to supplement current HIV therapeutics. IMPORTANCE Oregano essential oil has multiple benefits in traditional medicine, cosmetics, and food industries. Carvacrol and its analog, thymol, are well-described components of oregano oil. Here, we show that these compounds inhibit HIV-target cell fusion independently of viral tropism. Our results suggest that carvacrol and thymol alter the cholesterol content of the viral membrane, blocking HIV-1 entry into the target cell. Resistance to carvacrol has selected for viruses with mutations in the viral envelope glycoprotein, gp41. This protein is known for its interaction with cholesterol present in membrane lipid rafts. Together, these results demonstrate the potential of therapies targeting the viral envelope membrane, and oregano oil is a safe supplement to antiretrovirals, potentially delaying disease progression and resistance development.

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Functional Mapping of Regions Involved in the Negative Imprinting of Virion Particle Infectivity and in Target Cell Protection by Interferon-Induced Transmembrane Protein 3 against HIV-1

Journal of Virology ◽

10.1128/jvi.01716-18 ◽

2018 ◽

Vol 93 (2) ◽

Cited By ~ 6

Author(s):

Romain Appourchaux ◽

Mathilde Delpeuch ◽

Li Zhong ◽

Julien Burlaud-Gaillard ◽

Kevin Tartour ◽

...

Keyword(s):

Target Cell ◽

Transmembrane Protein ◽

Regulatory Role ◽

Target Cells ◽

Restriction Factors ◽

Cell Protection ◽

Differential Behavior ◽

Antiviral Activities ◽

Divergent Behavior ◽

Hiv 1

ABSTRACT The interferon-induced transmembrane proteins (IFITMs) are a family of highly related antiviral factors that affect numerous viruses at two steps: in target cells by sequestering incoming viruses in endosomes and in producing cells by leading to the production of virions that package IFITMs and exhibit decreased infectivity. While most studies have focused on the former, little is known about the regulation of the negative imprinting of virion particle infectivity by IFITMs and about its relationship with target cell protection. Using a panel of IFITM3 mutants against HIV-1, we have explored these issues as well as others related to the biology of IFITM3, in particular virion packaging, stability, the relation to CD63/multivesicular bodies (MVBs), the modulation of cholesterol levels, and the relationship between negative imprinting of virions and target cell protection. The results that we have obtained exclude a role for cholesterol and indicate that CD63 accumulation does not directly relate to an antiviral behavior. We have defined regions that modulate the two antiviral properties of IFITM3 as well as novel domains that modulate protein stability and that, in so doing, influence the extent of its packaging into virions. The results that we have obtained, however, indicate that, even in the context of an IFITM-susceptible virus, IFITM3 packaging is not sufficient for negative imprinting. Finally, while most mutations concomitantly affect target cell protection and negative imprinting, a region in the C-terminal domain (CTD) exhibits a differential behavior, potentially highlighting the regulatory role that this domain may play in the two antiviral activities of IFITM3. IMPORTANCE IFITM proteins have been associated with the sequestration of incoming virions in endosomes (target cell protection) and with the production of virion particles that incorporate IFITMs and exhibit decreased infectivity (negative imprinting of virion infectivity). How the latter is regulated and whether these two antiviral properties are related remain unknown. By examining the behavior of a large panel of IFITM3 mutants against HIV-1, we determined that IFITM3 mutants are essentially packaged into virions proportionally to their intracellular levels of expression. However, even in the context of an IFITM-susceptible virus, IFITM3 packaging is not sufficient for the antiviral effects. Most mutations were found to concomitantly affect both antiviral properties of IFITM3, but one CTD mutant exhibited a divergent behavior, possibly highlighting a novel regulatory role for this domain. These findings thus advance our comprehension of how this class of broad antiviral restriction factors acts.

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Effect of Gamma Radiation and Oregano Essential Oil on Murein and ATP Concentration of Escherichia coli O157:H7

Journal of Food Protection ◽

10.4315/0362-028x-68.12.2571 ◽

2005 ◽

Vol 68 (12) ◽

pp. 2571-2579 ◽

Cited By ~ 24

Author(s):

STÉPHANE CAILLET ◽

FRANÇOIS SHARECK ◽

MONIQUE LACROIX

Keyword(s):

Escherichia Coli ◽

Essential Oil ◽

Gamma Radiation ◽

Cell Damage ◽

Wall Structure ◽

High Dose ◽

Escherichia Coli O157 ◽

Oregano Essential Oil ◽

Intracellular Atp ◽

Oregano Oil

This study was carried out to evaluate the effects of gamma radiation alone or in combination with oregano essential oil on the murein composition of Escherichia coli O157:H7 and on the intracellular and extracellular concentrations of ATP. The bacterial strain was treated with three radiation doses: 0.4 kGy to induce cell damage, 1.1 kGy to obtain a viable but nonculturable state, and 1.3 kGy to cause cell death. Oregano essential oil was used at 0.006 and 0.025% (wt/vol), which is the MIC. All treatments had a significant effect (P ≤ 0.05) on the murein composition, although some muropeptides did not seem to be affected by the treatment. Each treatment had a different effect on the relative percentage and number of muropeptides. There was a significant correlation (P ≤ 0.05) between the decrease in intracellular ATP and the increase in extracellular ATP following treatment of the cells with oregano oil. The reduction of intracellular ATP was even more important when oregano oil was combined with irradiation, but irradiation alone at a high dose (≤1.1 kGy) significantly decreased (P ≤ 0.05) the internal ATP without affecting the external ATP. Transmission electron microscopic examination revealed that oregano oil and irradiation have an effect on cell wall structure.

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Effect of Gamma Radiation and Oregano Essential Oil on Murein and ATP Concentration of Listeria monocytogenes

Journal of Food Protection ◽

10.4315/0362-028x-69.12.2961 ◽

2006 ◽

Vol 69 (12) ◽

pp. 2961-2969 ◽

Cited By ~ 19

Author(s):

STÉPHANE CAILLET ◽

MONIQUE LACROIX

Keyword(s):

Listeria Monocytogenes ◽

Essential Oil ◽

Gamma Radiation ◽

Cell Damage ◽

Electron Microscopic Observation ◽

Wall Structure ◽

Electron Microscopic ◽

Oregano Essential Oil ◽

Transmission Electron ◽

Oregano Oil

The effects of gamma radiation and of oregano essential oil alone or in combination with radiation on murein composition of Listeria monocytogenes and on the intracellular and extracellular concentration of ATP were evaluated. The bacterial strain was treated with two radiation doses, 1.2 kGy to induce cell damage and 3.5 kGy to cause cell death. Oregano essential oil was used at 0.020 and 0.025% (wt/vol), which is the MIC. All treatments had a significant effect (P ≤ 0.05) on the murein composition, although some muropeptides did not seem to be affected by the treatment. Each treatment influenced differently the relative percentage and number of muropeptides. There was a significant correlation (P ≤ 0.05) between the reduction of intracellular ATP and increase in extracellular ATP, following treatment of the cells with oregano oil. The reduction of intra-cellular ATP was even more important when essential oil was combined with irradiation, but irradiation of L. monocytogenes alone induced a significant decrease (P ≤ 0.05) of the internal ATP without affecting the external ATP. Transmission electron microscopic observation revealed that oregano oil and irradiation have an effect on cell wall structure.

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Cholesterol Dependence of Varicella-Zoster Virion Entry into Target Cells

Journal of Virology ◽

10.1128/jvi.00486-07 ◽

2007 ◽

Vol 81 (14) ◽

pp. 7548-7558 ◽

Cited By ~ 45

Author(s):

S. Hambleton ◽

S. P. Steinberg ◽

M. D. Gershon ◽

A. A. Gershon

Keyword(s):

Lipid Rafts ◽

Viral Entry ◽

Membrane Lipid ◽

Viral Envelope ◽

Target Cells ◽

Cholesterol Depletion ◽

Dependent Manner ◽

Membrane Composition ◽

Focus Formation ◽

Varicella Zoster

ABSTRACT The entry of inhaled virions into airway cells is presumably the initiating step of varicella-zoster infection. In order to characterize viral entry, we studied the relative roles played by lipid rafts and clathrin-mediated transport. Virus and target cells were pretreated with agents designed to perturb selected aspects of endocytosis and membrane composition, and the effects of these perturbations on infectious focus formation were monitored. Infectivity was exquisitely sensitive to methyl-β-cyclodextrin (MβCD) and nystatin, which disrupt lipid rafts by removing cholesterol. These agents inhibited infection by enveloped, but not cell-associated, varicella-zoster virus (VZV) in a dose-dependent manner and exerted these effects on both target cell and viral membranes. Inhibition by MβCD, which could be reversed by cholesterol replenishment, rapidly declined as a function of time after exposure of target cells to VZV, suggesting that an early step in viral infection requires cholesterol. No effect of cholesterol depletion, however, was seen on viral binding; moreover, there was no reduction in the surface expression or internalization of mannose 6-phosphate receptors, which are required for VZV entry. Viral entry was energy dependent and showed concentration-dependent inhibition by chlorpromazine, which, among other actions, blocks clathrin-mediated endocytosis. These data suggest that both membrane lipid composition and clathrin-mediated transport are critical for VZV entry. Lipid rafts are likely to contribute directly to viral envelope integrity and, in the host membrane, may influence endocytosis, evoke downstream signaling, and/or facilitate membrane fusion.

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

Journal of Biological Chemistry ◽

10.1074/jbc.m114.574285 ◽

2014 ◽

Vol 289 (44) ◽

pp. 30842-30856 ◽

Cited By ~ 16

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.

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Cyanovirin-N Binds to gp120 To Interfere with CD4-Dependent Human Immunodeficiency Virus Type 1 Virion Binding, Fusion, and Infectivity but Does Not Affect the CD4 Binding Site on gp120 or Soluble CD4-Induced Conformational Changes in gp120

Journal of Virology ◽

10.1128/jvi.73.5.4360-4371.1999 ◽

1999 ◽

Vol 73 (5) ◽

pp. 4360-4371 ◽

Cited By ~ 93

Author(s):

Mark T. Esser ◽

Toshiyuki Mori ◽

Isabelle Mondor ◽

Quentin J. Sattentau ◽

Barna Dey ◽

...

Keyword(s):

Human Immunodeficiency Virus ◽

Conformational Changes ◽

Target Cell ◽

Envelope Glycoprotein ◽

Target Cells ◽

Immunodeficiency Virus ◽

Cd4 Binding Site ◽

Hiv 1 ◽

Soluble Cd4

ABSTRACT Cyanovirin-N (CV-N), an 11-kDa protein isolated from the cyanobacterium Nostoc ellipsosporum, potently inactivates diverse strains of human immunodeficiency virus type 1 (HIV-1), HIV-2, and simian immunodeficiency virus. While it has been well established that the viral surface envelope glycoprotein gp120 is a molecular target of CV-N, the detailed mechanism of action is of further interest. We compared matched native and CV-N-treated virus preparations in a panel of assays that measure viral replication, assessing successive stages of the viral life cycle. CV-N-treated virions failed to infect cells as detected by p24 production and quantitative PCR for HIV-1 reverse transcription products, whereas treatment of the target cells did not block infection, confirming that CV-N acts at the level of the virus, not the target cell, to abort the initial infection process. Compared to native HIV-1 preparations, CV-N-treated HIV-1 virions showed impaired CD4-dependent binding to CD4+ T cells and did not mediate “fusion from without” of CD4+ target cells. CV-N also blocked HIV envelope glycoprotein Env-induced, CD4-dependent cell-cell fusion. Mapping studies with monoclonal antibodies (MAbs) to defined epitopes on the HIV-1 envelope glycoprotein indicated that CV-N binds to gp120 in a manner that does not occlude or alter the CD4 binding site or V3 loop or other domains on gp120 recognized by defined MAbs and does not interfere with soluble CD4-induced conformational changes in gp120. Binding of CV-N to soluble gp120 or virions inhibited subsequent binding of the unique neutralizing MAb 2G12, which recognizes a glycosylation-dependent epitope. However, prior binding of 2G12 MAb to gp120 did not block subsequent binding by CV-N. These results help clarify the mechanism of action of CV-N and suggest that the compound may act in part by preventing essential interactions between the envelope glycoprotein and target cell receptors. This proposed mechanism is consistent with the extensive activity profile of CV-N against numerous isolates of HIV-1 and other lentiviruses and supports the potential broad utility of this protein as a microbicide to prevent the sexual transmission of HIV.

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The Polar Region of the HIV-1 Envelope Protein Determines Viral Fusion and Infectivity by Stabilizing the gp120-gp41 Association

Journal of Virology ◽

10.1128/jvi.02128-18 ◽

2019 ◽

Vol 93 (7) ◽

Cited By ~ 2

Author(s):

Wuxun Lu ◽

Shuliang Chen ◽

Jingyou Yu ◽

Ryan Behrens ◽

Joshua Wiggins ◽

...

Keyword(s):

Membrane Fusion ◽

Cell Fusion ◽

Viral Entry ◽

Polar Region ◽

Targeted Mutagenesis ◽

Target Cells ◽

Viral Fusion ◽

Underlying Mechanisms ◽

Hiv 1

ABSTRACT HIV-1 enters cells through binding between viral envelope glycoprotein (Env) and cellular receptors to initiate virus and cell fusion. HIV-1 Env precursor (gp160) is cleaved into two units noncovalently bound to form a trimer on virions, including a surface unit (gp120) and a transmembrane unit (gp41) responsible for virus binding and membrane fusion, respectively. The polar region (PR) at the N terminus of gp41 comprises 17 residues, including 7 polar amino acids. Previous studies suggested that the PR contributes to HIV-1 membrane fusion and infectivity; however, the precise role of the PR in Env-mediated viral entry and the underlying mechanisms remain unknown. Here, we show that the PR is critical for HIV-1 fusion and infectivity by stabilizing Env trimers. Through analyzing the PR sequences of 57,645 HIV-1 isolates, we performed targeted mutagenesis and functional studies of three highly conserved polar residues in the PR (S532P, T534A, and T536A) which have not been characterized previously. We found that single or combined mutations of these three residues abolished or significantly decreased HIV-1 infectivity without affecting viral production. These PR mutations abolished or significantly reduced HIV-1 fusion with target cells and also Env-mediated cell-cell fusion. Three PR mutations containing S532P substantially reduced gp120 and gp41 association, Env trimer stability, and increased gp120 shedding. Furthermore, S532A mutation significantly reduced HIV-1 infectivity and fusogenicity but not Env expression and cleavage. Our findings suggest that the PR of gp41, particularly the key residue S532, is structurally essential for maintaining HIV-1 Env trimer, viral fusogenicity, and infectivity. IMPORTANCE Although extensive studies of the transmembrane unit (gp41) of HIV-1 Env have led to a fusion inhibitor clinically used to block viral entry, the functions of different domains of gp41 in HIV-1 fusion and infectivity are not fully elucidated. The polar region (PR) of gp41 has been proposed to participate in HIV-1 membrane fusion in biochemical analyses, but its role in viral entry and infectivity remain unclear. In our effort to characterize three nucleotide mutations of an HIV-1 RNA element that partially overlaps the PR coding sequence, we identified a novel function of the PR that determines viral fusion and infectivity. We further demonstrated the structural and functional impact of six PR mutations on HIV-1 Env stability, viral fusion, and infectivity. Our findings reveal the previously unappreciated function of the PR and the underlying mechanisms, highlighting the important role of the PR in regulating HIV-1 fusion and infectivity.

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Nuclear Import Defect of Human Immunodeficiency Virus Type 1 DNA Flap Mutants Is Not Dependent on the Viral Strain or Target Cell Type

Journal of Virology ◽

10.1128/jvi.00974-06 ◽

2006 ◽

Vol 80 (20) ◽

pp. 10262-10269 ◽

Cited By ~ 26

Author(s):

Nathalie Arhel ◽

Sandie Munier ◽

Philippe Souque ◽

Karine Mollier ◽

Pierre Charneau

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Target Cell ◽

Nuclear Import ◽

Target Cells ◽

Virus Infectivity ◽

Immunodeficiency Virus ◽

Hiv 1

ABSTRACT We have previously established, using human immunodeficiency virus type 1 (HIV-1) strain LAI, that the HIV-1 central DNA Flap acts as a cis determinant of viral genome nuclear import. Although the impact of the DNA Flap on nuclear import has already found numerous independent confirmations in the context of lentivirus vectors, it has been claimed that it may be nonessential for infectious virus strains LAI, YU-2 (J. D. Dvorin et al., J. Virol. 76:12087-12096, 2002), HXB2, and NL4-3 (A. Limon et al., J. Virol. 76:12078-12086, 2002). We conducted a detailed analysis of virus infectivity using the provirus clones provided by the authors and analogous target cells. In contrast to published data, our results show that all cPPT mutant viruses exhibit reduced infectivity corresponding to a nuclear import defect irrespective of the viral genetic background or target cell.

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Oregano essential oil inhibits Candida spp. biofilms

Zeitschrift für Naturforschung C ◽

10.1515/znc-2021-0002 ◽

2021 ◽

Vol 0 (0) ◽

Author(s):

Mayram Hacioglu ◽

Ozlem Oyardi ◽

Alpcan Kirinti

Keyword(s):

Essential Oil ◽

Biofilm Formation ◽

The Body ◽

Candida Spp ◽

Antifungal Effect ◽

Oregano Essential Oil ◽

Mucosal Surfaces ◽

Staphyloccocus Aureus ◽

Oregano Oil ◽

First Time

Abstract Candida spp. can form biofilms on mucosal surfaces and epithelial cells as well as on devices implanted in the body such as catheters and dentures, which are thought to underlie the most recalcitrant infections. It was aimed to show antifungal and antibiofilm activities of oregano oil (Origanum onites). The antifungal activities of some essential oils were investigated against C. spp. and among them, oregano oil was found to be the most effective oil and further biofilm studies were conducted with it. Oregano oil inhibited biofilm adhesion and formation of C. spp. and mature biofilms and also displayed the ability to reduce biofilm formation when they were allowed to form on surfaces previously coated with oil (up to 50% inhibition rates). In addition, oregano oil was found to be effective against dual biofilms of Candida albicans + Staphyloccocus aureus at different concentrations. This study suggests that O. onites essential oil has useful antibiofilm effects against C. spp. The inhibitory effects of O. onites essential oil, against C. spp., were demonstrated for the first time. It also had antifungal effect on biofilm formation and established biofilm even at MIC level.

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Time-resolved Imaging of HIV-1 Env-mediated Lipid and Content Mixing between a Single Virion and Cell Membrane

Molecular Biology of the Cell ◽

10.1091/mbc.e05-06-0496 ◽

2005 ◽

Vol 16 (12) ◽

pp. 5502-5513 ◽

Cited By ~ 53

Author(s):

Ruben M. Markosyan ◽

Fredric S. Cohen ◽

Grigory B. Melikyan

Keyword(s):

Temperature Jump ◽

Viral Envelope ◽

Pseudotyped Virus ◽

Target Cells ◽

Time Resolved ◽

Virus Content ◽

Long Time ◽

Viral Envelopes ◽

Time Resolved Imaging ◽

Hiv 1

A method has been developed to follow fusion of individual pseudotyped virus expressing HIV-1 Env to cells by time-resolved fluorescence microscopy. Viral envelopes were labeled with a fluorescent lipid dye (DiD) and virus content was rendered visible by incorporating a Gag-GFP chimera. The Gag-GFP is naturally cleaved to the much smaller NC-GFP fragment in the mature virions. NC-GFP was readily released upon permeabilization of the viral envelope, whereas the capsid was retained. The NC-GFP thus provides a relatively small and mobile aqueous marker to follow viral content transfer. In fusion experiments, virions were bound to cells at low temperature, and fusion was synchronously triggered by a temperature jump. DiD transferred from virions to cells without a significant lag after the temperature jump. Some virions released DiD but retained NC-GFP. Surprisingly, the fraction of lipid mixing events yielding NC-GFP transfer was dependent on the type of target cell: of three infectable cell lines, only one permitted NC-GFP transfer within minutes of raising temperature. NC-GFP release did not correlate with the level of CD4 or coreceptor expression in the target cells. The data indicate that fusion pores formed by HIV-1 Env can remain small for a relatively long time before they enlarge.

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