Ion channels are vital components of filamentous fungi signaling in communication with their environment. We exploited the ability of the apical region of growing sporangiophores of Phycomyces ...blakesleeanus to form membrane-enveloped cytoplasmic droplets (CDs), to examine ion currents in the filamentous fungi native plasma membrane. In hypoosmotic conditions, the dominant current in the CDs is ORIC, an osmotically activated, anionic, outwardly rectified, fast inactivating instantaneous current that we have previously characterized. Here, we examined the effect of ATP on ORIC. We show that CDs contain active mitochondria, and that respiration inhibition by azide accelerates ORIC inactivation. ATP, added intracellularly, reduced ORIC run-down and shifted the voltage dependence of inactivation toward depolarized potentials, in a manner that did not require hydrolysis. Notably, ATP led to slowing down of ORIC inactivation, as evidenced by an increased time constant of inactivation, τ
, and slower decline of τ
during prolonged recordings. Flavonoids (genistein and quercetin) had the effect on ORIC opposite to ATP, acting as current inhibitors, possibly by disrupting the stabilizing effect of ATP on ORIC. The integration of osmotic sensing with ATP dependence of the anionic current, typical of vertebrate cells, is described here for the first time in filamentous fungi.
Mast cells (MCs) are an important part of the immune system, responding both to pathogens and toxins, but they also play an important role in allergic diseases, where recent data show that ...non-IgE-mediated activation is also of relevance, especially in chronic urticaria (CU) and atopic dermatitis (AD). Skin MCs express Mas-related G-protein-coupled receptor X2 (MRGPRX2), a key protein in non-IgE-dependent MC degranulation, and its overactivity is one of the triggering factors for the above-mentioned diseases, making MRGPRX2 a potential therapeutic target. Reviewing the latest literature revealed our need to focus on the discovery of MRGPRX2 activators as well as the ongoing vast research towards finding specific MRGPRX2 inhibitors for potential therapeutic approaches. Most of these studies are in their preliminary stages, with one drug currently being investigated in a clinical trial. Future studies and improved model systems are needed to verify whether any of these inhibitors may have the potential to be the next therapeutic treatment for CU, AD, and other pseudo-allergic reactions.
Studies of ion currents in filamentous fungi are a prerequisite for forming a complete understanding of their physiology. Cytoplasmic droplets (CDs), obtained from sporangiophores of Phycomyces ...blakesleeanus, are a model system that enables the characterization of ion currents in the native membrane, including the currents mediated by the channels not yet molecularly identified. Osmotically activated anionic current with outward rectification (ORIC) is a dominant current in the membrane of cytoplasmic droplets under the conditions of hypoosmotic stimulation. We have previously reported remarkable functional similarities of ORIC with the vertebrate volume regulated anionic current (VRAC), such as dose-dependent activation by osmotic difference, ion selectivity sequence, and time and voltage dependent profile of the current. Using the patch clamp method on the CD membrane, we further resolve VRAC-like ORIC characteristics in this paper. We examine the inhibition by extracellular ATP and carbenoxolone, the permeation of glutamate in presence of chloride, selectivity for nitrates, and activation by GTP, and we show its single channel behavior in excised membrane. We propose that ORIC is a functional counterpart of vertebrate VRAC in filamentous fungi, possibly with a similar essential role in anion efflux during cell volume regulation.
Role of IL‐17 in atopy—A systematic review Hofmann, Maja A.; Fluhr, Joachim W.; Ruwwe‐Glösenkamp, Christoph ...
Clinical and translational allergy,
August 2021, Volume:
11, Issue:
6
Journal Article
Peer reviewed
Open access
Purpose of Review
Atopy is defined as the genetic predisposition to react with type I allergic diseases such as food‐, skin‐, and respiratory allergies. Distinct molecular mechanisms have been ...described, including the known Th2 driven immune response. IL‐17A (IL‐17) is mainly produced by Th17 cells and belongs to the IL‐17 family of cytokines, IL‐17A to F. While IL‐17 plays a major role in inflammatory and autoimmune disorders, more data was published in recent years elucidating the role of IL‐17 in allergic diseases. The present study aimed to elaborate specifically the role of IL‐17 in atopy.
Methods
A systematic literature search was conducted in MEDLINE, Embase, and Cochrane Central Register of Controlled Trials, regarding IL‐17 and atopy/allergic diseases.
Results
In total, 31 novel publications could be identified (food allergy n = 3, allergic asthma n = 7, allergic rhinitis AR n = 10, atopic dermatitis AD n = 11). In all allergic diseases, the IL‐17 pathway has been investigated. Serum IL‐17 was elevated in all allergic diseases. In AR, serum and nasal IL‐17 levels correlated with the severity of the disease. In food allergies, serum IL‐17E was also elevated in children. In AD, there is a trend for higher IL‐17 values in the serum and skin specimen, while it is more expressed in acute lesions. In allergic asthma, serum IL‐17 levels were increased. In two studies, higher serum IL‐17 levels were found in severe persistent asthmatic patients than in intermittent asthmatics or healthy controls. Only one therapeutic clinical study exists on allergic diseases (asthma patients) using a monoclonal antibody against the IL‐17 receptor A. No clinical efficacy was found in the total study population, except for a subgroup of patients with (post‐bronchodilator) high reversibility.
Summary
The role of IL 17 in the pathogenesis of allergic diseases is evident, but the involvement of the Th17 cytokine in the pathophysiological pathway is not conclusively defined. IL‐17 is most likely relevant and will be a clinical target in subgroups of patients. The current data indicates that IL‐17 is elevated more often in acute and severe forms of allergic diseases.
ABSTRACT
Equilibrative nucleoside transporters (ENTs) translocate nucleosides and nucleobases across plasma membranes, as well as a variety of anti‐cancer, ‐viral, and ‐parasite nucleoside analogs. ...They are also key members of the purinome complex and regulate the protective and anti‐inflammatory effects of adenosine. Despite their important role, little is known about the mechanisms involved in their regulation. We conducted membrane yeast 2‐hybrid and coimmunoprecipitation studies and identified, for the first time to our knowledge, the existence of protein‐protein interactions between human ENT1 and ENT2 (hENT1 and hENT2) proteins in human cells and the formation of hetero‐ and homo‐oligomers at the plasma membrane and the submembrane region. The use of NanoLuc Binary Technology allowed us to analyze changes in the oligomeric status of hENT1 and hENT2 and how they rapidly modify the uptake profile for nucleosides and nucleobases and allow cells to respond promptly to external signals or changes in the extracellular environment. These changes in hENTs oligomerization are triggered by PKC activation and subsequent action of protein phosphatase 1.—Grañe‐Boladeras, N., Williams, D., Tarmakova, Z., Stevanovic, K., Villani, L. A., Mehrabi, P., Siu, K. W. M., Pastor‐Anglada, M., Coe, I. R. Oligomerization of equilibrative nucleoside transporters: a novel regulatory and functional mechanism involving PKC and PP1. FASEB J. 33, 3841–3850 (2019). www.fasebj.org
Background
GA2LEN‐ADCARE is a branch of the largest multidisciplinary network of research centres and clinical care in allergy and asthma, GA2LEN, focussing on the field of atopic dermatitis (AD). AD ...is a chronic inflammatory skin disease with high burden and many comorbidities requiring different levels of treatment. The need for aligned information from all involved healthcare providers led to the discussion of an integrated care pathway (ICP) plan for AD patient care involving all stakeholders and considering the complexity and variability of the disease, with a particular focus placed on the large number of patients with milder forms of AD.
Methods
The GA2LEN ADCARE network and all stakeholders, abbreviated the AD‐ICPs working group, were involved in the discussion and preparation of the AD‐ICPs during a series of subgroup workshops and meetings in years 2020 and 2021.
Results
Here we discuss the unmet needs in AD, the methodology for devising an AD‐ICP and the ICP action plan.
Conclusion
The GA2LEN ADCARE network has outlined the unmet needs in AD and provided an action plan for devising AD‐ICPs, considering the complexity and variability of the disease.
Long interspersed nuclear elements (LINE-1), a type of retrotransposon, are genetic elements in the human genome that can self-amplify and integrate back into the genome, increasing their copy ...number. Their mode of action is called retrotransposition, and it is known to affect the human genome in many ways: generating insertional mutations, genomic instability, alterations in gene expression, and contribute to genetic innovation. These elements are usually silenced in somatic tissue because of their mutagenic potential. I have identified that the evolutionary youngest and human-specific LINE-1 elements are expressed and active in human placental trophoblast cells and that LINE-1 elements are over-expressed and possibly over-active in certain patients of the pregnancy disorder pre-eclampsia (PE).PE is a pregnancy-specific complex disorder in which a combination of poor placentation and placental stress leads to hypertension and proteinuria in the mother following the 20th week of gestation. PE affects 2-8% of all pregnancies. Despite PE being the leading cause of maternal and neonatal mortality and morbidity, the etiology remains unknown. One major problem in PE research is that PE is a multi origin disorder, exhibiting different phenotypes before the onset of the disease. I hypothesized that LINE-1 overexpression and activity in an immune-mediated subtype of PE patients contributes to PE pathogenesis.I conducted RNA sequencing in 10 PE and 8 control primary human trophoblast and have identified that human-specific LINE-1 elements, L1HS, are over expressed in 3/10 patients. Using RT-PCR, western blot, and immunohistochemistry methods, I have shown that LINE1s are expressed in primary human trophoblasts at the RNA and protein levels and in all three subtypes of trophoblast cells. Using a LINE-1 retrotransposition reporter assay, I showed that trophoblast cells contain all the necessary conditions to support LINE-1 retrotransposition and that there is greater retrotransposition activity in the PE-model trophoblast cell line. In addition, I identified that LINE-1 over expression in a subset of PE patients induces the expression innate immune response, suggesting a mechanism of early pathogenesis induction in a subtype of PE patients. Lastly, I also showed that in PE, a TE-derived miRNA, has-miR-576-5p, is upregulated, among other newly discovered dyregulated miRNAs. The expression of placenta fuction-relevant genes that are predicted targets of has-miR-576-5p was investigated, but no dysregulation was determined.Overall, this research revealed novel information about the somatic expression and activity of LINE-1 in the human placenta, and for the first time provided insight into the potential role of LINE-1 in PE pathogenesis.
Compared to plants, nowadays mushrooms attract more attention as functional foods, due to a number of advantages in manipulating them. This study aimed to screen the chemical composition (fatty acids ...and phenolics) and antioxidant potential (OH•, 2,2-diphenyl-1-picrylhydrazyl (DPPH•) and ferric reducing ability of plasma (FRAP)) of two edible mushrooms, Coprinus comatus and Coprinellus truncorum, collected from nature and submerged cultivation. Partial least square regression analysis has pointed out the importance of some fatty acids—more precisely, unsaturated fatty acids (UFAs) followed by fatty acids possessing both short (C6:0 and C8:0) and long (C23:0 and C24:0) saturated chains—and phenolic compounds (such as protocatechuic acid, daidzein, p-hydroxybenzoic acid, genistein and vanillic acid) for promising anti-OH•, FRAP and anti-DPPH• activities, respectively. However, other fatty acids (C16:0, C18:0 and C18:3n3) along with the flavonol isorhamnetin are actually suspected to negatively affect (by acting pro-oxidative) the aforementioned parameters, respectively. Taken together, design of new food supplements targeting oxidative stress might be predominantly based on the various UFAs combinations (C18:2n6, C20:1, C20:2, C20:4n6, C22:2, C22:1n9, etc.), particularly if OH• is suspected to play an important role.
Studying the membrane physiology of filamentous fungi is key to understanding their interactions with the environment and crucial for developing new therapeutic strategies for disease-causing ...pathogens. However, their plasma membrane has been inaccessible for a micron-sized patch-clamp pipette for pA current recordings due to the rigid chitinous cell wall. Here, we report the first femtosecond IR laser nanosurgery of the cell wall of the filamentous fungi, which enabled patch-clamp measurements on protoplasts released from hyphae. A reproducible and highly precise (diffraction-limited, submicron resolution) method for obtaining viable released protoplasts was developed. Protoplast release from the nanosurgery-generated incisions in the cell wall was achieved from different regions of the hyphae. The plasma membrane of the obtained protoplasts formed tight and high-resistance (GΩ) contacts with the recording pipette. The entire nanosurgical procedure followed by the patch-clamp technique could be completed in less than 1 hour. Compared to previous studies using heterologously expressed channels, this technique provides the opportunity to identify new ionic currents and to study the properties of the ion channels in the protoplasts of filamentous fungi in their native environment.
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