Biology subjects

Solis, N.

Publications and source records attributed to Solis, N..

5 recordsLinked to original sources

bioRxiv2024

YibN, a bona fide interactor of YidC with implications in membrane protein insertion and membrane lipid production

YidC, a prominent member of the Oxa1 superfamily, is essential for the biogenesis of the bacterial inner membrane, significantly influencing its protein composition and lipid organization. It interacts with the Sec translocon, aiding the proper folding of multi-pass membrane proteins. It also functions independently, serving as an insertase and lipid scramblase, augmenting the insertion of smaller membrane proteins while contributing to the organization of the bilayer. Despite the wealth of structural and biochemical data available, how YidC operates remains unclear. To investigate this, we employed proximity-dependent biotin labeling (BioID), leading to the identification of YibN as a crucial component within the YidC protein environment. We then demonstrated the association between YidC and YibN by affinity purification-mass spectrometry assays conducted on native membranes, with further confirmation using on-gel binding assays with purified proteins. Co-expression studies and in vitro assays indicated that YibN enhances the production and membrane insertion of YidC substrates, such as M13 and Pf3 phage coat proteins, ATP synthase subunit c, and various small membrane proteins like SecG. Additionally, the overproduction of YibN was found to stimulate membrane lipid production and promote inner membrane proliferation, perhaps by interfering with YidC lipid scramblase activity. Consequently, YibN emerges as a significant physical and functional interactor of YidC, influencing membrane protein insertion and lipid organization.

biochemistry↗

bioRxiv2022

Plasma membrane phosphatidylinositol-4-phosphate is not necessary for Candida albicans viability, yet is key for cell wall integrity and systemic infection

Phosphatidylinositol phosphates are key phospholipids with a range of regulatory roles, including membrane trafficking and cell polarity. Phosphatidylinositol-4-phosphate [PI(4)P] at the Golgi is required for the budding to filamentous growth transition in the human pathogenic fungus Candida albicans, however the role of plasma membrane PI(4)P is unclear. We have investigated the importance of this phospholipid in C. albicans growth, stress response, and virulence by generating mutant strains with decreased levels of plasma membrane PI(4)P, via deletion of components of the PI-4-kinase complex, i.e. Efr3, Ypp1 and Stt4. The amount of plasma membrane PI(4)P in the efr3{Delta}/{Delta} and ypp1{Delta}/{Delta} mutant was [~]60% and [~]40% of the wild-type strain, respectively, whereas it was nearly undetectable in the stt4{Delta}/{Delta} mutant. All three mutants had reduced plasma membrane phosphatidylserine (PS). Although these mutants had normal yeast phase growth, they were defective in filamentous growth, exhibited defects in cell wall integrity and had an increased exposure of cell wall {beta}(1,3)-glucan, yet they induced a range of hyphal specific genes. In a mouse model of hematogenously disseminated candidiasis, fungal plasma membrane PI(4)P levels directly correlated with virulence; the efr3{Delta}/{Delta} had wild-type virulence, the ypp1{Delta}/{Delta} mutant had attenuated virulence and the stt4{Delta}/{Delta} mutant caused no lethality. In the mouse model of orpharyngeal candidiasis, only the ypp1{Delta}/{Delta} mutant had reduced virulence, indicating that plasma membrane PI(4)P is less important for proliferation in the oropharynx. Collectively, these results demonstrate that plasma membrane PI(4)P levels play a central role in filamentation, cell wall integrity and virulence in C. albicans. ImportanceWhile the PI-4-kinases Pik1 and Stt4 both produce PI(4)P, the former generates PI(4)P at the Golgi and the latter at the plasma membrane and these two pools are functionally distinct. To address the importance of plasma membrane PI(4)P in Candida albicans, we have generated deletion mutants of the three putative plasma membrane PI-4-kinase complex components and quantified the levels of plasma membrane PI(4)P in each of these strains. Our work reveals that this phosphatidylinositol phosphate is specifically critical for the yeast-to-hyphal transition, cell wall integrity and virulence in a mouse systemic infection model. The significance of this work is in identifying a plasma membrane phospholipid that has an infection specific role, which is attributed to the loss of plasma membrane PI(4)P resulting in {beta}(1,3)-glucan unmasking.

cell biology↗

bioRxiv2020

Control of ss-glucan exposure by the endo-1,3-glucanase Eng1 in Candida albicans modulates virulence

Candida albicans is a major cause of invasive candidiasis, which has a high mortality rate. The hyphal form of C. albicans is virulent and activates the host innate immune response, while the yeast form is hypovirulent and less immunogenic. The innate immune response is critical for host defense, but overactivation can cause tissue damage and sepsis. The innate immune response can be triggered when the C-type lectin receptor Dectin-1 recognizes {beta}-glucans, which is protected by the outer mannan layer of the cell wall on C. albicans. Here, we demonstrate that there is low level of Dectin-1 binding at the septum of yeast cells, but high level of Dectin-1 binding over the entire surface of hyphae. We find that {beta}-glucan masking in yeast is controlled by two highly expressed yeast proteins, the endo-1,3-{beta}-glucanase Eng1 and the Yeast Wall Protein Ywp1. An eng1 deletion mutant shows enhanced Dectin-1 binding at the septa, while an eng1 ywp1 double mutant, but not an ywp1 single mutant, shows strong overall Dectin-1 binding. Thus, Eng1-mediated {beta}-glucan trimming and Ywp1-mediated {beta}-glucan masking are two parallel mechanisms utilized by C. albicans yeast to minimize recognition by Dectin-1. In the model of disseminated candidiasis, mice infected with the eng1 deletion mutant showed delayed mortality with an increased renal immune response in males compared to mice infected with the wild-type strain, but earlier mortality with a higher renal immune response in females. Using the eng1 mutant that is specifically defective in {beta}-glucan masking in yeast, this study demonstrates that the level of {beta}-glucan exposure is important for modulating the balance between immune protection and immunopathogenesis. Abstract ImportanceCandida albicans is a major opportunistic fungal pathogen of humans. Systemic Candidiasis has high mortality rates. C. albicans is also a constituent of the human microbiome and found in gastrointestinal and genitourinary tracts of most healthy individuals. C. albicans is able to switch reversibly between yeast and hyphae in response to environmental cues. The hyphal form is virulent, while the yeast form is hypovirulent and less immunogenic. This study demonstrates that {beta}-glucan exposure in yeast is protected by two highly expressed yeast proteins, the endo-1,3-{beta}-glucanase Eng1 and the Yeast Wall Protein Ywp1. Eng1-mediated {beta}-glucan trimming and Ywp1-mediated {beta}-glucan masking are two parallel mechanisms utilized by C. albicans yeast to minimize recognition by the host C-type lectin receptor Dectin-1. The eng1 mutant triggers a higher immune response and leads to earlier mortality compared to the wild-type strain. Thus, {beta}-glucan masking in yeast keeps yeast cells less immunogenic and hypovirulent.

microbiology↗

bioRxiv2018

Activation of the epidermal growth factor receptor initiates innate immune responses during oropharyngeal candidiasis

During oropharyngeal candidiasis (OPC), Candida albicans invades and damages oral epithelial cells, which respond by producing proinflammatory mediators that recruit phagocytes to foci of infection. The ephrin type-A receptor 2 (EphA2) detects {beta}-glucan and plays a central role in stimulating epithelial cells to release proinflammatory mediators during OPC. The epidermal growth factor receptor (EGFR) also interacts with C. albicans and is known to be activated by the Als3 adhesin/invasin and the Ece1/Candidalysin pore-forming toxin. Here, we investigated the interactions among EphA2, EGFR, Als3 and Ece1/Candidalysin during OPC. We found that Als3 and Ece1/Candidalysin function in the same pathway to damage epithelial cells in vitro. They also work together to cause OPC in mice. EGFR and EphA2 constitutively associate with each other as part of a physical complex and are mutually dependent for C. albicans-induced activation. In vitro, either Als3 or Ece1/Candidalysin is required for C. albicans to activate EGFR, sustain EphA2 activation, and stimulate epithelial cells to secrete CXCL8/IL-8 and CCL20. In the mouse model of OPC, Ece1/Candidalysin alone activates EGFR and induces CXCL1/KC and CCL20 production. Ece1/Candidalysin is also necessary for the production of IL-1 and IL-17A independently of Als3 and EGFR. These results delineate the complex interplay between host cell receptors and C. albicans virulence factors during the induction of OPC and the resulting oral inflammatory response. Author summaryOropharyngeal candidiasis occurs when the fungus Candida albicans proliferates in the mouth. The disease is characterized by fungal invasion of the superficial epithelium and a localized inflammatory response. Two C. albicans virulence factors contribute to the pathogenesis of OPC, Als3 which enables the organisms to adhere to and invade host cells and Ece1/Candidalysin which is pore-forming toxin that damages host cells. Two epithelial cell receptors, ephrin type-A receptor 2 (EphA2) and the epidermal growth factor receptor (EGFR) are activated by C. albicans. Here, we show that EphA2 and EGFR form part of complex and that each receptor is required to activate the other. Als3 and Ece1/Candidalysin function in the same pathway to damage epithelial cells. In isolated epithelial cells, both of these virulence factors activate EphA2 and EGFR, and stimulate the production of inflammatory mediators. In the mouse model of OPC, Ece1/Candidalysin elicits of a subset of the oral inflammatory response. Of the cytokines and chemokines induced by this toxin, some require the activation of EGFR while others are induced independently of EGFR. This work provides a deeper understanding of the interactions among C. albicans virulence factors and host cell receptors during OPC.

microbiology↗

bioRxiv2018

Intersection of phosphate transport, oxidative stress and TOR signalling in Candida albicans virulence

Phosphate is an essential macronutrient required for cell growth and division. Pho84 is the major high-affinity cell-surface phosphate importer of Saccharomyces cerevisiae and a crucial element in the phosphate homeostatic system of this model yeast. We found that loss of Candida albicans Pho84 attenuated virulence in Drosophila and murine oropharyngeal and disseminated models of invasive infection, and conferred hypersensitivity to neutrophil killing. Susceptibility of cells lacking Pho84 to neutrophil attack depended on reactive oxygen species (ROS): pho84-/- cells were no more susceptible than wild type C. albicans to neutrophils from a patient with chronic granulomatous disease, or to those whose oxidative burst was pharmacologically inhibited or neutralized. pho84-/- mutants hyperactivated oxidative stress signalling. They accumulated intracellular ROS in the absence of extrinsic oxidative stress, in high as well as low ambient phosphate conditions. ROS accumulation correlated with diminished levels of the unique superoxide dismutase Sod3 in pho84-/- cells, while SOD3 overexpression from a conditional promoter substantially restored these cells oxidative stress resistance in vitro. Repression of SOD3 expression sharply increased their oxidative stress hypersensitivity. Neither of these oxidative stress management effects of manipulating SOD3 transcription was observed in PHO84 wild type cells. Sod3 levels were not the only factor driving oxidative stress effects on pho84-/- cells, though, because overexpressing SOD3 did not ameliorate these cells hypersensitivity to neutrophil killing ex vivo, indicating Pho84 has further roles in oxidative stress resistance and virulence. Measurement of cellular metal concentrations demonstrated that diminished Sod3 expression was not due to decreased import of its metal cofactor manganese, as predicted from the function of S. cerevisiae Pho84 as a low-affinity manganese transporter. Instead of a role of Pho84 in metal transport, we found its role in TORC1 activation to impact oxidative stress management: overexpression of the TORC1-activating GTPase Gtr1 relieved the Sod3 deficit and ROS excess in pho84-/- null mutant cells, though it did not suppress their hypersensitivity to neutrophil killing or hyphal growth defect. Pharmacologic inhibition of Pho84 by small molecules including the FDA-approved drug foscarnet also induced ROS accumulation. Inhibiting Pho84 could hence support host defenses by sensitizing C. albicans to oxidative stress.

microbiology↗