Biology subjects

Dhillon, G.

Publications and source records attributed to Dhillon, G..

3 recordsLinked to original sources

bioRxiv2026

Local Confinement within Plasma Membrane Nanodomains Drives Constitutive Activity of GPCRs

Many G protein coupled receptors (GPCRs) exhibit constitutive (basal) activity, where they can signal in the absence of ligand binding through spontaneous conformational transitions that facilitate G protein coupling and downstream signaling. This intrinsic baseline activity is critical for cellular homeostasis and can be modulated by the receptors conformational ensemble, membrane organization, and interactions with intracellular effectors. In this study, we use live-cell signaling assays, fluorescence cross-correlation spectroscopy (FCCS), and single-particle tracking (SPT) to investigate how membrane organization influences the basal activity of two class A GPCRs: the M1 muscarinic receptor (M1R) and the adenosine A2A receptor (A2AR). In live-cell signalling assays, M1R showed minimal agonist-independent Ca{superscript 2} responses, while A2AR exhibited significant basal cAMP production that was eliminated by an inverse agonist. FCCS showed that, without ligand, only a small portion of M1R co-diffuses with its cognate G11 protein, whereas a much larger fraction of A2AR co-diffuses with the GS protein. SPT revealed that A2AR, but not M1R, is enriched in slowly diffusing, confined states with spatial scales around 150-200 nm and sensitivity to cholesterol- and raft-modulating agents, consistent with localization in lipid-raft nanodomains. Dual-color tracking and diffusion mapping demonstrated that a significant portion of A2AR and GS share confinement domains under basal conditions, while M1R and G11 only show such co-confinement in the active state. These findings support a model where the co-confinement of GPCRs and G proteins within plasma membrane nanodomains--rather than stable pre-coupled RG complexes-- determines the level of constitutive GPCR activity.

biophysics↗

bioRxiv2025

Macrophage ferroptosis inhibits Aspergillus conidial killing in lung transplantation

Immune suppression heightens the risk for fungal infections, but the mechanisms that result in clinical disease are poorly understood. Here we demonstrate that macrophage ferroptosis, an iron-dependent form of regulated cell death, inhibits Aspergillus fumigatus (Af) killing. In a mouse tracheal transplant model of Af infection, we observed an increase in macrophage lipid peroxidation, a decreased expression of negative ferroptosis regulators Gpx4 and Slc7a11, and an increase in positive regulators Ptgs2 and Nox2, relative to syntransplants. Depletion of macrophages in transplant recipients decreased Af invasion. In vitro, iron overload reduced macrophage viability and decreased their capability to kill Af spores, through a decrease in lysosomal acidification and lysosomal loss. Treatment with ferrostatin-1, a ferroptosis inhibitor, and deferasirox (an iron chelator) restored Af killing. Ferroptotic alveolar macrophages isolated from lung transplant patients also showed a decreased ability to kill Af spores and the patients bronchoalveolar lavage was characterized by higher iron levels and markers of ferroptotic stress compared to non-lung transplants. These characteristics were strongly correlated with a clinical history of fungal infections, independent of immune suppressive medications. Our findings indicate that macrophage ferroptosis augments the risk of invasive aspergillosis, representing a novel mechanism for host immune dysfunction. Graphical AbstractSchematic of proposed mechanism underlying ferroptosis induced immune dysregulation and increased Af invasion in lung transplantation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/643092v1_ufig1.gif" ALT="Figure 1"> View larger version (104K): org.highwire.dtl.DTLVardef@1112102org.highwire.dtl.DTLVardef@18e2ec1org.highwire.dtl.DTLVardef@19f9c5forg.highwire.dtl.DTLVardef@1d325e8_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

Forward genetic screen in zebrafish identifies new fungal regulators that limit host-protective Candida-innate immune interaction

Candida is one of the most frequent causes of bloodstream infections, and our first line of defense against these invasive infections is the innate immune system. The early immune response is critical in controlling C. albicans infection, but C. albicans has several strategies to evade host immune attack. Phagocytosis of C. albicans blocks hyphal growth, limiting host damage and virulence, but how C. albicans limits early recruitment and phagocytosis in vertebrate infection is poorly understood. To study innate immune evasion by intravital imaging, we utilized the transparent larval zebrafish infection model to screen 131 C. albicans mutants for altered virulence and phagocyte response. Infections with each of seven hypovirulent mutants led to altered phagocyte recruitment and/or phagocytosis, falling into four categories. Of particular interest among these is NMD5, a predicted {beta}-importin and newly-identified virulence factor. The nmd5{Delta}/{Delta} mutant fails to limit phagocytosis and its virulence defects are eliminated when phagocyte activity is compromised, suggesting that its role in virulence is limited to immune evasion. These quantitative intravital imaging experiments are the first to document altered Candida-phagocyte interactions for several additional mutants, and clearly distinguish recruitment from phagocytic uptake, suggesting that Candida modulates both events. This initial large-scale screen of individual C. albicans mutants in a vertebrate, coupled with high-resolution imaging of Candida-phagocyte interactions, provides a more nuanced view of how diverse mutations can lead to more effective phagocytosis, a key immune process which blocks germination and drives anti-fungal immunity. ImportanceCandida albicans is part of the human microbial community and is a dangerous opportunistic pathogen, able to prevent its elimination by the host immune system. Although Candida avoids immune attack through several strategies, we still understand little about how it regulates when immune phagocytes get recruited to the infection site and when they engulf fungal cells. We tested over 130 selected Candida mutants for their ability to cause lethal infection and found several avirulent mutants which provoked altered innate immune responses, resulting in lower overall inflammation and greater host survival. Of particular interest is NMD5, which acts to limit fungal phagocytosis and is predicted to regulate the activity of stress-associated transcription factors. Our high-content screening was enabled by modeling Candida infection in transparent vertebrate zebrafish larva. Our findings help us understand how Candida survives immune attack during commensal and pathogenic growth, and may eventually inform new strategies for controlling disease.

microbiology↗