Biology subjects

Ayscough, K. R.

Publications and source records attributed to Ayscough, K. R..

3 recordsLinked to original sources

bioRxiv2024

CryoEM Reconstruction of Yeast ADP-Actin Filament at 2.5 Angstrom resolution. A comparison with mammalian and avian F-actin.

The core component of the actin cytoskeleton is the globular protein G-actin, which reversibly polymerises into filaments (F-actin). Budding yeast possesses a single actin which shares 87-89% sequence identity with vertebrate actin isoforms. Previous structural studies indicate very close overlap of main-chain backbones. Intriguingly however, substitution of yeast ACT1 with vertebrate {beta}-cytoplasmic actin severely disrupts cell function and substitution with a skeletal muscle isoform is lethal. Here we report a 2.5 [A] structure of budding yeast F-actin. Previously unresolved side-chain information now highlights four main differences in the comparison of yeast and vertebrate ADP F-actins: a more open nucleotide binding pocket; a more solvent exposed C-terminus; a rearrangement of intersubunit binding interactions in the vicinity of the D-loop and changes in the hydrogen bonding network in the vicinity of histidine 73 (yeast actin) and methyl-histidine 73 (vertebrate actin).

biochemistry↗

bioRxiv2023

Candida albicans suppression of neutrophil reactive nitrogen species is rescued by host Hif-1alpha in vivo

Candida albicans is a human commensal that can cause life-threatening invasive infection in immunocompromised individuals. Human immunity to C. albicans infection is thought to be largely dependent on neutrophil reactive oxygen and nitrogen species (ROS/RNS) generation by neutrophils. Despite this, our understanding of innate immune killing and escape by C. albicans is primarily studied in macrophages and the precise mechanisms of evasion are unclear in neutrophils. Here we sought to determine the importance of neutrophil reactive nitrogen species (RNS) production during C. albicans infection in vivo. Using a zebrafish model, we found that C. albicans rapidly downregulated neutrophil RNS below basal levels during the first day post infection, a time at which neutrophil RNS is upregulated in bacterial infections as an important host-defense mechanism, indicating fungal evasion of host neutrophils. We confirmed downregulation of RNS in human primary neutrophils and with clinical Candida spp. isolates, including emerging human pathogens C. auris and C. glabrata. Using a zebrafish arginase 2 transgenic line and a C. albicans car1{Delta} mutant, we show that both host and fungal arginase contribute the reduction in neutrophil RNS. Despite pathogen downregulation, upregulation of neutrophil RNS via Hypoxia inducible factor (Hif)-1 stabilisation, was sufficient to improve C. albicans infection survival, dependent on the presence of neutrophils and Nitric oxide synthase 2 (Nos2). Finally, restoration of neutrophil RNS, via Hif-1 stabilisation, was synergistic with clinically relevant antifungal treatment, increasing survival and clearance of C. albicans infections. Together, these data demonstrate that restoration of the neutrophil RNS response in C. albicans infection improves infection outcomes, highlighting the potential of targeting Hif-1 and RNS in host directed therapies against fungal infections.

immunology↗

bioRxiv2023

Candida albicans cells lacking AP-2 have defective hyphae and are avirulent despite increased host uptake and intracellular proliferation in macrophages

Candida albicans is a commensal microbe and opportunistic human pathogen. The yeast can be recognised and taken up by macrophages via interactions with its cell wall, a complex polysaccharide structure containing several components that are specifically recognised by immune cell receptors. Following uptake Candida can respond in the host environment by switching from a yeast to hyphal morphology which facilitates escape from macrophages and allows subsequent invasion of host tissues. Disruption of Candidas ability to form hyphae results in reduced virulence and fitness for survival in the host environment. Candida albicans cells lacking AP-2, an endocytic adaptor complex, have increased cell wall chitin and morphologically defective hyphae in vitro. Previous studies have correlated increased chitin with decreased recognition by macrophages, possibly due to masking of cell wall beta-glucan which is recognised by dectin receptors. Despite the cell wall changes in the mutant strain there was an unexpected increased uptake of the mutant. Increased chitin did not reduce phagocytosis and additional uptake was not due to compensatory elevated exposure of beta-glucan, highlighting the importance of cell wall components beyond chitin and glucan for macrophage engagement and uptake. Furthermore, the apm4 mutant exhibited parasitism of macrophages, surviving and proliferating within the phagosome, a phenotype that was then replicated with a well-characterised yeast locked mutant. Finally, the combined phenotype of reduced hyphal formation but continued proliferation resulted in reduced virulence despite an equivalent burden of infection to a wild-type Candida infection, as determined using a zebrafish larval model of candidiasis.

microbiology↗