Biology subjects

Whitfield, P.

Publications and source records attributed to Whitfield, P..

3 recordsLinked to original sources

bioRxiv2025

The Type VI Secretion System Antifungal Effector Tfe2 Inhibits Protein Translation and Drives Hyperactivation of TORC1.

Type VI Secretion Systems (T6SS) are utilised by many bacteria to deliver toxic effectors into neighbouring bacterial, fungal or host cells. Whilst many antibacterial effectors are well characterised, much less is known regarding the identity or mode-of-action of antifungal effectors. Here we combine structural modelling with proteomics and in vivo approaches, to show that the Serratia marcescens antifungal effector Tfe2 adopts a novel fold and functions as a potent inhibitor of protein translation leading to hyperactivation of the TORC1 kinase. We show that Tfe2 expression in Saccharomyces cerevisiae, or treatment with the protein translation inhibitor cycloheximide, drive identical increases in free intracellular amino acids and hyperactivation of TORC1. This, in turn, triggers the Tfe2 and cycloheximide-mediated rapid turnover of amino acid transporters through stimulating substrate-independent endocytosis. Polysome profiling, however, revealed differences in Tfe2 and cycloheximide-mediated protein translation inhibition, with Tfe2 inhibiting initiation of translation. Tfe2-mediated hyperactivation of TORC1 may also underpin adaptive responses to this effector which include significant remodelling of the lipidome and notable alterations in organelle and cell wall structures. Collectively this study has provided new insight into the mode-of-action of a structurally novel antifungal effector Tfe2.

microbiology↗

bioRxiv2025

Inhibition of systemic mammalian metabolism by carnitine mimics from the gut microbiota

BackgroundThe gut microbiota and microbiome-derived metabolites are implicated in various aspects of human health. Here we sought to determine the systemic effects, and mechanism of action, of microbiome-derived carnitine analogues in germ free and conventionally colonised mice. ResultsHere we report the systemic localization of the microbiome-derived carnitine analogues, 3-methyl-4-(trimethylammonio)butanoate (3M-4-TMAB) and 5-aminovalerate betaine (5-AVAB), post-administration to germ free mice, with systemic carnitine depletion and mitochondrial dysregulation, reflected in altered acylcarnitine profiles due to incomplete carnitine-mediated fatty acid oxidation. Studies on the inhibitory potency of 3M-4-TMAB at the enzymatic, cellular, and organism levels indicate that, in part, this is a result of inhibition of gammabutyrobetaine hydroxylase, which catalyses the final step in carnitine biosynthesis. Systemic administration of 13C- labelled 3M-4-TMAB to conventionally colonised animals to further investigate the physiological relevance of this inhibition, identified a significant reduction in systemic carnitine levels due to increased excretion in urine and faeces and disruption of carnitine-mediated metabolism across ten organs. ConclusionsThese results highlight the physiological relevance and significance of microbiome-derived metabolites in vivo. The depletion of carnitine and inhibition of its function have potentially long-term impacts on both mammalian energy generation and the protective, signalling and immune regulatory effects of this critical molecule.

microbiology↗

bioRxiv2022

Cell overgrowth during G1 arrest triggers an osmotic stress response and chronic p38 activation to promote cell cycle exit

Cell size and the cell cycle are intrinsically coupled and abnormal increases in cell size are associated with senescence. The mechanism by which overgrowth primes cells to exit the cell cycle remains unclear. We investigate this using CDK4/6 inhibitors that arrest cell cycle progression in G0/G1 and are used to treat ER+/HER2-metastatic breast cancer. We demonstrate that long-term CDK4/6 inhibition promotes cellular overgrowth during the G0/G1 arrest, causing widespread proteome remodeling and p38-p53-p21-dependent cell cycle exit. Cell cycle exit is triggered by two waves of p21 induction. First, overgrowth during a G0/G1 arrest induces an osmotic stress response, producing the first wave of p21 induction. Second, when CDK4/6 inhibitors are removed, a fraction of cells escape G0/G1 arrest and enter S-phase where overgrowth-driven replication stress results in a second wave of p21 induction that causes cell cycle withdrawal from G2, or the subsequent G1. This could explain why cellular hypertrophy is associated with senescence and why CDK4/6 inhibitors have long-lasting anti-proliferative effects in patients.

cell biology↗