Biology subjects

Fascione, M.

Publications and source records attributed to Fascione, M..

4 recordsLinked to original sources

bioRxiv2026

Characterisation of prostate cancer sialome re-engineering via CMAH transfection reveals a bystander effect that propagates Neu5Gc presentation to neighbouring cells

Sialic acids are a family of nine-carbon -keto sugars that play essential roles in human health and disease. In mammals, they are found in two main forms: N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc), with interactions between Neu5Ac-containing glycans and Siglec receptors on immune cells increasingly recognised as glyco-immune checkpoints, promoting immunosuppression. However, humans do not synthesise Neu5Gc due pseudogenisation of the CMAH gene which encodes the cytidine monophospho-N-acetylneuraminic acid hydroxylase enzyme responsible for CMP-Neu5Ac conversion into CMP-Neu5Gc, which then serves as the donor substrate for sialyltransferases. Here, we investigated the effects of re-engineering tumour cell-surface glycans in a prostate cancer cell model by expressing rat CMAH, thereby enabling the conversion of CMP-Neu5Ac to CMP-Neu5Gc. LNCaP cells transfected with the rat CMAH gene predominantly incorporated Neu5Gc into mucin-associated O-glycans implicated in immune suppression. Treatment with sialidase significantly reduced Neu5Gc expression, indicating that Neu5Gc was presented on cell-surface glycans, while cell-tracing experiments demonstrated the transfer of Neu5Gc to neighbouring cells, revealing a potential bystander effect capable of propagating Neu5Gc expression within the tumour microenvironment.

cancer biology↗

bioRxiv2026

Structural basis for covalent inhibition of sulfatases by sulfamate warheads

The enzymatic removal of sulfate groups regulates processes ranging from steroid metabolism to carbohydrate degradation. Most sulfatases belong to the S1 family, whose members use a co-translationally installed formylglycine residue to hydrolyse sulfate esters. Arylsulfamates are potent covalent inhibitors of aryl and steroid sulfatases, including the clinical steroid sulfatase inhibitor Irosustat, yet the structure and stability of the inhibited complex remain unresolved. Arylsulfamates and carbohydrate sulfamates do not covalently inhibit many S1 carbohydrate sulfatases despite conservation of their sulfate-binding sites and formylglycine residue. Using enzyme kinetics, X-ray crystallography, molecular dynamics simulations and density functional theory calculations, we define the basis of these contrasting behaviours. High-resolution structures of the Pseudomonas aeruginosa arylsulfatase PaAtsA treated with two arylsulfamates reveal a long-lived tetrahedral, O-linked -hydroxysulfamate adduct attached to formylglycine. Molecular simulations show that replacing sulfate with sulfamate disrupts the favourable Ca2+-oxyanion interaction and alters ligand binding geometry. The permissive hydrophobic binding site of PaAtsA accommodates this rearrangement while retaining a trajectory compatible with nucleophilic attack. By contrast, in the Bacteroides thetaiotaomicron carbohydrate sulfatase BT16363S-Gal, sulfate-to-sulfamate substitution weakens binding and displaces the sulfamate from a reactive pose near the catalytic nucleophile due to a restrictive active site with conserved sugar binding. These findings define the structure and persistence of the arylsulfamate-derived covalent intermediate and explain why sulfamate warheads are tolerated by aryl sulfatases but not carbohydrate sulfatases.

biochemistry↗

bioRxiv2025

Nicotine biosynthesis completed by cryptic activating glucosylation

Nicotine is a neuroactive alkaloid produced by tobacco (Nicotiana tabacum) as a defense against herbivory, and an addictive stimulant that has been used by humans for millennia. Despite its significance, the core steps of its biosynthesis have remained elusive. Here, we demonstrate in vitro reconstruction of nicotine synthase, a four-enzyme stereoselective biocatalytic cascade that forms (S)-nicotine from nicotinic acid and N-methylpyrrolinium. This cascade includes two glucose-processing enzymes that participate in a cryptic activating glucosylation step. We also reconstruct this pathway in planta and present high resolution X-ray structures of the key oxidoreductases A622 and BBL bound to their substrate and product, respectively. This work establishes the complete biosynthetic pathway to nicotine, providing new gene targets for controlling alkaloid production in Nicotiana and unlocking enzymatic routes to pyridine alkaloids.

biochemistry↗

bioRxiv2025

The Androgen Receptor and MYC synergise to modulate the synthesis of Siglec-7 ligands in prostate cancer

Glyco-immune checkpoints have recently been shown to be critical mediators of immunotherapy resistance across multiple cancer types. In clinical trials, immunotherapeutic treatments for prostate cancer have failed to elicit durable clinical responses. PCa progression is driven by transcriptional networks regulated by key transcription factors including the androgen receptor (AR) and the oncogene MYC. How this crossover between hormone and oncogene-driven signalling pathways regulates tumour glyco-immune checkpoints remains unclear. Here, we show that O-glycans are the major substrates for sialylation in prostate cancer and that sialyltransferases that have preferences for O-glycans are differentially regulated by androgens. We show that supraphysiological levels of androgens produce distinct glycopeptide profiles in prostate cancer cells compared with cells exposed to physiological androgens. Additionally, we identify a direct and coordinated role for AR and MYC in regulating ST3Gal1 and the synthesis of Siglec-7 ligands in prostate cancer. Both transcription factors converge to repress ST3GAL1, thereby limiting the generation of Siglec-7 ligands. These findings highlight a context-dependent, cooperative relationship between the AR and MYC in shaping the tumour sialome, linking hormonal signalling and oncogenic transcription to Siglec biology. Our study highlights how cell-type specific differences in transcriptional networks has important downstream effects for immune modulating glycans and has tumour specific clinical implications.

cancer biology↗