Biology subjects

Fregno, I.

Publications and source records attributed to Fregno, I..

5 recordsLinked to original sources

bioRxiv2026

The role of N-glycans and their processing in ER-to-lysosome-associated degradation of disease-causing mutant Neuroserpin

Most proteins synthesized in the endoplasmic reticulum (ER) are covalently modified upon addition of pre-assembled oligosaccharides to side chains of asparagine (N) residues. Processing of N-linked oligosaccharides by ER-resident glucosidases, mannosidases and glucosyltransferases determines the fate of the associated polypeptides. Terminally glucose residues are removed from N-glycans to hamper engagement of ER-resident glucose-binding chaperones and promote secretion of native polypeptides. Mannose residues are removed to target terminally misfolded proteins for dislocation across the ER membrane and clearance by the cytoplasmic ubiquitin proteasome system (ER-associated degradation, ERAD). Recent evidence highlights the role of persistent N-glycan glucosylation as a signal that promotes segregation of misfolded proteins in ER subdomains that are eventually delivered to endolysosomal compartments for ER-to-Lysosome-Associated Degradation (ERLAD). Here we show that the polymerization-prone Portland variant of Neuroserpin (NS_PL) associated with familial encephalopathy with NS inclusion bodies (FENIB) is a client of the ERLAD machinery. Its lysosomal clearance relies on the LC3-dependent delivery branch of ERLAD involving the lectin chaperone Calnexin (CNX), the ERphagy receptor FAM134B and the SNARE protein Syntaxin17 (STX17), which is engaged upon persistent glucosylation of the NS_PL oligosaccharide linked at the asparagine residue at position 321.

cell biology↗

bioRxiv2025

The involvement of the ER-phagy receptor FAM134B in membrane contact sites between ER and endolysosomes promotes ERLAD

Membrane contact sites (MCS) between organelles maintain the proximity required for controlled exchange of small molecules and ions yet preventing fusion events that would compromise organelles identity and integrity. Here, by investigating the intracellular fate of the disease-causing Z-variant of alpha1 antitrypsin (ATZ), we report on a novel function of MCS between the endoplasmic reticulum (ER) and RAB7/LAMP1-positive endolysosomes in ER-to-lysosome-associated degradation (ERLAD). For this function, the VAPA:ORP1L:RAB7 multi-protein complex forming MCS between the ER and endolysosomes engages, in an ERLAD client-driven manner, the misfolded protein segregation complex formed by the lectin chaperone Calnexin (CNX), the ER-phagy receptor FAM134B and the ubiquitin-like protein LC3. Generation of this supramolecular complex facilitates the membrane fusion events regulated by the SNARE proteins STX17 and VAMP8 that ensure efficient delivery of ATZ polymers from their site of generation, the ER, to the site of their intracellular clearance, the degradative RAB7/LAMP1-positive endolysosomes.

cell biology↗

bioRxiv2024

A novel class of allosteric glucosylceramidase beta 1 correctors that reduce cellular stress and enhance lysosomal function

Mutations in glucosylceramidase beta 1 (GCase) disrupt the proteins conformational maturation in the endoplasmic reticulum (ER) and hinder its transport to the lysosome. The intralysosomal accumulation of glucocerebrosides, which are substrates of the GCase enzyme, impairs lysosomal function and is linked to Gaucher disease (GD). GCase mutations also increase the risk of Parkinsons disease (PD) and Dementia with Lewy Bodies. We used Site-directed Enzyme Enhancement Therapy (SEE-Tx(R)) technology to design two structurally targeted allosteric regulators (STARs) of GCase. Administration of GT-02287 and GT-02329 to cultured GD patient-derived primary human fibroblasts enhances folding and protects the two most common disease-causing GCase variants, GCaseAsn370Ser and GCaseLeu444Pro, from proteasomal degradation. Mechanistically, these treatments facilitate the lysosomal delivery of enzymatically active forms of mutant GCase, leading to improved lysosomal function and reduced cellular stress in GD patient-derived fibroblasts. The findings suggest that the allosteric pharmacologic regulators GT-02287 and GT-02329 hold promise for further development as potential therapeutic agents for GCase-related disorders, including GD, PD and Dementia with Lewy Bodies.

cell biology↗

bioRxiv2024

Pharmacological GCase Activity Enhancement Inhibits Tau Accumulation

A slow decline in the autophagy-lysosomal pathway is a hallmark of the normal aging brain. Yet, an acceleration of this cellular function may propel neurodegenerative events. In fact, mutations in genes associated with the autophagy-lysosomal pathway can lead to Parkinsons disease. Also, amyloidogenic protein deposition is observed in lysosomal storage disorders, which are caused by genetic mutations representing risk factors for Parkinsons disease. For example, Gauchers disease GBA1 mutations leading to defects in lysosomal sphingolipid metabolism cause -synuclein accumulation. We observed that increased lysosomal Tau accumulation is found in human dermal fibroblasts engineered for inducible Tau expression. Inhibition of the GBA1 product GCase augmented Tau-dependent lysosomal stress and Tau accumulation. Here, we show increased Tau seed-induced Tau accumulation in Gauchers fibroblasts carrying GBA1 mutations when compared to normal fibroblasts. Pharmacological enhancement of GCase reversed this effect, notably, also in normal fibroblasts. This suggests that boosting GCase activity may represent a therapeutic strategy to slow down aging-dependent lysosomal deficits and brain protein deposition.

cell biology↗

bioRxiv2023

FAM134B regulates ER-to-lysosome-associated degradation of misfolded proteins upon pharmacologic or genetic inactivation of ER-associated degradation

About 40% of the eukaryotic cells proteome is synthesized and assembled in the endoplasmic reticulum (ER). Native proteins are transported to their intra- or extra-cellular site of activity. Folding-defective polypeptides are dislocated across the ER membrane into the cytoplasm, poly-ubiquitylated and degraded by 26S proteasomes (ER-associated degradation, ERAD). Large misfolded proteins like mutant forms of collagen or aggregation-prone mutant forms of alpha1 antitrypsin cannot be dislocated across the ER membrane for ERAD. Rather, they are segregated in ER subdomains that vesiculate and deliver their cargo to endolysosomal compartments for clearance by ER-to-lysosome-associated degradation (ERLAD). Here, we show the lysosomal delivery of a canonical ERAD substrate upon pharmacologic and genetic inhibition of the ERAD pathways. This highlights the surrogate intervention of ERLAD to remove defective gene products upon dysfunctional ERAD.

cell biology↗