Fluorescence Methods to Measure Pexophagy.

We outline our approach for studying the selective autophagy of peroxisomes (pexophagy), using fluorescence microscopy in tissue cell culture models. Ratiometric reporters, which specifically localize to peroxisomes, allow a quantitative assessment of pexophagy in fixed and live cells, as well as whole organisms. We discuss chemical and physiological inducers of pexophagy and any overlap with the induction of mitophagy.

Quantifying neutralising antibody responses against SARS-CoV-2 in dried blood spots (DBS) and paired sera.

The ongoing SARS-CoV-2 pandemic was initially managed by non-pharmaceutical interventions such as diagnostic testing, isolation of positive cases, physical distancing and lockdowns. The advent of vaccines has provided crucial protection against SARS-CoV-2. Neutralising antibody (nAb) responses are a key correlate of protection, and therefore measuring nAb responses is essential for monitoring vaccine efficacy.

Segregation of pathways leading to pexophagy.

Peroxisomes are organelles with key roles in metabolism including long-chain fatty acid production. Their metabolic functions overlap and interconnect with those of mitochondria, with which they share an overlapping but distinct proteome. Both organelles are degraded by selective autophagy processes termed pexophagy and mitophagy.

Benchmarking a highly selective USP30 inhibitor for enhancement of mitophagy and pexophagy.

The deubiquitylase USP30 is an actionable target considered for treatment of conditions associated with defects in the PINK1-PRKN pathway leading to mitophagy. We provide a detailed cell biological characterization of a benzosulphonamide molecule, compound 39, that has previously been reported to inhibit USP30 in an in vitro enzymatic assay. The current compound offers increased selectivity over previously described inhibitors.

USP30 sets a trigger threshold for PINK1-PARKIN amplification of mitochondrial ubiquitylation.

The mitochondrial deubiquitylase USP30 negatively regulates the selective autophagy of damaged mitochondria. We present the characterisation of an N-cyano pyrrolidine compound, FT3967385, with high selectivity for USP30. We demonstrate that ubiquitylation of TOM20, a component of the outer mitochondrial membrane import machinery, represents a robust biomarker for both USP30 loss and inhibition.