Oliver Florey

Research Summary

Research in our lab focuses on a non-canonical autophagy pathway, associated with CASM (conjugation of ATG8 to single-membranes), and its role in lysosomal biology. We aim to understand the molecular mechanisms underlying the regulation and function of this pathway in cellular processes such as cell stress responses and infection.

Our work exploits a combination of molecular and cellular biology, state-of-the-art microscopy (long-term time-lapse imaging, spinning disk confocal and electron microscopy) and proteomics (mass spectrometry).

Existing projects aim to define the molecular mechanisms which underlie non-canonical autophagy, and exploring the potential to manipulate the pathway for therapeutic benefit.

Latest Publications

Subtractive CRISPR screen identifies the ATG16L1/vacuolar ATPase axis as required for non-canonical LC3 lipidation.
Ulferts R, Marcassa E, Timimi L, Lee LC, Daley A, Montaner B, Turner SD, Florey O, Baillie JK, Beale R

Although commonly associated with autophagosomes, LC3 can also be recruited to membranes by covalent lipidation in a variety of non-canonical contexts. These include responses to ionophores such as the M2 proton channel of influenza A virus. We report a subtractive CRISPR screen that identifies factors required for non-canonical LC3 lipidation. As well as the enzyme complexes directly responsible for LC3 lipidation in all contexts, we show the RALGAP complex is important for M2-induced, but not ionophore drug-induced, LC3 lipidation. In contrast, ATG4D is responsible for LC3 recycling in M2-induced and basal LC3 lipidation. Identification of a vacuolar ATPase subunit in the screen suggests a common mechanism for non-canonical LC3 recruitment. Influenza-induced and ionophore drug-induced LC3 lipidation lead to association of the vacuolar ATPase and ATG16L1 and can be antagonized by Salmonella SopF. LC3 recruitment to erroneously neutral compartments may therefore represent a response to damage caused by diverse invasive pathogens.

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Cell reports, 37, 4, 26 Oct 2021

PMID: 34706226

GABARAP sequesters the FLCN-FNIP tumor suppressor complex to couple autophagy with lysosomal biogenesis.
Goodwin JM, Walkup WG, Hooper K, Li T, Kishi-Itakura C, Ng A, Lehmberg T, Jha A, Kommineni S, Fletcher K, Garcia-Fortanet J, Fan Y, Tang Q, Wei M, Agrawal A, Budhe SR, Rouduri SR, Baird D, Saunders J, Kiselar J, Chance MR, Ballabio A, Appleton BA, Brumell JH, Florey O, Murphy LO

[Figure: see text].

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Science advances, 7, 40, Oct 2021

PMID: 34597140

A new flavor of cellular Atg8-family protein lipidation - alternative conjugation to phosphatidylserine during CASM.
Durgan J, Florey O

Atg8-family protein lipidation is the most commonly used marker for monitoring autophagy. During macroautophagy, Atg8-family proteins are specifically conjugated to phosphatidylethanolamine (PE) in forming, double-membrane autophagosomes. A distinct, non-canonical autophagy pathway also operates, characterized by the Conjugation of ATG8s to endolysosomal Single Membranes (CASM). In our new study, we show that CASM is associated with the alternative conjugation of Atg8-family proteins to phosphatidylserine (PS), and PE, in response to various cellular stimuli. We also discover differences in the regulation of conjugation to PE and PS by ATG4s, and altered dynamics between the two species. The identification of alternative Atg8-family protein PS lipidation opens up exciting new questions on the roles, regulation and biology of Atg8-family proteins during non-canonical autophagy.

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Autophagy, 1, 1, 12 Jul 2021

PMID: 34251968