Biology subjects

Tettamanti, M. G.

Publications and source records attributed to Tettamanti, M. G..

3 recordsLinked to original sources

bioRxiv2024

TORC2-regulated sterol redistribution mediates recovery from membrane perturbation by small amphipathic molecules.

To maintain plasma membrane (PM) integrity, cells need to acutely regulate PM lipid composition. The Target Of Rapamycin (TOR) complex 2 is a protein kinase that acts as a central regulator of PM homeostasis, but the mechanisms by which it monitors and reacts to membrane stresses are poorly understood. To address this knowledge gap, we characterized a family of amphiphilic molecules that physically perturb PM organization and in doing so inhibit TORC2 in yeast and mammalian cells. Using fluorescent lipid associated reporters in budding yeast, we show that these small molecules first cause a transient increase in the amount of biochemically accessible ergosterol at the PM. Contemporaneous TORC2 inhibition stimulates a rapid removal of accessible ergosterol from the PM by the PM-ER sterol transporters Lam2 and Lam4, necessary for TORC2 reactivation. Thus, we show that TORC2 acts in a feedback loop to control active sterol levels at the PM and introduce sterols as possible TORC2 signalling modulators.

cell biology↗

bioRxiv2024

The Dsc ubiquitin ligase complex identifies transmembrane degrons to degrade orphaned proteins at the Golgi

The Golgi apparatus is essential for protein sorting, yet its quality control mechanisms are poorly understood. Here we show that the Dsc ubiquitin ligase complex, particularly the rhomboid pseudo-protease subunit, Dsc2, assesses the hydrophobic length of -helical transmembrane domains (TMDs) at the Golgi. Thereby the Dsc complex interacts with orphaned ER and Golgi proteins that have shorter TMDs and ubiquitinates them for targeted degradation. Some Dsc substrates will be K63 polyubiquitinated for ESCRT dependent vacuolar degradation or K48 polyubiquitinated for endosome and Golgi associated proteasomal degradation (EGAD). Other Dsc substrates are exclusively extracted by Cdc48 for EGAD. The accumulation of Dsc substrates entails a specific increase in glycerophospholipids with shorter and asymmetric fatty acyl chains. Hence, the Dsc complex mediates the selective degradation of orphaned proteins at the sorting center of cells, which prevents their spreading across other organelles and thus preserves cellular membrane protein and lipid composition.

cell biology↗

bioRxiv2023

CryoEM architecture of a native stretch-sensitive membrane microdomain

Biological membranes are partitioned into functional zones containing specific lipids and proteins, termed membrane microdomains. Their composition and organization remain controversial owing to a paucity of techniques that can visualize lipids in situ without disrupting their native behavior1,2. The yeast eisosome, a membrane compartment scaffolded by the BAR-domain proteins Pil1 and Lsp1, senses and responds to mechanical stress by flattening and releasing sequestered factors3-7. Here, we isolated native eisosomes as helical filaments of Pil1/Lsp1 lattice bound to plasma membrane lipids and solved their structures by helical reconstruction. We observe remarkable organization within the lipid bilayer density from which we could assign headgroups of PI(4,5)P2 and phosphatidylserine bound to Pil1/Lsp1 and a pattern of membrane voids, signatures of sterols, beneath an amphipathic helix. We verified these assignments using in vitro reconstitutions and molecular dynamics simulations. 3D variability analysis of the native eisosomes revealed a dynamic stretching of the Pil1/Lsp1 lattice that affects functionally important lipid sequestration, supporting a mechanism in which membrane stretching liberates lipids otherwise anchored by the Pil1/Lsp1 coat. Our results provide mechanistic insight into how eisosome BAR-domain proteins create a mechanosensitive membrane microdomain and, more globally, resolve long-standing controversies about the architecture and nature of lipid microdomains.

biophysics↗