Non-canonical roles of ATG8 for TFEB activation.

Autophagy is an evolutionally conserved cytoplasmic degradation pathway in which the double membrane structure, autophagosome sequesters cytoplasmic material and delivers them to lysosomes for degradation. Many autophagy related (ATG) proteins participate in the regulation of the several steps of autophagic process. Among ATGs, ubiquitin-like protein, ATG8 plays a pivotal role in autophagy. ATG8 is directly conjugated on lipid in autophagosome membrane upon induction of autophagy thus providing a good marker to monitor and analyze autophagy process. However, recent discoveries suggest that ATG8 has autophagy independent non-canonical functions and ATG8 positive structures are not always autophagosomes. This review briefly overviews canonical and non-canonical roles of ATG8 and introduce novel function of ATG8 to activate Transcriptional Factor EB(TFEB), a master transcription factor of autophagy and lysosome function during lysosomal damage.

De novo design of a nanopore for single-molecule detection that incorporates a ß-hairpin peptide.

The amino-acid sequence of a protein encodes information on its three-dimensional structure and specific functionality. De novo design has emerged as a method to manipulate the primary structure for the development of artificial proteins and peptides with desired functionality. This paper describes the de novo design of a pore-forming peptide, named SV28, that has a ß-hairpin structure and assembles to form a stable nanopore in a bilayer lipid membrane. This large synthetic nanopore is an entirely artificial device for practical applications. The peptide forms multidispersely sized nanopore structures ranging from 1.7 to 6.3 nm in diameter and can detect DNAs. To form a monodispersely sized nanopore, we redesigned the SV28 by introducing a glycine-kink mutation. The resulting redesigned peptide forms a monodisperse pore with a diameter of 1.7 nm leading to detection of a single polypeptide chain. Such de novo design of a ß-hairpin peptide has the potential to create artificial nanopores, which can be size adjusted to a target molecule.

Autophagy-independent function of lipidated LC3 essential for TFEB activation during the lysosomal damage responses.

Lysosomes are digestive organelles in cells containing many hydrolases, and also serve as a signaling hub to integrate intracellular and extracellular inputs; therefore, the integrity of lysosomes is critical for cellular homeostasis. Many agents and conditions can damage lysosomal membranes, which lead to leakage of lysosomal acidic contents into the cytosol thus becoming harmful for cells. Accordingly, cells have developed several defense systems to cope with damaged lysosomes, but underlying mechanisms of each system and their cross-talks are unclear. In our recent study, we found that a master transcription factor regulating autophagy and lysosomal biogenesis, TFEB (transcription factor EB) is activated during lysosomal damage, and this activation depends on an autophagy-independent function of lipidated LC3, which localizes on lysosomes. We further showed that this regulatory mechanism is essential to prevent the progression of the crystal nephropathy that accompanies lysosomal damage.

LC3 lipidation is essential for TFEB activation during the lysosomal damage response to kidney injury.

Sensing and clearance of dysfunctional lysosomes is critical for cellular homeostasis. Here we show that transcription factor EB (TFEB)-a master transcriptional regulator of lysosomal biogenesis and autophagy-is activated during the lysosomal damage response, and its activation is dependent on the function of the ATG conjugation system, which mediates LC3 lipidation. In addition, lysosomal damage triggers LC3 recruitment on lysosomes, where lipidated LC3 interacts with the lysosomal calcium channel TRPML1, facilitating calcium efflux essential for TFEB activation. Furthermore, we demonstrate the presence and importance of this TFEB activation mechanism in kidneys in a mouse model of oxalate nephropathy accompanying lysosomal damage. A proximal tubule-specific TFEB-knockout mouse exhibited progression of kidney injury induced by oxalate crystals. Together, our results reveal unexpected mechanisms of TFEB activation by LC3 lipidation and their physiological relevance during the lysosomal damage response.