Autophagy reports最新文献

The KEAP1 (kelch like ECH associated protein 1)- NFE2L2/NRF2 (NFE2 like bZIP transcription factor 2) pathway is a major antioxidative stress pathway that contributes to cellular homeostasis. KEAP1 acts as a sensor and attenuates degradation of the transcription factor NRF2, which induces gene expression for a network of enzymes involved in the antioxidant response. When cells are exposed to various electrophiles and reactive oxidative species, they modify one or more selective cysteine residues in KEAP1, resulting in conformational changes that disable its NRF2-inhibitory function. In addition to this redox-dependent pathway, SQSTM1/p62 (sequestosome 1), which is a selective autophagy receptor for ubiquitinated proteins and a driver of liquid-liquid phase separation (LLPS) upon binding to ubiquitinated proteins, competitively inhibits the binding between KEAP1 and NRF2, thereby disabling the NRF2-repressive function of KEAP1. Our study showed that phase-separated SQSTM1/p62 bodies are phosphorylated by ULK1 (Unc-51 like autophagy activating kinase 1) and that KEAP1 is retained in the SQSTM1/p62 body, resulting in NRF2-activation in a redox-independent manner.

Autophagy has two distinct pathways, degradation and secretion. Autophagic degradation plays a pivotal role in cellular homeostasis by the formation of a double-membrane autophagosome in concert with numerous ATG (autophagy-related) proteins. However, the mechanism that mediates autophagic secretion is not fully understood. To explore how autophagic secretion is physiologically triggered and regulated in neurons, we investigated whether neuronal activity affected autophagic secretion by analyzing SNCA secretion in mouse primary cortical neurons and SH-SY5Y cells. In primary neurons, rapamycin promoted SNCA secretion, while the effect was canceled in primary neurons of Becn1 ^+/-deficient mice. Stimulating neuronal activity by glutamate promoted SNCA secretion, autophagic flux, and colocalization of SNCA with LC3 (microtubule-associated proteins 1 light chain 3). These effects were inhibited by the intracellular Ca²⁺ chelator BAPTA-AM. Additionally, glutamate-induced SNCA secretion was suppressed by Atg5 or Rab8a knockdown in SH-SY5Y cells, and mainly occurred in the fashion associated with extracellular vesicles in primary neurons. These results suggest that neuronal activity triggers autophagic secretion for releasing SNCA via an increase in intracellular Ca²⁺ concentration.

Glycogen-autophagy ('glycophagy') is a selective autophagy process involved in delivering glycogen to the lysosome for bulk degradation. Glycophagy protein intermediaries include STBD1 as a glycogen tagging receptor, delivering the glycogen cargo into the forming phagosome by partnering with the Atg8 homolog, GABARAPL1. Glycophagy is emerging as a key process of energy metabolism and development of reliable tools for assessment of glycophagy activity is an important priority. Here we show that antibodies raised against the N-terminus of the GABARAPL1 protein (but not the full-length protein) detected a specific endogenous GABARAPL1 immunoblot band at 18kDa. A stable GFP-GABARAPL1 cardiac cell line was used to quantify GABARAPL1 lysosomal flux via measurement of GFP puncta in response to lysosomal inhibition with bafilomycin. Endogenous glycophagy flux was quantified in primary rat ventricular myocytes by the extent of glycogen accumulation with bafilomycin combined with chloroquine treatment (no effect observed with bafilomycin or chloroquine alone). In wild-type isolated mouse hearts, bafilomycin alone and bafilomycin combined with chloroquine (but not chloroquine alone) elicited a significant increase in glycogen content signifying basal glycophagy flux. Collectively, these methodologies provide a comprehensive toolbox for tracking cardiac glycophagy activity to advance research into the role of glycophagy in health and disease.

Avian metapneumovirus subgroup C (aMPV/C) is an emerging pathogen that causes acute respiratory infection in chickens and turkeys. Sequestosome 1 (SQSTM1), a selective autophagy receptor, regulates cellular activity or viral replication by recognizing ubiquitinated substrates. Here, we found that SQSTM1 expression inhibits aMPV/C replication through selective autophagy. In particular, SQSTM1 interacts with the aMPV/C M2-2 protein via its PB1 domain, and by recognizing a ubiquitinated lysine at position 67 of viral M2-2 protein. This recognition leads to the autophagic degradation of the aMPV/C M2-2 protein, suppressing viral replication.

Excessive exposure to sunlight, especially to ultraviolet B (UVB), results in DNA damage and a cutaneous inflammatory reaction commonly known as sunburn, which increases skin cancer risks. UVB-induced inflammasome activation in epidermal keratinocytes mediates the cutaneous inflammatory response, but the intracellular machinery that maintains skin homeostasis by suppressing UVB-induced inflammasome activation is unclear. Here, we summarize our recent work on the protective role of alternative autophagy against UVB-induced NLRP3 (NLR family pyrin domain containing 3) inflammasome activation in human keratinocytes. We found that UVB radiation induces ATG5/ATG7-independent alternative (noncanonical) autophagy, which leads to suppression of NLRP3 inflammasome activation through the clearance of damaged mitochondria in UVB-irradiated keratinocytes. Our findings indicate that ATG5/ATG7-independent alternative autophagy, rather than conventional autophagy, may play a key role in mitigating inflammatory responses, and restoring skin homeostasis after UV radiation.

Beiging of adipocytes is characteristic of a higher number of mitochondria, the central hub of metabolism in the cell. However, studies show that beiging can improve metabolic health or cause metabolic disorders. Here we discuss a liver-fat crosstalk for iron flux associated with healthy beiging of adipocytes. Deletion of the transcription factor FoxO1 in adipocytes (adO1KO mice) induces a higher iron flux from the liver to white adipose tissue, concurrent with augmented mitochondrial biogenesis that increases iron demands. In addition, adO1KO mice adopt an alternate mechanism to sustain mitophagy, which enhances mitochondrial quality control, thereby improving mitochondrial respiratory capacity and metabolic health. However, the liver-fat crosstalk is not detectable in adipose Atg7 knockout (ad7KO) mice, which undergo beiging of adipocytes but have metabolic dysregulation. Autophagic clearance of mitochondria is blocked in ad7KO mice, which accumulates dysfunctional mitochondria and elevates mitochondrial content but lowers mitochondrial respiratory capacity. Mitochondrial biogenesis is comparable in the control and ad7KO mice, and the iron influx into adipocytes and iron efflux from the liver remain unchanged. Therefore, activation of the liver-fat crosstalk is critical for mitochondrial quality control that underlies healthy beiging of adipocytes.

Most autophagy-related (ATG) genes have been identified and characterized through studies using the budding yeast Saccharomyces cerevisiae. However, there are no studies that comprehensively compare the contribution of each ATG gene to non-selective bulk autophagy and various types of selective autophagy, including the cytoplasm-to-vacuole targeting (Cvt) pathway, mitophagy, reticulophagy, and pexophagy. Our recent study quantified these types of autophagy in all atg mutants and showed that (i) autophagy is not completely impaired in specific deletants such as lacking the components of the two ubiquitin conjugation-like (UBL) systems, traditionally regarded as essential for autophagy in yeast, and (ii) residual autophagic activity is especially prominent in mutants of the Atg8 UBL system, which display small autophagic body-like vesicles at a low frequency. Alongside these findings, our comprehensive analysis suggested a link between mitophagy and pexophagy, and a differential contribution of Atg proteins to cargo specificity. We discuss how our datasets are useful for future autophagy research. Abbreviations: Ape1: aminopeptidase I; ATG: autophagy related; Cvt: cytoplasm-to-vacuole targeting; ER: endoplasmic reticulum; ESCRT: endosomal sorting complex required for transport; GFP: green fluorescent protein; PI3K: phosphatidylinositol 3-kinase; UBL: ubiquitin conjugation-like.