The soluble reticulophagy receptor CALCOCO1 is also a Golgiphagy receptor.

Cellular stress response mechanisms typically increase organellar quantity and volume. To restore cellular homeostasis and organellar integrity, the surplus organelles are cleared by macroautophagy/autophagy, an intracellular process that shuttles cytoplasmic material to the lysosomes for degradation. The degradation is mediated by autophagy receptors that selectively link the degradable cargo to the autophagy machinery. Studies have identified receptors for the degradation of mitochondria, endoplasmic reticulum, lysosomes, and peroxisomes. The autophagic degradation of the Golgi, named Golgiphagy, however, has remained undefined. The Golgi is essential for the processing, sorting and trafficking of proteins and lipids in the secretory pathway. In a recent study, we identified CALCOCO1 as a Golgiphagy receptor in response to nutrient deprivation. CALCOCO1 interacts with Golgi membranes by binding to cytoplasmic Ankyrin repeat (AR) domains of Golgi resident ZDHHC17 and ZDHHC13 palmitoyltransferases (PATs) via a defined zDHHC-AR-binding motif (zDABM) to recruit autophagy machinery. Lack of CALCOCO1 in cells causes an impaired Golgiphagy and expansion of the Golgi.

SAMM50 acts with p62 in piecemeal basal- and OXPHOS-induced mitophagy of SAM and MICOS components.

Mitophagy is the degradation of surplus or damaged mitochondria by autophagy. In addition to programmed and stress-induced mitophagy, basal mitophagy processes exert organelle quality control. Here, we show that the sorting and assembly machinery (SAM) complex protein SAMM50 interacts directly with ATG8 family proteins and p62/SQSTM1 to act as a receptor for a basal mitophagy of components of the SAM and mitochondrial contact site and cristae organizing system (MICOS) complexes. SAMM50 regulates mitochondrial architecture by controlling formation and assembly of the MICOS complex decisive for normal cristae morphology and exerts quality control of MICOS components. To this end, SAMM50 recruits ATG8 family proteins through a canonical LIR motif and interacts with p62/SQSTM1 to mediate basal mitophagy of SAM and MICOS components. Upon metabolic switch to oxidative phosphorylation, SAMM50 and p62 cooperate to mediate efficient mitophagy.

Regulation of Golgi turnover by CALCOCO1-mediated selective autophagy.

The Golgi complex is essential for the processing, sorting, and trafficking of newly synthesized proteins and lipids. Golgi turnover is regulated to meet different cellular physiological demands. The role of autophagy in the turnover of Golgi, however, has not been clarified. Here we show that CALCOCO1 binds the Golgi-resident palmitoyltransferase ZDHHC17 to facilitate Golgi degradation by autophagy during starvation. Depletion of CALCOCO1 in cells causes expansion of the Golgi and accumulation of its structural and membrane proteins. ZDHHC17 itself is degraded by autophagy together with other Golgi membrane proteins such as TMEM165. Taken together, our data suggest a model in which CALCOCO1 mediates selective Golgiphagy to control Golgi size and morphology in eukaryotic cells via its interaction with ZDHHC17.

Occurrence of Dirofilaria immitis in Stray Dogs from Nepal.

BACKGROUND: Nematodes of the genus Dirofilaria are widespread vector-borne helminths (VBH) of veterinary relevance. Dirofilariosis caused by D. immitis is frequently diagnosed in dogs often showing a severe clinical condition known as heartworm disease. Assessing the distribution pattern of canine dirofilariosis is pivotal to undertake appropriate control measures and define the risk of infection in animals and humans. This study provides original data on the occurrence of D. immitis in naive stray dogs from Nepal. METHODS: An epidemiological study was performed on stray dogs from Siddharthanagar, Lumbini region, Nepal. Alive animals were blood sampled and examined for microfilariae using direct microscopy, buffy coat centrifugation and modified Knott's method. In addition, hematobiochemical parameters, including packed cell volume (PCV), alanine aminotransferase (SGPT), aspartate aminotransferase (SGOT), alkaline phosphate (ALP), creatinine, blood urea nitrogen (BUN), urea and total protein were analysed, along with a clinical scoring (body temperature and respiratory rate). During the observation period, data on autopsies performed on stray dogs showing canine heartworm disease were also included. RESULTS: The overall prevalence of D. immitis in both alive and necropsied dogs was 22.49% (38/169). Out of 150 alive dogs, 29 (19.3%) were positive for D. immitis microfilariae. The prevalence of the infection varied according to the technique used, ranging from 16.0 to 19.3% based on direct blood smear microscopy and modified Knott's technique, respectively. The infection rate was significantly higher (p < 0.05) in dogs aging more than 3 years. A positive correlation was observed between SGPT and SGOT and the presence of microfilariae (p < 0.05). Among the 19 dead stray dogs, 9 animals showed adult D. immitis nematodes. CONCLUSION: Results of this survey account for the presence of D. immitis in stray dogs from Nepal, thus supporting the existence of a local transmission cycle of diroflariosis in the region. Further investigations are warranted to accurately define the prevalence of the infection in the canine population and to undertake appropriate control measures to reduce the burden on the infection in dogs and, accordingly, the risk of transmission to humans.

Susceptibility to SARS, MERS, and COVID-19 from animal health perspective.

Viruses are having great time as they seem to have bogged humans down. Severe acute respiratory syndrome (SARS), Middle East respiratory syndrome (MERS), and novel coronavirus (COVID-19) are the three major coronaviruses of present-day global human and animal health concern. COVID-19 caused by SARS-CoV-2 is identified as the newest disease, presumably of bat origin. Different theories on the evolution of viruses are in circulation, yet there is no denying the fact that the animal source is the skeleton. The whole world is witnessing the terror of the COVID-19 pandemic that is following the same path of SARS and MERS, and seems to be more severe. In addition to humans, several species of animals are reported to have been infected with these life-threatening viruses. The possible routes of transmission and their zoonotic potentialities are the subjects of intense research. This review article aims to overview the link of all these three deadly coronaviruses among animals along with their phylogenic evolution and cross-species transmission. This is essential since animals as pets or food are said to pose some risk, and their better understanding is a must in order to prepare a possible plan for future havoc in both human and animal health. Although COVID-19 is causing a human health hazard globally, its reporting in animals are limited compared to SARS and MERS. Non-human primates and carnivores are most susceptible to SARS-coronavirus and SARS-CoV-2, respectively, whereas the dromedary camel is susceptible to MERS-coronavirus. Phylogenetically, the trio viruses are reported to have originated from bats and have special capacity to undergo mutation and genomic recombination in order to infect humans through its reservoir or replication host. However, it is difficult to analyze how the genomic pattern of coronaviruses occurs. Thus, increased possibility of new virus-variants infecting humans and animals in the upcoming days seems to be the biggest challenge for the future of the world. One health approach is portrayed as our best way ahead, and understanding the animal dimension will go a long way in formulating such preparedness plans.

CALCOCO1 is a soluble reticulophagy receptor.

The endoplasmic reticulum (ER) is the largest membrane-bound organelle in eukaryotic cells and plays critical roles in diverse processes in metabolism, signaling and intracellular organization. In response to stress stimuli such as nutrient deprivation, accumulation of misfolded proteins or exposure to chemicals, the ER increases in size through upregulated synthesis of its components to counteract the stress. To restore physiological size, the excess ER components are continuously dismantled and degraded by reticulophagy, a form of autophagy that targets, via adaptor molecules called reticulophagy receptors, specific ER portions to the lysosome for degradation. Previous studies have identified several ER resident proteins as reticulophagy receptors. In a recent study, we identified CALCOCO1 as a soluble reticulophagy receptor for the degradation of tubular ER in response to proteotoxic and starvation-induced stress. On the ER membrane, CALCOCO1 interacts with VAPA and VAPB via a FFAT-like motif and recruits autophagy machinery by binding directly to Atg8-family proteins via LIR and UDS interacting region (UIR) motifs acting co-dependently. Depletion of CALCOCO1 in cultured cells led to an impaired ER degradation during stress.

CALCOCO1 acts with VAMP-associated proteins to mediate ER-phagy.

The endoplasmic reticulum (ER) plays important roles in protein synthesis and folding, and calcium storage. The volume of the ER and expression of its resident proteins are increased in response to nutrient stress. ER-phagy, a selective form of autophagy, is involved in the degradation of the excess components of the ER to restore homeostasis. Six ER-resident proteins have been identified as ER-phagy receptors so far. In this study, we have identified CALCOCO1 as a novel ER-phagy receptor for the degradation of the tubular ER in response to proteotoxic and nutrient stress. CALCOCO1 is a homomeric protein that binds directly to ATG8 proteins via LIR- and UDS-interacting region (UIR) motifs acting co-dependently. CALCOCO1-mediated ER-phagy requires interaction with VAMP-associated proteins VAPA and VAPB on the ER membranes via a conserved FFAT-like motif. Depletion of CALCOCO1 causes expansion of the ER and inefficient basal autophagy flux. Unlike the other ER-phagy receptors, CALCOCO1 is peripherally associated with the ER. Therefore, we define CALCOCO1 as a soluble ER-phagy receptor.

Structural basis of p62/SQSTM1 helical filaments and their role in cellular cargo uptake.

p62/SQSTM1 is an autophagy receptor and signaling adaptor with an N-terminal PB1 domain that forms the scaffold of phase-separated p62 bodies in the cell. The molecular determinants that govern PB1 domain filament formation in vitro remain to be determined and the role of p62 filaments inside the cell is currently unclear. We here determine four high-resolution cryo-EM structures of different human and Arabidopsis PB1 domain assemblies and observed a filamentous ultrastructure of p62/SQSTM1 bodies using correlative cellular EM. We show that oligomerization or polymerization, driven by a double arginine finger in the PB1 domain, is a general requirement for lysosomal targeting of p62. Furthermore, the filamentous assembly state of p62 is required for autophagosomal processing of the p62-specific cargo KEAP1. Our results show that using such mechanisms, p62 filaments can be critical for cargo uptake in autophagy and are an integral part of phase-separated p62 bodies.

NIMA-related kinase 9-mediated phosphorylation of the microtubule-associated LC3B protein at Thr-50 suppresses selective autophagy of p62/sequestosome 1.

Human ATG8 family proteins (ATG8s) are active in all steps of the macroautophagy pathway, and their lipidation is essential for autophagosome formation. Lipidated ATG8s anchored to the outer surface of the phagophore serve as scaffolds for binding of other core autophagy proteins and various effector proteins involved in trafficking or fusion events, whereas those at the inner surface are needed for assembly of selective autophagy substrates. Their scaffolding role depends on specific interactions between the LC3-interacting region (LIR) docking site (LDS) in ATG8s and LIR motifs in various interaction partners. LC3B is phosphorylated at Thr-50 within the LDS by serine/threonine kinase (STK) 3 and STK4. Here, we identified LIR motifs in STK3 and atypical protein kinase Cζ (PKCζ) and never in mitosis A (NIMA)-related kinase 9 (NEK9). All three kinases phosphorylated LC3B Thr-50 in vitro A phospho-mimicking substitution of Thr-50 impaired binding of several LIR-containing proteins, such as ATG4B, FYVE, and coiled-coil domain-containing 1 (FYCO1), and autophagy cargo receptors p62/sequestosome 1 (SQSTM1) and neighbor of BRCA1 gene (NBR1). NEK9 knockdown or knockout enhanced degradation of the autophagy receptor and substrate p62. Of note, the suppression of p62 degradation was mediated by NEK9-mediated phosphorylation of LC3B Thr-50. Consistently, reconstitution of LC3B-KO cells with the phospho-mimicking T50E variant inhibited autophagic p62 degradation. PKCζ knockdown did not affect autophagic p62 degradation, whereas STK3/4 knockouts inhibited autophagic p62 degradation independently of LC3B Thr-50 phosphorylation. Our findings suggest that NEK9 suppresses LC3B-mediated autophagy of p62 by phosphorylating Thr-50 within the LDS of LC3B.

ATG4B contains a C-terminal LIR motif important for binding and efficient cleavage of mammalian orthologs of yeast Atg8.

The cysteine protease ATG4B cleaves off one or more C-terminal residues of the inactive proform of proteins of the ortholog and paralog LC3 and GABARAP subfamilies of yeast Atg8 to expose a C-terminal glycine that is conjugated to phosphatidylethanolamine during autophagosome formation. We show that ATG4B contains a C-terminal LC3-interacting region (LIR) motif important for efficient binding to and cleavage of LC3 and GABARAP proteins. We solved the crystal structures of the GABARAPL1-ATG4B C-terminal LIR complex. Analyses of the structures and in vitro binding assays, using specific point mutants, clearly showed that the ATG4B LIR binds via electrostatic-, aromatic HP1 and hydrophobic HP2 pocket interactions. Both these interactions and the catalytic site-substrate interaction contribute to binding between LC3s or GABARAPs and ATG4B. We also reveal an unexpected role for ATG4B in stabilizing the unlipidated forms of GABARAP and GABARAPL1. In mouse embryonic fibroblast (MEF) atg4b knockout cells, GABARAP and GABARAPL1 were unstable and degraded by the proteasome. Strikingly, the LIR motif of ATG4B was required for stabilization of the unlipidated forms of GABARAP and GABARAPL1 in cells.

Measles outbreak in a vaccinated population in Dhankutta.

A measles outbreak occurred in Dhankutta town in year 2056 Falgun to 2057 Baishak (3 months). A total of 130 cases were registered at the Dhankutta District Hospital. A large proportion of the cases had severe disease with complications and were admitted at the hospital. None of the patients registered at the hospital died. Interestingly, 50.0% of cases were in higher age group (10 years and above) which is an unusual trend in developing countries. All the cases gave history of measles vaccination, which points towards the need for investigation of vaccine efficacy and cold chain efficiency.