Selective autophagy of mitochondria on a ubiquitin-endoplasmic reticulum platform

Correction: Developmental Cell, Volume 55, Issue 2 https://doi.org/10.1016/j.devcel.2020.10.002The dynamics and co-ordination between autophagy machinery and selective receptors during mitophagy are unknown. Also unknown is whether mitophagy depends on pre-existing membranes, or is triggered on the surface of damaged mitochondria. Using a ubiquitin-dependent mitophagy inducer, the lactone ivermectin, we have combined genetic and imaging experiments to address these questions. Ubiquitination of mitochondrial fragments is required earliest followed by autophosphorylation of TBK1. Next, early essential autophagy proteins FIP200 and ATG13 act at different steps whereas ULK1/2 are dispensable. Receptors act temporally and mechanistically upstream of ATG13 but downstream of FIP200. The VPS34 complex functions at the omegasome step. ATG13 and optineurin target mitochondria in a discontinuous oscillatory way suggesting multiple initiation events. Targeted ubiquitinated mitochondrial are cradled by endoplasmic reticulum strands even without functional autophagy machinery and mitophagy adaptors. We propose that damaged mitochondria are ubiquitinated and dynamically encased in ER strands providing platforms for formation of the mitophagosomes.Peer reviewe

Prostate MRI quality: clinical impact of the PI-QUAL score in prostate cancer diagnostic work-up.

OBJECTIVE: To assess the reproducibility and impact of prostate imaging quality (PI-QUAL) scores in a clinical cohort undergoing prostate multiparametric MRI. METHODS: PI-QUAL scores were independently recorded by three radiologists (two senior, one junior). Readers also recorded whether MRI was sufficient to rule-in/out cancer and if repeat imaging was required. Inter-reader agreement was assessed using Cohen's κ. PI-QUAL scores were further correlated to PI-RADS score, number of biopsy procedures, and need for repeat imaging. RESULTS: Image quality was sufficient (≥PI-QUAL-3) in 237/247 (96%) and optimal (≥PI-QUAL-4) in 206/247 (83%) of males undergoing 3T-MRI. Overall PI-QUAL scores showed moderate inter-reader agreement for senior (K = 0.51) and junior-senior readers (K = 0.47), with DCE showing highest agreement (K = 0.47). With PI-QUAL-5 studies, the negative MRI calls increased from 50 to 87% and indeterminate PI-RADS-3 rates decreased from 31.8. to 10.4% compared to lower quality PI-QUAL-3 studies. More patients with PI-QUAL scores 1-3 underwent biopsy for negative (47%) and indeterminate probability (100%) MRIs compared to PI-QUAL score 4-5 (30 and 75%, respectively). Ability to rule-in cancer increased with PI-QUAL score, from 50% at PI-QUAL 1-2 to 90% for PI-QUAL 4-5, with a similarly, but greater effect for ruling-out cancer and at a lower threshold, from 0% for scans of PI-QUAL 1-2 to 67.1% for PI-QUAL 4 and 100% for PI-QUAL-5. CONCLUSION: Higher PI-QUAL scores for image quality are associated with decreased uncertainty in MRI decision-making and improved efficiency of diagnostic pathway delivery. ADVANCES IN KNOWLEDGE: This study demonstrates moderate inter-reader agreement for PI-QUAL scoring and validates the score in a clinical setting, showing correlation of image quality to certainty of decision making and clinical outcomes of repeat imaging and biopsy of low-to-intermediate risk cases

Building arks for tRNA: Structure and function of the Arc1p family of non-catalytic tRNA-binding proteins

Following the intricate architecture of the eukaryotic cell, protein synthesis involves formation of many macromolecular assemblies, some of which are composed by tRNA-aminoacylation enzymes. Protein-protein and protein-tRNA interactions in these complexes can be facilitated by non-catalytic tRNA-binding proteins. This review focuses on the dissection of the molecular, structural and functional properties of a particular family of such proteins: yeast Arc1p and its homologues in pro-karyotes and higher eukaryotes. They represent paradigms of the strategies employed for the organization of sophisticated and dynamic nanostructures supporting spatio-temporal cellular organization. (C) 2010 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved

Phosphorylation of Phosphatidate Phosphatase Regulates Its Membrane Association and Physiological Functions in Saccharomyces cerevisiae: IDENTIFICATION OF SER602, THR723, AND SER744 AS THE SITES PHOSPHORYLATED BY CDC28 (CDK1)-ENCODED CYCLIN-DEPENDENT KINASE*

The Saccharomyces cerevisiae PAH1-encoded phosphatidate phosphatase (PAP) catalyzes the penultimate step in the synthesis of triacylglycerol and plays a role in the transcriptional regulation of phospholipid synthesis genes. PAP is phosphorylated at multiple Ser and Thr residues and is dephosphorylated for in vivo function by the Nem1p-Spo7p protein phosphatase complex localized in the nuclear/endoplasmic reticulum membrane. In this work, we characterized seven previously identified phosphorylation sites of PAP that are within the Ser/Thr-Pro motif. When expressed on a low copy plasmid, wild type PAP could not complement the pah1Δ mutant in the absence of the Nem1p-Spo7p complex. However, phosphorylation-deficient PAP (PAP-7A) containing alanine substitutions for the seven phosphorylation sites bypassed the requirement of the phosphatase complex and complemented the pah1Δ nem1Δ mutant phenotypes, such as temperature sensitivity, nuclear/endoplasmic reticulum membrane expansion, decreased triacylglycerol synthesis, and derepression of INO1 expression. Subcellular fractionation coupled with immunoblot analysis showed that PAP-7A was highly enriched in the membrane fraction. In fluorescence spectroscopy analysis, the PAP-7A showed tighter association with phospholipid vesicles than wild type PAP. Using site-directed mutagenesis of PAP, we identified Ser602, Thr723, and Ser744, which belong to the seven phosphorylation sites, as the sites phosphorylated by the CDC28 (CDK1)-encoded cyclin-dependent kinase. Compared with the dephosphorylation mimic of the seven phosphorylation sites, alanine substitution for Ser602, Thr723, and/or Ser744 had a partial effect on circumventing the requirement for the Nem1p-Spo7p complex