Research in autophagy continues to accelerate,(1) and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.(2,3) There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to verify an autophagic response.

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Protein folding in the endoplasmic reticulum (ER) is monitored by ER quality control (ERQC) mechanisms. Proteins that pass ERQC criteria traffic to their final destinations through the secretory pathway, whereas non-native and unassembled subunits of multimeric proteins are degraded by the ER-associated degradation (ERAD) pathway. During ERAD, molecular chaperones and associated factors recognize and target substrates for retrotranslocation to the cytoplasm, where they are degraded by the ubiquitin-proteasome machinery. The discovery of diseases that are associated with • Endoplasmic reticulum (ER)-associated degradation (ERAD) is a secretory protein quality control process that results in the removal of aberrant proteins from the ER.• ERAD substrates are selected by molecular chaperones that identify proteins that might be unable to fold, that fold slowly or contain a misfolded domain, or those that lack specific protein partners.• Nearly all ERAD substrates are modified with ubiquitin, a 76 amino-acid peptide that helps target proteins to the proteasome. Specific E3 ubiquitin ligases are required for ERAD and reside in or near the ER membrane.• ERAD substrates are degraded by the proteasome, a large multicatalytic protease that resides in the cytoplasm. Although integral membrane proteins in the ER can readily access the proteasome, soluble ERAD substrates (that reside within the lumen) must be retrotranslocated or dislocated from the ER to the cytoplasm before they are degraded. •The ERAD pathway is conserved from yeast to humans, and indeed many of the factors that contribute to this pathway were first identified in the yeast Saccharomyces cerevisiae.• A growing number of links between the ERAD pathway and human diseases have been identified.Jeffrey Brodsky holds the Avinoff Chair in the

Highlights d optoTDP43 is a light-inducible model of TDP-43 proteinopathy and is neurotoxic d RNA binding antagonizes aberrant liquid-liquid phase separation (LLPS) of TDP-43 d Acute recruitment to RNA-containing stress granules maintains TDP-43 solubility d TDP-43 targeting oligonucleotides prevent aberrant LLPS and rescues neurotoxicity

More than 2000 mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) have been described that confer a range of molecular cell biological and functional phenotypes. Most of these mutations lead to compromised anion conductance at the apical plasma membrane of secretory epithelia and cause cystic fibrosis (CF) with variable disease severity. Based on the molecular phenotypic complexity of CFTR mutants and their susceptibility to pharmacotherapy, it has been recognized that mutations may impose combinatorial defects in CFTR channel biology. This notion led to the conclusion that the combination of pharmacotherapies addressing single defects (e.g., transcription, translation, folding, and/or gating) may show improved clinical benefit over available low-efficacy monotherapies. Indeed, recent phase 3 clinical trials combining ivacaftor (a gating potentiator) and lumacaftor (a folding corrector) have proven efficacious in CF patients harboring the most common mutation (deletion of residue F508, ΔF508, or Phe508del). This drug combination was recently approved by the U.S. Food and Drug Administration for patients homozygous for ΔF508. Emerging studies of the structural, cell biological, and functional defects caused by rare mutations provide a new framework that reveals a mixture of deficiencies in different CFTR alleles. Establishment of a set of combinatorial categories of the previously defined basic defects in CF alleles will aid the design of even more efficacious therapeutic interventions for CF patients.