Reproductive regulation of the mitochondrial stress response in Caenorhabditis elegans.

Proteome integrity is fundamental for cellular and organismal homeostasis. The mitochondrial unfolded protein response (UPRmt), a key component of the proteostasis network, is activated in a non-cell-autonomous manner in response to mitochondrial stress in distal tissues. However, the importance of inter-tissue communication for UPRmt inducibility under physiological conditions remains elusive. Here, we show that an intact germline is essential for robust UPRmt induction in the Caenorhabditis elegans somatic tissues. A series of nematode mutants with germline defects are unable to respond to genetic or chemical UPRmt inducers. Our genetic analysis suggests that reproductive signals, rather than germline stem cells, are responsible for somatic UPRmt induction. Consistent with this observation, we show that UPRmt is sexually dimorphic, as male nematodes are inherently unresponsive to mitochondrial stress. Our findings highlight a paradigm of germline-somatic communication and suggest that reproductive cessation is a primary cause of age-related UPRmt decline.

Assessing polyglutamine tract aggregation in the nematode Caenorhabditis elegans.

Proteome integrity is a prerequisite for cellular functionality and organismal viability. Its compromise is considered an inherent part of the aging process and has been associated with the onset of age-related, neurodegenerative pathologies. Although the molecular underpinnings of protein homeostasis (proteostasis) have been extensively studied, several aspects of its regulation remain elusive. The nematode Caenorhabditis elegans has emerged as a versatile, heterologous model organism to study the dynamics of aggregation-prone human proteins in vivo. Here, we describe an experimental pipeline for the analysis of polyglutamine (polyQ) tract aggregation, as a measure of the state of proteostasis, during aging.

The interplay between selective types of (macro)autophagy: Mitophagy and xenophagy.

Autophagy is a physiological response, activated by a myriad of endogenous and exogenous cues, including DNA damage, perturbation of proteostasis, depletion of nutrients or oxygen and pathogen infection. Upon sensing those stimuli, cells employ multiple non-selective and selective autophagy pathways to promote fitness and survival. Importantly, there are a variety of selective types of autophagy. In this review we will focus on autophagy of bacteria (xenophagy) and autophagy of mitochondria (mitophagy). We provide a brief introduction to bulk autophagy, as well as xenophagy and mitophagy, highlighting their common molecular factors. We also describe the role of xenophagy and mitophagy in the detection and elimination of pathogens by the immune system and the adaptive mechanisms that some pathogens have developed through evolution to escape the host autophagic response. Finally, we summarize the recent articles (from the last five years) linking bulk autophagy with xenophagy and/or mitophagy in the context on developmental biology, cancer and metabolism.

The κ-opioid receptor-induced autophagy is implicated in stress-driven synaptic alterations.

Recent evidence has shown that G protein-coupled receptors (GPCRs) are direct sensors of the autophagic machinery and opioid receptors regulate neuronal plasticity and neurotransmission with an as yet unclarified mechanism. Using in vitro and in vivo experimental approaches, this study aims to clarify the potential role of autophagy and κ-opioid receptor (κ-OR) signaling in synaptic alterations. We hereby demonstrate that the selective κ-OR agonist U50,488H, induces autophagy in a time-and dose-dependent manner in Neuro-2A cells stably expressing the human κ-OR by upregulating microtubule-associated protein Light Chain 3-II (LC3-II), Beclin 1 and Autophagy Related Gene 5 (ATG5). Pretreatment of neuronal cells with pertussis toxin blocked the above κ-OR-mediated cellular responses. Our molecular analysis also revealed a κ-OR-driven upregulation of becn1 gene through ERK1,2-dependent activation of the transcription factor CREB in Neuro-2A cells. Moreover, our studies demonstrated that sub-chronic U50,488H administration in mice causes profound increases of specific autophagic markers in the hippocampus with a concomitant decrease of several pre-and post-synaptic proteins, such as spinophilin, postsynaptic density protein 95 (PSD-95) and synaptosomal associated protein 25 (SNAP25). Finally, using acute stress, a stimulus known to increase the levels of the endogenous κ-OR ligand dynorphin, we are demonstrating that administration of the κ-ΟR selective antagonist, nor-binaltorphimine (norBNI), blocks the induction of autophagy and the stress-evoked reduction of synaptic proteins in the hippocampus. These findings provide novel insights about the essential role of autophagic machinery into the mechanisms through which κ-OR signaling regulates brain plasticity.

Sustained intracellular calcium rise mediates neuronal mitophagy in models of autosomal dominant optic atrophy.

Mitochondrial dysfunction and mitophagy are often hallmarks of neurodegenerative diseases such as autosomal dominant optic atrophy (ADOA) caused by mutations in the key mitochondrial dynamics protein optic atrophy 1 (Opa1). However, the second messengers linking mitochondrial dysfunction to initiation of mitophagy remain poorly characterized. Here, we show in mammalian and nematode neurons that Opa1 mutations trigger Ca2+-dependent mitophagy. Deletion or expression of mutated Opa1 in mouse retinal ganglion cells and Caenorhabditis elegans motor neurons lead to mitochondrial dysfunction, increased cytosolic Ca2+ levels, and decreased axonal mitochondrial density. Chelation of Ca2+ restores mitochondrial density in neuronal processes, neuronal function, and viability. Mechanistically, sustained Ca2+ levels activate calcineurin and AMPK, placed in the same genetic pathway regulating axonal mitochondrial density. Our data reveal that mitophagy in ADOA depends on Ca2+-calcineurin-AMPK signaling cascade.

Monitoring autophagic flux in Caenorhabditis elegans using a p62/SQST-1 reporter.

Autophagy is a well-conserved self-degrading mechanism, which involves the elimination of unnecessary or damaged cellular constituents. Although extensively studied, many aspects regarding its tight regulation and its implication in health and disease remain elusive. The nematode Caenorhabditis elegans has been widely used as a simple multicellular model organism for studying the autophagic machinery per se, and uncover its multidimensional roles in the maintenance of cellular and organismal homeostasis. The current protocol describes the in vivo detection and biochemical analysis of the autophagic substrate SQST-1, as an indicator of autophagic flux in C. elegans.

The Synthetic Microneurotrophin BNN27 Affects Retinal Function in Rats With Streptozotocin-Induced Diabetes.

BNN27, a C17-spiroepoxy derivative of DHEA, was shown to have antiapoptotic properties via mechanisms involving the nerve growth factor receptors (tropomyosin-related kinase A [TrkA]/neurotrophin receptor p75 [p75NTR]). In this study, we examined the effects of BNN27 on neural/glial cell function, apoptosis, and inflammation in the experimental rat streptozotocin (STZ) model of diabetic retinopathy (DR). The ability of BNN27 to activate the TrkA receptor and regulate p75NTR expression was investigated. BNN27 (2,10, and 50 mg/kg i.p. for 7 days) administration 4 weeks post-STZ injection (paradigm A) reversed the diabetes-induced glial activation and loss of function of amacrine cells (brain nitric oxide synthetase/tyrosine hydroxylase expression) and ganglion cell axons via a TrkA receptor (TrkAR)-dependent mechanism. BNN27 activated/phosphorylated the TrkAY490 residue in the absence but not the presence of TrkAR inhibitor and abolished the diabetes-induced increase in p75NTR expression. However, it had no effect on retinal cell death (TUNEL+ cells). A similar result was observed when BNN27 (10 mg/kg i.p.) was administered at the onset of diabetes, every other day for 4 weeks (paradigm B). However, BNN27 decreased the activation of caspase-3 in both paradigms. Finally, BNN27 reduced the proinflammatory (TNFα and IL-1ß) and increased the anti-inflammatory (IL-10 and IL-4) cytokine levels. These findings suggest that BNN27 has the pharmacological profile of a therapeutic for DR, since it targets both the neurodegenerative and inflammatory components of the disease.