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Human LC3 Polyclonal Antibody

RRID:AB_2274121

Antibody ID

AB_2274121

Target Antigen

Human LC3 human

Proper Citation

(MBL International Cat# PM036, RRID:AB_2274121)

Clonality

polyclonal antibody

Comments

manufacturer recommendations: IgG; IgG Immunocytochemistry; Western Blot; WB, ICC

Host Organism

rabbit

Vendor

MBL International

ROS-Mediated 15-Hydroxyprostaglandin Dehydrogenase Degradation via Cysteine Oxidation Promotes NAD+-Mediated Epithelial-Mesenchymal Transition.

  • Wang W
  • Cell Chem Biol
  • 2018 Mar 15

Literature context:


Abstract:

Nicotinamide adenine dinucleotide (NAD) levels decrease with aging as a result of aging-associated CD38 upregulation. Here, we established a cell model with decreased cellular NAD levels by overexpressing CD38 or treating cells with FK866, an inhibitor of nicotinamide phosphoribosyltransferase. We revealed that decreased NAD triggered reactive oxygen species (ROS)-mediated degradation of 15-hydroxyprostaglandin dehydrogenase (15-PGDH), which drove cells to undergo epithelial-mesenchymal transition (EMT). Moreover, we showed that oxidation of the Cys44 residue to sulfonic acid in 15-PGDH led to its degradation via non-canonical ubiquitination-proteasome and autophagy pathways. Mutation of Cys44 to alanine abolished ROS-induced 15-PGDH degradation. We demonstrated that 15-PGDH silencing promoted EMT, whereas supplementation with NAD precursors increased NAD and 15-PGDH stability, and reversed the EMT process. Taken together, these results suggest that declining NAD levels contribute to age-dependent increases in cancer incidence, and repletion of NAD precursors is beneficial for increasing 15-PGDH expression.

Funding information:
  • National Health and Medical Research Council - 1061122(United States)

Identification of New Activators of Mitochondrial Fusion Reveals a Link between Mitochondrial Morphology and Pyrimidine Metabolism.

  • Miret-Casals L
  • Cell Chem Biol
  • 2018 Mar 15

Literature context:


Abstract:

Mitochondria are dynamic organelles that produce most of the cellular ATP, and are involved in many other cellular functions such as Ca2+ signaling, differentiation, apoptosis, cell cycle, and cell growth. One key process of mitochondrial dynamics is mitochondrial fusion, which is catalyzed by mitofusins (MFN1 and MFN2) and OPA1. The outer mitochondrial membrane protein MFN2 plays a relevant role in the maintenance of mitochondrial metabolism, insulin signaling, and mutations that cause neurodegenerative disorders. Therefore, modulation of proteins involved in mitochondrial dynamics has emerged as a potential pharmacological strategy. Here, we report the identification of small molecules by high-throughput screen that promote mitochondrial elongation in an MFN1/MFN2-dependent manner. Detailed analysis of their mode of action reveals a previously unknown connection between pyrimidine metabolism and mitochondrial dynamics. Our data indicate a link between pyrimidine biosynthesis and mitochondrial dynamics, which maintains cell survival under stress conditions characterized by loss of pyrimidine synthesis.

Mitochondrial Fission Promotes the Continued Clearance of Apoptotic Cells by Macrophages.

  • Wang Y
  • Cell
  • 2017 Oct 5

Literature context:


Abstract:

Clearance of apoptotic cells (ACs) by phagocytes (efferocytosis) prevents post-apoptotic necrosis and dampens inflammation. Defective efferocytosis drives important diseases, including atherosclerosis. For efficient efferocytosis, phagocytes must be able to internalize multiple ACs. We show here that uptake of multiple ACs by macrophages requires dynamin-related protein 1 (Drp1)-mediated mitochondrial fission, which is triggered by AC uptake. When mitochondrial fission is disabled, AC-induced increase in cytosolic calcium is blunted owing to mitochondrial calcium sequestration, and calcium-dependent phagosome formation around secondarily encountered ACs is impaired. These defects can be corrected by silencing the mitochondrial calcium uniporter (MCU). Mice lacking myeloid Drp1 showed defective efferocytosis and its pathologic consequences in the thymus after dexamethasone treatment and in advanced atherosclerotic lesions in fat-fed Ldlr-/- mice. Thus, mitochondrial fission in response to AC uptake is a critical process that enables macrophages to clear multiple ACs and to avoid the pathologic consequences of defective efferocytosis in vivo.

Viral Replication Complexes Are Targeted by LC3-Guided Interferon-Inducible GTPases.

  • Biering SB
  • Cell Host Microbe
  • 2017 Jul 12

Literature context:


Abstract:

All viruses with positive-sense RNA genomes replicate on membranous structures in the cytoplasm called replication complexes (RCs). RCs provide an advantageous microenvironment for viral replication, but it is unknown how the host immune system counteracts these structures. Here we show that interferon-gamma (IFNG) disrupts the RC of murine norovirus (MNV) via evolutionarily conserved autophagy proteins and the induction of IFN-inducible GTPases, which are known to destroy the membrane of vacuoles containing bacteria, protists, or fungi. The MNV RC was marked by the microtubule-associated-protein-1-light-chain-3 (LC3) conjugation system of autophagy and then targeted by immunity-related GTPases (IRGs) and guanylate-binding proteins (GBPs) upon their induction by IFNG. Further, the LC3 conjugation system and the IFN-inducible GTPases were necessary to inhibit MNV replication in mice and human cells. These data suggest that viral RCs can be marked and antagonized by a universal immune defense mechanism targeting diverse pathogens replicating in cytosolic membrane structures.

Funding information:
  • NIAID NIH HHS - R01 AI103197()

Parkinson Sac Domain Mutation in Synaptojanin 1 Impairs Clathrin Uncoating at Synapses and Triggers Dystrophic Changes in Dopaminergic Axons.

  • Cao M
  • Neuron
  • 2017 Feb 22

Literature context:


Abstract:

Synaptojanin 1 (SJ1) is a major presynaptic phosphatase that couples synaptic vesicle endocytosis to the dephosphorylation of PI(4,5)P2, a reaction needed for the shedding of endocytic factors from their membranes. While the role of SJ1's 5-phosphatase module in this process is well recognized, the contribution of its Sac phosphatase domain, whose preferred substrate is PI4P, remains unclear. Recently a homozygous mutation in its Sac domain was identified in early-onset parkinsonism patients. We show that mice carrying this mutation developed neurological manifestations similar to those of human patients. Synapses of these mice displayed endocytic defects and a striking accumulation of clathrin-coated intermediates, strongly implicating Sac domain's activity in endocytic protein dynamics. Mutant brains had elevated auxilin (PARK19) and parkin (PARK2) levels. Moreover, dystrophic axonal terminal changes were selectively observed in dopaminergic axons in the dorsal striatum. These results strengthen evidence for a link between synaptic endocytic dysfunction and Parkinson's disease.

Funding information:
  • NCATS NIH HHS - UL1 TR001863()
  • NIDA NIH HHS - P30 DA018343()
  • NIGMS NIH HHS - P41 GM103412()
  • NINDS NIH HHS - R01 NS036251()
  • NINDS NIH HHS - R01 NS036942()
  • NINDS NIH HHS - R37 NS036251()
  • NINDS NIH HHS - R37 NS036942()

Multiplex image-based autophagy RNAi screening identifies SMCR8 as ULK1 kinase activity and gene expression regulator.

  • Jung J
  • Elife
  • 2017 Feb 14

Literature context:


Abstract:

Autophagy is an intracellular recycling and degradation pathway that depends on membrane trafficking. Rab GTPases are central for autophagy but their regulation especially through the activity of Rab GEFs remains largely elusive. We employed a RNAi screen simultaneously monitoring different populations of autophagosomes and identified 34 out of 186 Rab GTPase, GAP and GEF family members as potential autophagy regulators, amongst them SMCR8. SMCR8 uses overlapping binding regions to associate with C9ORF72 or with a C9ORF72-ULK1 kinase complex holo-assembly, which function in maturation and formation of autophagosomes, respectively. While focusing on the role of SMCR8 during autophagy initiation, we found that kinase activity and gene expression of ULK1 are increased upon SMCR8 depletion. The latter phenotype involved association of SMCR8 with the ULK1 gene locus. Global mRNA expression analysis revealed that SMCR8 regulates transcription of several other autophagy genes including WIPI2. Collectively, we established SMCR8 as multifaceted negative autophagy regulator.

Genetic screen in Drosophila muscle identifies autophagy-mediated T-tubule remodeling and a Rab2 role in autophagy.

  • Fujita N
  • Elife
  • 2017 Jan 7

Literature context:


Abstract:

Transverse (T)-tubules make-up a specialized network of tubulated muscle cell membranes involved in excitation-contraction coupling for power of contraction. Little is known about how T-tubules maintain highly organized structures and contacts throughout the contractile system despite the ongoing muscle remodeling that occurs with muscle atrophy, damage and aging. We uncovered an essential role for autophagy in T-tubule remodeling with genetic screens of a developmentally regulated remodeling program in Drosophila abdominal muscles. Here, we show that autophagy is both upregulated with and required for progression through T-tubule disassembly stages. Along with known mediators of autophagosome-lysosome fusion, our screens uncovered an unexpected shared role for Rab2 with a broadly conserved function in autophagic clearance. Rab2 localizes to autophagosomes and binds to HOPS complex members, suggesting a direct role in autophagosome tethering/fusion. Together, the high membrane flux with muscle remodeling permits unprecedented analysis both of T-tubule dynamics and fundamental trafficking mechanisms.