X
Forgot Password

If you have forgotten your password you can enter your email here and get a temporary password sent to your email.

Anti-Puromycin Antibody, clone 12D10

RRID:AB_2566826

Antibody ID

AB_2566826

Target Antigen

Puromycin from Streptomyces alboniger

Proper Citation

(Millipore Cat# MABE343, RRID:AB_2566826)

Clonality

monoclonal antibody

Clone ID

12D10

Host Organism

mouse

Vendor

Millipore

Ubiquitination of ABCE1 by NOT4 in Response to Mitochondrial Damage Links Co-translational Quality Control to PINK1-Directed Mitophagy.

  • Wu Z
  • Cell Metab.
  • 2018 Jul 3

Literature context: MABE343; RRID:AB_2566826 Rabbit anti-ABCE1 Dr. R Hegde N


Abstract:

Translation of mRNAs is tightly regulated and constantly surveyed for errors. Aberrant translation can trigger co-translational protein and RNA quality control processes, impairments of which cause neurodegeneration by still poorly understood mechanism(s). Here we show that quality control of translation of mitochondrial outer membrane (MOM)-localized mRNA intersects with the turnover of damaged mitochondria, both orchestrated by the mitochondrial kinase PINK1. Mitochondrial damage causes stalled translation of complex-I 30 kDa subunit (C-I30) mRNA on MOM, triggering the recruitment of co-translational quality control factors Pelo, ABCE1, and NOT4 to the ribosome/mRNA-ribonucleoprotein complex. Damage-induced ubiquitination of ABCE1 by NOT4 generates poly-ubiquitin signals that attract autophagy receptors to MOM to initiate mitophagy. In the Drosophila PINK1 model, these factors act synergistically to restore mitophagy and neuromuscular tissue integrity. Thus ribosome-associated co-translational quality control generates an early signal to trigger mitophagy. Our results have broad therapeutic implications for the understanding and treatment of neurodegenerative diseases.

Funding information:
  • NCI NIH HHS - CA 19014(United States)
  • NIMH NIH HHS - R01 MH080378()
  • NINDS NIH HHS - R01 NS083417()
  • NINDS NIH HHS - R01 NS084412()

Pseudouridylation of tRNA-Derived Fragments Steers Translational Control in Stem Cells.

  • Guzzi N
  • Cell
  • 2018 May 17

Literature context: RRID:AB_2566826 Anti-eIF4A rabbit monoclonal Ce


Abstract:

Pseudouridylation (Ψ) is the most abundant and widespread type of RNA epigenetic modification in living organisms; however, the biological role of Ψ remains poorly understood. Here, we show that a Ψ-driven posttranscriptional program steers translation control to impact stem cell commitment during early embryogenesis. Mechanistically, the Ψ "writer" PUS7 modifies and activates a novel network of tRNA-derived small fragments (tRFs) targeting the translation initiation complex. PUS7 inactivation in embryonic stem cells impairs tRF-mediated translation regulation, leading to increased protein biosynthesis and defective germ layer specification. Remarkably, dysregulation of this posttranscriptional regulatory circuitry impairs hematopoietic stem cell commitment and is common to aggressive subtypes of human myelodysplastic syndromes. Our findings unveil a critical function of Ψ in directing translation control in stem cells with important implications for development and disease.

Funding information:
  • NIDDK NIH HHS - DK068164(United States)

Stress Granule Assembly Disrupts Nucleocytoplasmic Transport.

  • Zhang K
  • Cell
  • 2018 May 3

Literature context: Cat#MABE343; RRID:AB_2566826 Rabbit anti-GR Proteintech Cat#


Abstract:

Defects in nucleocytoplasmic transport have been identified as a key pathogenic event in amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) mediated by a GGGGCC hexanucleotide repeat expansion in C9ORF72, the most common genetic cause of ALS/FTD. Furthermore, nucleocytoplasmic transport disruption has also been implicated in other neurodegenerative diseases with protein aggregation, suggesting a shared mechanism by which protein stress disrupts nucleocytoplasmic transport. Here, we show that cellular stress disrupts nucleocytoplasmic transport by localizing critical nucleocytoplasmic transport factors into stress granules, RNA/protein complexes that play a crucial role in ALS pathogenesis. Importantly, inhibiting stress granule assembly, such as by knocking down Ataxin-2, suppresses nucleocytoplasmic transport defects as well as neurodegeneration in C9ORF72-mediated ALS/FTD. Our findings identify a link between stress granule assembly and nucleocytoplasmic transport, two fundamental cellular processes implicated in the pathogenesis of C9ORF72-mediated ALS/FTD and other neurodegenerative diseases.

Funding information:
  • Canadian Institutes of Health Research - (Canada)

mTORC1 Is Transiently Reactivated in Injured Nerves to Promote c-Jun Elevation and Schwann Cell Dedifferentiation.

  • Norrmén C
  • J. Neurosci.
  • 2018 May 16

Literature context: y (Millipore, catalog #MABE343, RRID:AB_2566826, 1:1000) to detect newly synthe


Abstract:

Schwann cells (SCs) are endowed with a remarkable plasticity. When peripheral nerves are injured, SCs dedifferentiate and acquire new functions to coordinate nerve repair as so-called repair SCs. Subsequently, SCs redifferentiate to remyelinate regenerated axons. Given the similarities between SC dedifferentiation/redifferentiation in injured nerves and in demyelinating neuropathies, elucidating the signals involved in SC plasticity after nerve injury has potentially wider implications. c-Jun has emerged as a key transcription factor regulating SC dedifferentiation and the acquisition of repair SC features. However, the upstream pathways that control c-Jun activity after nerve injury are largely unknown. We report that the mTORC1 pathway is transiently but robustly reactivated in dedifferentiating SCs. By inducible genetic deletion of the functionally crucial mTORC1-subunit Raptor in mouse SCs (including male and female animals), we found that mTORC1 reactivation is necessary for proper myelin clearance, SC dedifferentiation, and consequently remyelination, without major alterations in the inflammatory response. In the absence of mTORC1 signaling, c-Jun failed to be upregulated correctly. Accordingly, a c-Jun binding motif was found to be enriched in promoters of genes with reduced expression in injured mutants. Furthermore, using cultured SCs, we found that mTORC1 is involved in c-Jun regulation by promoting its translation, possibly via the eIF4F-subunit eIF4A. These results provide evidence that proper c-Jun elevation after nerve injury involves also mTORC1-dependent post-transcriptional regulation to ensure timely dedifferentiation of SCs.SIGNIFICANCE STATEMENT A crucial evolutionary acquisition of vertebrates is the envelopment of axons in myelin sheaths produced by oligodendrocytes in the CNS and Schwann cells (SCs) in the PNS. When myelin is damaged, conduction of action potentials along axons slows down or is blocked, leading to debilitating diseases. Unlike oligodendrocytes, SCs have a high regenerative potential, granted by their remarkable plasticity. Thus, understanding the mechanisms underlying SC plasticity may uncover new therapeutic targets in nerve regeneration and demyelinating diseases. Our work reveals that reactivation of the mTORC1 pathway in SCs is essential for efficient SC dedifferentiation after nerve injury. Accordingly, modulating this signaling pathway might be of therapeutic relevance in peripheral nerve injury and other diseases.

Funding information:
  • Medical Research Council - MC_U120081321(United Kingdom)

Axon-Axon Interactions Regulate Topographic Optic Tract Sorting via CYFIP2-Dependent WAVE Complex Function.

  • Cioni JM
  • Neuron
  • 2018 Mar 7

Literature context: ABE343, RRID:AB_2566826 anti-tubulin Sigma-Aldrich Cat#


Abstract:

The axons of retinal ganglion cells (RGCs) are topographically sorted before they arrive at the optic tectum. This pre-target sorting, typical of axon tracts throughout the brain, is poorly understood. Here, we show that cytoplasmic FMR1-interacting proteins (CYFIPs) fulfill non-redundant functions in RGCs, with CYFIP1 mediating axon growth and CYFIP2 specifically involved in axon sorting. We find that CYFIP2 mediates homotypic and heterotypic contact-triggered fasciculation and repulsion responses between dorsal and ventral axons. CYFIP2 associates with transporting ribonucleoprotein particles in axons and regulates translation. Axon-axon contact stimulates CYFIP2 to move into growth cones where it joins the actin nucleating WAVE regulatory complex (WRC) in the periphery and regulates actin remodeling and filopodial dynamics. CYFIP2's function in axon sorting is mediated by its binding to the WRC but not its translational regulation. Together, these findings uncover CYFIP2 as a key regulatory link between axon-axon interactions, filopodial dynamics, and optic tract sorting.

Funding information:
  • NICHD NIH HHS - R01 HD032067(United States)

As Extracellular Glutamine Levels Decline, Asparagine Becomes an Essential Amino Acid.

  • Pavlova NN
  • Cell Metab.
  • 2018 Feb 6

Literature context: 0) EMD Millipore Cat. #MABE343; RRID:AB_2566826 Mouse monoclonal α-Vinculin Sig


Abstract:

When mammalian cells are deprived of glutamine, exogenous asparagine rescues cell survival and growth. Here we report that this rescue results from use of asparagine in protein synthesis. All mammalian cell lines tested lacked cytosolic asparaginase activity and could not utilize asparagine to produce other amino acids or biosynthetic intermediates. Instead, most glutamine-deprived cell lines are capable of sufficient glutamine synthesis to maintain essential amino acid uptake and production of glutamine-dependent biosynthetic precursors, with the exception of asparagine. While experimental introduction of cytosolic asparaginase could enhance the synthesis of glutamine and increase tricarboxylic acid cycle anaplerosis and the synthesis of nucleotide precursors, cytosolic asparaginase suppressed the growth and survival of cells in glutamine-depleted medium in vitro and severely compromised the in vivo growth of tumor xenografts. These results suggest that the lack of asparaginase activity represents an evolutionary adaptation to allow mammalian cells to survive pathophysiologic variations in extracellular glutamine.

Funding information:
  • NCI NIH HHS - P30 CA008748()
  • NIAID NIH HHS - R21 AI091457(United States)

Epitranscriptomic m6A Regulation of Axon Regeneration in the Adult Mammalian Nervous System.

  • Weng YL
  • Neuron
  • 2018 Jan 17

Literature context: ti-Puromycin Millipore MABE343; RRID:AB_2566826 Rabbit anti-ATF3 Santa Cruz sc-


Abstract:

N6-methyladenosine (m6A) affects multiple aspects of mRNA metabolism and regulates developmental transitions by promoting mRNA decay. Little is known about the role of m6A in the adult mammalian nervous system. Here we report that sciatic nerve lesion elevates levels of m6A-tagged transcripts encoding many regeneration-associated genes and protein translation machinery components in the adult mouse dorsal root ganglion (DRG). Single-base resolution m6A-CLIP mapping further reveals a dynamic m6A landscape in the adult DRG upon injury. Loss of either m6A methyltransferase complex component Mettl14 or m6A-binding protein Ythdf1 globally attenuates injury-induced protein translation in adult DRGs and reduces functional axon regeneration in the peripheral nervous system in vivo. Furthermore, Pten deletion-induced axon regeneration of retinal ganglion neurons in the adult central nervous system is attenuated upon Mettl14 knockdown. Our study reveals a critical epitranscriptomic mechanism in promoting injury-induced protein synthesis and axon regeneration in the adult mammalian nervous system.

Funding information:
  • NCI NIH HHS - U01 CA84243(United States)
  • NHGRI NIH HHS - RM1 HG008935()
  • NINDS NIH HHS - P01 NS097206()
  • NINDS NIH HHS - R35 NS097370()

Insulin-Like Growth Factor II Targets the mTOR Pathway to Reverse Autism-Like Phenotypes in Mice.

  • Steinmetz AB
  • J. Neurosci.
  • 2018 Jan 24

Literature context: al antibody (1/5000; Millipore; RRID:AB_2566826). Signal intensities were norma


Abstract:

Autism spectrum disorder (ASD) is a developmental disability characterized by impairments in social interaction and repetitive behavior, and is also associated with cognitive deficits. There is no current treatment that can ameliorate most of the ASD symptomatology; thus, identifying novel therapies is urgently needed. We used male BTBR T+Itpr3tf /J (BTBR) mice, a model that reproduces most of the core behavioral phenotypes of ASD, to test the effects of systemic administration of insulin-like growth factor II (IGF-II), a polypeptide that crosses the blood-brain barrier and acts as a cognitive enhancer. We show that systemic IGF-II treatments reverse the typical defects in social interaction, cognitive/executive functions, and repetitive behaviors reflective of ASD-like phenotypes. In BTBR mice, IGF-II, via IGF-II receptor, but not via IGF-I receptor, reverses the abnormal levels of the AMPK-mTOR-S6K pathway and of active translation at synapses. Thus, IGF-II may represent a novel potential therapy for ASD.SIGNIFICANCE STATEMENT Currently, there is no effective treatment for autism spectrum disorder (ASD), a developmental disability affecting a high number of children. Using a mouse model that expresses most of the key core as well as associated behavioral deficits of ASD, that are, social, cognitive, and repetitive behaviors, we report that a systemic administration of the polypeptide insulin-like growth factor II (IGF-II) reverses all these deficits. The effects of IGF-II occur via IGF-II receptors, and not IGF-I receptors, and target both basal and learning-dependent molecular abnormalities found in several ASD mice models, including those of identified genetic mutations. We suggest that IGF-II represents a potential novel therapeutic target for ASD.

Funding information:
  • NCATS NIH HHS - UL1 TR000135(United States)
  • NIMH NIH HHS - F31 MH090636()
  • NIMH NIH HHS - R01 MH074736()
  • NIMH NIH HHS - R37 MH065635()
  • NIMH NIH HHS - T32 MH019524()
  • NIMH NIH HHS - T32 MH087004()

Nuclear localization of EIF4G3 suggests a role for the XY body in translational regulation during spermatogenesis in mice.

  • Hu J
  • Biol. Reprod.
  • 2018 Jan 1

Literature context: omycin Mo Millipore MABE343 RRID:AB_2566826 1:10000 n/a


Abstract:

Eukaryotic translation initiation factor 4G (EIF4G) is an important scaffold protein in the translation initiation complex. In mice, mutation of the Eif4g3 gene causes male infertility, with arrest of meiosis at the end of meiotic prophase. This study documents features of the developmental expression and subcellular localization of EIF4G3 that might contribute to its highly specific role in meiosis and spermatogenesis. Quite unexpectedly, EIF4G3 is located in the nucleus of spermatocytes, where it is highly enriched in the XY body, the chromatin domain formed by the transcriptionally inactive sex chromosomes. Moreover, many other, but not all, translation-related proteins are also localized in the XY body. These unanticipated observations implicate roles for the XY body in controlling mRNA metabolism and/or "poising" protein translation complexes before the meiotic division phase in spermatocytes.

Funding information:
  • Medical Research Council - MC_UU_12012/5(United Kingdom)

RNA Docking and Local Translation Regulate Site-Specific Axon Remodeling In Vivo.

  • Wong HH
  • Neuron
  • 2017 Aug 16

Literature context: n Millipore Cat# MABE343-AF488; RRID:AB_2566826 Rabbit anti-β-actin Abcam Cat#


Abstract:

Nascent proteins can be positioned rapidly at precise subcellular locations by local protein synthesis (LPS) to facilitate localized growth responses. Axon arbor architecture, a major determinant of synaptic connectivity, is shaped by localized growth responses, but it is unknown whether LPS influences these responses in vivo. Using high-resolution live imaging, we examined the spatiotemporal dynamics of RNA and LPS in retinal axons during arborization in vivo. Endogenous RNA tracking reveals that RNA granules dock at sites of branch emergence and invade stabilized branches. Live translation reporter analysis reveals that de novo β-actin hotspots colocalize with docked RNA granules at the bases and tips of new branches. Inhibition of axonal β-actin mRNA translation disrupts arbor dynamics primarily by reducing new branch emergence and leads to impoverished terminal arbors. The results demonstrate a requirement for LPS in building arbor complexity and suggest a key role for pre-synaptic LPS in assembling neural circuits.

The Mammalian Ribo-interactome Reveals Ribosome Functional Diversity and Heterogeneity.

  • Simsek D
  • Cell
  • 2017 Jun 1

Literature context: 12D10)EMD MilliporeCat#MABE343; RRID: AB_2566826Rabbit polyclonal anti-RPS3/uS3B


Abstract:

During eukaryotic evolution, ribosomes have considerably increased in size, forming a surface-exposed ribosomal RNA (rRNA) shell of unknown function, which may create an interface for yet uncharacterized interacting proteins. To investigate such protein interactions, we establish a ribosome affinity purification method that unexpectedly identifies hundreds of ribosome-associated proteins (RAPs) from categories including metabolism and cell cycle, as well as RNA- and protein-modifying enzymes that functionally diversify mammalian ribosomes. By further characterizing RAPs, we discover the presence of ufmylation, a metazoan-specific post-translational modification (PTM), on ribosomes and define its direct substrates. Moreover, we show that the metabolic enzyme, pyruvate kinase muscle (PKM), interacts with sub-pools of endoplasmic reticulum (ER)-associated ribosomes, exerting a non-canonical function as an RNA-binding protein in the translation of ER-destined mRNAs. Therefore, RAPs interconnect one of life's most ancient molecular machines with diverse cellular processes, providing an additional layer of regulatory potential to protein expression.

Funding information:
  • NICHD NIH HHS - R21 HD086730()
  • NIH HHS - DP2 OD008509()

The PERK arm of the unfolded protein response regulates satellite cell-mediated skeletal muscle regeneration.

  • Xiong G
  • Elife
  • 2017 Mar 23

Literature context: # MABE343 RRID:AB_2566826).


Abstract:

Regeneration of skeletal muscle in adults is mediated by satellite stem cells. Accumulation of misfolded proteins triggers endoplasmic reticulum stress that leads to unfolded protein response (UPR). The UPR is relayed to the cell through the activation of PERK, IRE1/XBP1, and ATF6. Here, we demonstrate that levels of PERK and IRE1 are increased in satellite cells upon muscle injury. Inhibition of PERK, but not the IRE1 arm of the UPR in satellite cells inhibits myofiber regeneration in adult mice. PERK is essential for the survival and differentiation of activated satellite cells into the myogenic lineage. Deletion of PERK causes hyper-activation of p38 MAPK during myogenesis. Blocking p38 MAPK activity improves the survival and differentiation of PERK-deficient satellite cells in vitro and muscle formation in vivo. Collectively, our results suggest that the PERK arm of the UPR plays a pivotal role in the regulation of satellite cell homeostasis during regenerative myogenesis.

Funding information:
  • NIA NIH HHS - R01 AG029623()
  • NIAMS NIH HHS - R01 AR059810()
  • NIAMS NIH HHS - R01 AR068313()

Hermes Regulates Axon Sorting in the Optic Tract by Post-Trancriptional Regulation of Neuropilin 1.

  • Hörnberg H
  • J. Neurosci.
  • 2016 Dec 14

Literature context: (1:1000, RRID:AB_2566826, catalog #


Abstract:

The establishment of precise topographic maps during neural development is facilitated by the presorting of axons in the pathway before they reach their targets. In the vertebrate visual system, such topography is seen clearly in the optic tract (OT) and in the optic radiations. However, the molecular mechanisms involved in pretarget axon sorting are poorly understood. Here, we show in zebrafish that the RNA-binding protein Hermes, which is expressed exclusively in retinal ganglion cells (RGCs), is involved in this process. Using a RiboTag approach, we show that Hermes acts as a negative translational regulator of specific mRNAs in RGCs. One of these targets is the guidance cue receptor Neuropilin 1 (Nrp1), which is sensitive to the repellent cue Semaphorin 3A (Sema3A). Hermes knock-down leads to topographic missorting in the OT through the upregulation of Nrp1. Restoring Nrp1 to appropriate levels in Hermes-depleted embryos rescues this effect and corrects the axon-sorting defect in the OT. Our data indicate that axon sorting relies on Hermes-regulated translation of Nrp1. SIGNIFICANCE STATEMENT: An important mechanism governing the formation of the mature neural map is pretarget axon sorting within the sensory tract; however, the molecular mechanisms involved in this process remain largely unknown. The work presented here reveals a novel function for the RNA-binding protein Hermes in regulating the topographic sorting of retinal ganglion cell (RGC) axons in the optic tract and tectum. We find that Hermes negatively controls the translation of the guidance cue receptor Neuropilin-1 in RGCs, with Hermes knock-down resulting in aberrant growth cone cue sensitivity and axonal topographic misprojections. We characterize a novel RNA-based mechanism by which axons restrict their translatome developmentally to achieve proper targeting.

Funding information:
  • European Research Council - 322817()

Translational control of auditory imprinting and structural plasticity by eIF2α.

  • Batista G
  • Elife
  • 2016 Dec 23

Literature context: MABE343, RRID:AB_2566826) and S6 (C


Abstract:

The formation of imprinted memories during a critical period is crucial for vital behaviors, including filial attachment. Yet, little is known about the underlying molecular mechanisms. Using a combination of behavior, pharmacology, in vivo surface sensing of translation (SUnSET) and DiOlistic labeling we found that, translational control by the eukaryotic translation initiation factor 2 alpha (eIF2α) bidirectionally regulates auditory but not visual imprinting and related changes in structural plasticity in chickens. Increasing phosphorylation of eIF2α (p-eIF2α) reduces translation rates and spine plasticity, and selectively impairs auditory imprinting. By contrast, inhibition of an eIF2α kinase or blocking the translational program controlled by p-eIF2α enhances auditory imprinting. Importantly, these manipulations are able to reopen the critical period. Thus, we have identified a translational control mechanism that selectively underlies auditory imprinting. Restoring translational control of eIF2α holds the promise to rejuvenate adult brain plasticity and restore learning and memory in a variety of cognitive disorders.

Funding information:
  • NINDS NIH HHS - NS94668(United States)

Impaired eukaryotic translation initiation factor 2B activity specifically in oligodendrocytes reproduces the pathology of vanishing white matter disease in mice.

  • Lin Y
  • J. Neurosci.
  • 2014 Sep 3

Literature context: The blots were incubated with a primary antibody against puromycin (1:5000, EMD Millipore, Catalog


Abstract:

Vanishing white matter disease (VWMD) is an inherited autosomal-recessive hypomyelinating disease caused by mutations in eukaryotic translation initiation factor 2B (eIF2B). eIF2B mutations predominantly affect the brain white matter, and the characteristic features of VWMD pathology include myelin loss and foamy oligodendrocytes. Activation of pancreatic endoplasmic reticulum kinase (PERK) has been observed in oligodendrocytes in VWMD. PERK activation in response to endoplasmic reticulum stress attenuates eIF2B activity by phosphorylating eIF2α, suggesting that impaired eIF2B activity in oligodendrocytes induced by VWMD mutations or PERK activation exploit similar mechanisms to promote selective white matter pathology in VWMD. Using transgenic mice that allow for temporally controlled activation of PERK specifically in oligodendrocytes, we discovered that strong PERK activation in oligodendrocytes during development suppressed eIF2B activity and reproduced the characteristic features of VWMD in mice, including hypomyelinating phenotype, foamy oligodendrocytes, and myelin loss. Notably, impaired eIF2B activity induced by PERK activation in oligodendrocytes of fully myelinated adult mice had minimal effects on morphology or function. Our observations point to a cell-autonomous role of impaired eIF2B activity in myelinating oligodendrocytes in the pathogenesis of VWMD.