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Anti-Ubiquitin, Lys63-Specific, clone Apu3 antibody


Antibody ID


Target Antigen

Ubiquitin Lys63-Specific clone Apu3 h, m, r, rb, rabbit

Proper Citation

(Millipore Cat# 05-1308, RRID:AB_1587580)


monoclonal antibody


seller recommendations: IgG; IgG Functional Assay; Cell Function Assay, IC, IP, IH, WB; Immunohistochemistry; Immunoprecipitation; Immunocytochemistry; Western Blot

Host Organism




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: anti-K63-Ub Millipore 05-1308; RRID:AB_1587580 Rabbit anti-OPTN Abcam ab23666;


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()

Discovery and Characterization of ZUFSP/ZUP1, a Distinct Deubiquitinase Class Important for Genome Stability.

  • Kwasna D
  • Mol. Cell
  • 2018 Apr 5

Literature context: pu3) Millipore Cat# 05-1308, RRID:AB_1587580 Rabbit anti-Ub Dako Cat# Z0458,


Deubiquitinating enzymes (DUBs) are important regulators of ubiquitin signaling. Here, we report the discovery of deubiquitinating activity in ZUFSP/C6orf113. High-resolution crystal structures of ZUFSP in complex with ubiquitin reveal several distinctive features of ubiquitin recognition and catalysis. Our analyses reveal that ZUFSP is a novel DUB with no homology to any known DUBs, leading us to classify ZUFSP as the seventh DUB family. Intriguingly, the minimal catalytic domain does not cleave polyubiquitin. We identify two ubiquitin binding domains in ZUFSP: a ZHA (ZUFSP helical arm) that binds to the distal ubiquitin and an atypical UBZ domain in ZUFSP that binds to polyubiquitin. Importantly, both domains are essential for ZUFSP to selectively cleave K63-linked polyubiquitin. We show that ZUFSP localizes to DNA lesions, where it plays an important role in genome stability pathways, functioning to prevent spontaneous DNA damage and also promote cellular survival in response to exogenous DNA damage.

Funding information:
  • NIA NIH HHS - R21 AG040683(United States)

Metal Sensing by the IRT1 Transporter-Receptor Orchestrates Its Own Degradation and Plant Metal Nutrition.

  • Dubeaux G
  • Mol. Cell
  • 2018 Mar 15

Literature context: Cat# 05-1308, RRID:AB_1587580 Anti-Phospho-Thr Cell Signaling


Plant roots forage the soil for iron, the concentration of which can be dramatically lower than those needed for growth. Soil iron uptake uses the broad metal spectrum IRT1 transporter that also transports zinc, manganese, cobalt, and cadmium. Sophisticated iron-dependent transcriptional regulatory mechanisms allow plants to tightly control the abundance of IRT1, ensuring optimal absorption of iron. Here, we uncover that IRT1 acts as a transporter and receptor (transceptor), directly sensing excess of its non-iron metal substrates in the cytoplasm, to regulate its own degradation. Direct metal binding to a histidine-rich stretch in IRT1 triggers its phosphorylation by the CIPK23 kinase and facilitates the subsequent recruitment of the IDF1 E3 ligase. CIPK23-driven phosphorylation and IDF1-mediated lysine-63 polyubiquitination are jointly required for efficient endosomal sorting and vacuolar degradation of IRT1. Thus, IRT1 directly senses elevated non-iron metal concentrations and integrates multiple substrate-dependent regulations to optimize iron uptake and protect plants from highly reactive metals.

Funding information:
  • Wellcome Trust - 090532(United Kingdom)

Ubiquitin Linkage-Specific Affimers Reveal Insights into K6-Linked Ubiquitin Signaling.

  • Michel MA
  • Mol. Cell
  • 2017 Oct 5

Literature context: RRID:AB_1587580 α-TOM20 Santa Cruz Biotechnolog


Several ubiquitin chain types have remained unstudied, mainly because tools and techniques to detect these posttranslational modifications are scarce. Linkage-specific antibodies have shaped our understanding of the roles and dynamics of polyubiquitin signals but are available for only five out of eight linkage types. We here characterize K6- and K33-linkage-specific "affimer" reagents as high-affinity ubiquitin interactors. Crystal structures of affimers bound to their cognate chain types reveal mechanisms of specificity and a K11 cross-reactivity in the K33 affimer. Structure-guided improvements yield superior affinity reagents suitable for western blotting, confocal fluorescence microscopy and pull-down applications. This allowed us to identify RNF144A and RNF144B as E3 ligases that assemble K6-, K11-, and K48-linked polyubiquitin in vitro. A protocol to enrich K6-ubiquitinated proteins from cells identifies HUWE1 as a main E3 ligase for this chain type, and we show that mitofusin-2 is modified with K6-linked polyubiquitin in a HUWE1-dependent manner.

Funding information:
  • European Research Council - 309756()

Centriolar Satellites Control GABARAP Ubiquitination and GABARAP-Mediated Autophagy.

  • Joachim J
  • Curr. Biol.
  • 2017 Jul 24

Literature context: #05-1308; RRID:AB_1587580 Rabbit ant


Autophagy maintains cellular health and homeostasis during stress by delivering cytosolic material captured by autophagosomes to lysosomes for degradation. Autophagosome formation is complex: initiated by the recruitment of autophagy (Atg) proteins to the formation site, it is sustained by activation of Atg proteins to allow growth and closure of the autophagosome. How Atg proteins are translocated to the forming autophagosome is not fully understood. Transport of the ATG8 family member GABARAP from the centrosome occurs during starvation-induced autophagosome biogenesis, but how centrosomal proteins regulate GABARAP localization is unknown. We show that the centriolar satellite protein PCM1 regulates the recruitment of GABARAP to the pericentriolar material. In addition to residing on the pericentriolar material, GABARAP marks a subtype of PCM1-positive centriolar satellites. GABARAP, but not another ATG8 family member LC3B, binds directly to PCM1 through a canonical LIR motif. Loss of PCM1 results in destabilization of GABARAP, but not LC3B, through proteasomal degradation. GABARAP instability is mediated through the centriolar satellite E3 ligase Mib1, which interacts with GABARAP through its substrate-binding region and promotes K48-linked ubiquitination of GABARAP. Ubiquitination of GABARAP occurs in the N terminus, a domain associated with ATG8-family-specific functions during autophagosome formation, on residues absent in the LC3 family. Furthermore, PCM1-GABARAP-positive centriolar satellites colocalize with forming autophagosomes. PCM1 enhances GABARAP/WIPI2/p62-positive autophagosome formation and flux but has no significant effect on LC3B-positive autophagosome formation. These data suggest a mechanism for how centriolar satellites can specifically regulate an ATG8 ortholog, the centrosomal GABARAP reservoir, and centrosome-autophagosome crosstalk.

Funding information:
  • NIGMS NIH HHS - R01 HD069647()
  • Wellcome Trust - R01 GM120776()

Cyclophilin A-regulated ubiquitination is critical for RIG-I-mediated antiviral immune responses.

  • Liu W
  • Elife
  • 2017 Jun 8

Literature context: illipore, RRID:AB_1587580). For immu


RIG-I is a key cytosolic pattern recognition receptor that interacts with MAVS to induce type I interferons (IFNs) against RNA virus infection. In this study, we found that cyclophilin A (CypA), a peptidyl-prolyl cis/trans isomerase, functioned as a critical positive regulator of RIG-I-mediated antiviral immune responses. Deficiency of CypA impaired RIG-I-mediated type I IFN production and promoted viral replication in human cells and mice. Upon Sendai virus infection, CypA increased the interaction between RIG-I and its E3 ubiquitin ligase TRIM25, leading to enhanced TRIM25-mediated K63-linked ubiquitination of RIG-I that facilitated recruitment of RIG-I to MAVS. In addition, CypA and TRIM25 competitively interacted with MAVS, thereby inhibiting TRIM25-induced K48-linked ubiquitination of MAVS. Taken together, our findings reveal an essential role of CypA in boosting RIG-I-mediated antiviral immune responses by controlling the ubiquitination of RIG-I and MAVS.

YOD1/TRAF6 association balances p62-dependent IL-1 signaling to NF-κB.

  • Schimmack G
  • Elife
  • 2017 Feb 28

Literature context: itin K63 (RRID:AB_1587580, Millipore


The ubiquitin ligase TRAF6 is a key regulator of canonical IκB kinase (IKK)/NF-κB signaling in response to interleukin-1 (IL-1) stimulation. Here, we identified the deubiquitinating enzyme YOD1 (OTUD2) as a novel interactor of TRAF6 in human cells. YOD1 binds to the C-terminal TRAF homology domain of TRAF6 that also serves as the interaction surface for the adaptor p62/Sequestosome-1, which is required for IL-1 signaling to NF-κB. We show that YOD1 competes with p62 for TRAF6 association and abolishes the sequestration of TRAF6 to cytosolic p62 aggregates by a non-catalytic mechanism. YOD1 associates with TRAF6 in unstimulated cells but is released upon IL-1β stimulation, thereby facilitating TRAF6 auto-ubiquitination as well as NEMO/IKKγ substrate ubiquitination. Further, IL-1 triggered IKK/NF-κB signaling and induction of target genes is decreased by YOD1 overexpression and augmented after YOD1 depletion. Hence, our data define that YOD1 antagonizes TRAF6/p62-dependent IL-1 signaling to NF-κB.

The Deubiquitinase OTULIN Is an Essential Negative Regulator of Inflammation and Autoimmunity.

  • Damgaard RB
  • Cell
  • 2016 Aug 25

Literature context: #05-1308; RRID:AB_1587580 anti-Lys48


Methionine-1 (M1)-linked ubiquitin chains regulate the activity of NF-κB, immune homeostasis, and responses to infection. The importance of negative regulators of M1-linked chains in vivo remains poorly understood. Here, we show that the M1-specific deubiquitinase OTULIN is essential for preventing TNF-associated systemic inflammation in humans and mice. A homozygous hypomorphic mutation in human OTULIN causes a potentially fatal autoinflammatory condition termed OTULIN-related autoinflammatory syndrome (ORAS). Four independent OTULIN mouse models reveal that OTULIN deficiency in immune cells results in cell-type-specific effects, ranging from over-production of inflammatory cytokines and autoimmunity due to accumulation of M1-linked polyubiquitin and spontaneous NF-κB activation in myeloid cells to downregulation of M1-polyubiquitin signaling by degradation of LUBAC in B and T cells. Remarkably, treatment with anti-TNF neutralizing antibodies ameliorates inflammation in ORAS patients and rescues mouse phenotypes. Hence, OTULIN is critical for restraining life-threatening spontaneous inflammation and maintaining immune homeostasis.