Identification of LRP1B-interacting proteins and inhibition of protein kinase Cα-phosphorylation of LRP1B by association with PICK1

Recent studies show LDL receptor-related protein 1B, LRP1B as a transducer of extracellular signals. Here, we identify six interacting partners of the LRP1B cytoplasmic region by yeast two-hybrid screen and confirmed their in vivo binding by immunoprecipitation. One of the partners, PICK1 recognizes the C-terminus of LRP1B and LRP1. The cytoplasmic domains of LRP1B are phosphorylated by PKCα about 100 times more efficiently than LRP1. Binding of PICK1 inhibits phosphorylation of LRP1B, but does not affect LRP1 phosphorylation.This study presents the possibility that LRP1B participates in signal transduction which PICK1 may regulate by inhibiting PKCα phosphorylation of LRP1B.

Structured summary

MINT-6801075: Lrp1b (uniprotkb:Q9JI18) physically interacts (MI:0218) with SNTG2 (uniprotkb:Q925E0) by two hybrid (MI:0018)MINT-6801030, MINT-6801468: Lrp1b (uniprotkb:Q9JI18) physically interacts (MI:0218) with Pick1 (uniprotkb:Q80VC8) by two hybrid (MI:0018)MINT-6801284: LRP1B4 (uniprotkb:Q9JI18) physically interacts (MI:0218) with RanBPM (uniprotkb:P69566) by anti tag coimmunoprecipitation (MI:0007)MINT-6801108: Lrp1b (uniprotkb:Q9JI18) physically interacts (MI:0218) with Grb7 (uniprotkb:Q03160) by two hybrid (MI:0018)MINT-6801090: Lrp1b (uniprotkb:Q9JI18) physically interacts (MI:0218) with RanBPM (uniprotkb:P69566) by two hybrid (MI:0018)MINT-6801008: Lrp1b (uniprotkb:Q9JI18) physically interacts (MI:0218) with Jip-1b (uniprotkb:Q9WVI9-1) by two hybrid (MI:0018)MINT-6801052: Lrp1b (uniprotkb:Q9JI18) physically interacts (MI:0218) with Jip-2 (uniprotkb:Q9ERE9) by two hybrid (MI:0018)MINT-6801258, MINT-6801271: LRP1B4 (uniprotkb:Q9JI18) physically interacts (MI:0218) with Pick1 (uniprotkb:Q80VC8) by anti tag coimmunoprecipitation (MI:0007)MINT-6801244: RanBPM (uniprotkb:P69566) physically interacts (MI:0218) with mLRP4 (uniprotkb:Q8VI56) by anti tag coimmunoprecipitation (MI:0007)MINT-6801131, MINT-6801158: LRP1B4 (uniprotkb:Q9JI18) physically interacts (MI:0218) with Jip-1b (uniprotkb:Q9WVI9-1) by anti tag coimmunoprecipitation (MI:0007)MINT-6801231: PICK1 (uniprotkb:Q80VC8) physically interacts (MI:0218) with mLRP4 (uniprotkb:Q8VI56) by anti tag coimmunoprecipitation (MI:0007)MINT-6801173: Jip-1b (uniprotkb:Q9WVI9-1) physically interacts (MI:0218) with mLRP4 (uniprotkb:Q8VI56) by anti tag coimmunoprecipitation (MI:0007)     

Interactions with LC3 and polyubiquitin chains link nbr1 to autophagic protein turnover

Nbr1, a ubiquitous kinase scaffold protein, contains a PB1, and a ubiquitin-associated (UBA) domain. We show here that the nbr1 UBA domain binds to lysine-48 and -63 linked polyubiquitin-B chains. Nbr1 also binds to the autophagic effector protein LC3-A via a novel binding site. Ubiquitin-binding, but not PB1-mediated p62/SQSTM1 interaction, is required to target nbr1 to LC3 and polyubiquitin-positive bodies. Nbr1 binds additionally to proteins implicated in ubiquitin-mediated protein turnover and vesicle trafficking: ubiquitin-specific peptidases USP8, and the endosomal transport regulator p14/Robld3. Nbr1 thus contributes to specific steps in protein turnover regulation disrupted in several hereditary human diseases.

Structured summary

MINT-7034452: USP8 (uniprotkb:P40818) physically interacts (MI:0218) with NBR1 (uniprotkb:Q14596) by pull down (MI:0096)MINT-7034438: SQSTM1 (uniprotkb:Q13501) and LC3 (uniprotkb:Q9H492) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7034309: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with Ubiquitin (uniprotkb:P62988) by pull down (MI:0096)MINT-7034323: NBR1 (uniprotkb:P97432) physically interacts (MI:0218) with Ubiquitin (uniprotkb:P62988) by pull down (MI:0096)MINT-7034233: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with USP8 (uniprotkb:P40818) by two hybrid (MI:0018)MINT-7034207: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with Robld3 (uniprotkb:Q9JHS3) by two hybrid (MI:0018)MINT-7034400, MINT-7034418: NBR1 (uniprotkb:Q14596) and LC3 (uniprotkb:Q9H492) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7034167: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with Ubiquitin B (uniprotkb:Q78XY9) by two hybrid (MI:0018)MINT-7034470: NBR1 (uniprotkb:Q14596) and USP8 (uniprotkb:P40818) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7034194: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with LC3-A (uniprotkb:Q91VR7) by two hybrid (MI:0018)MINT-7034336: SQSTM1 (uniprotkb:Q13501) physically interacts (MI:0218) with Ubiquitin (uniprotkb:P62988) by pull down (MI:0096)MINT-7034375: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with LC3 (uniprotkb:Q9H492) by pull down (MI:0096)MINT-7034350: NBR1 (uniprotkb:Q14596) and Ubiquitin (uniprotkb:P62988) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7034181: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with Tmed10 (uniprotkb:Q9D1D4) by two hybrid (MI:0018)MINT-7034220: NBR1 (uniprotkb:Q14596) physically interacts (MI:0218) with ube2o (uniprotkb:Q6ZPJ3) by two hybrid (MI:0018)     

Interactions between membrane-bound cellulose synthases involved in the synthesis of the secondary cell wall

It has not yet been reported how the secondary CESA (cellulose synthase) proteins are organized in the rosette structure. A membrane-based yeast two-hybrid (MbYTH) approach was used to analyze the interactions between the CESA proteins involved in secondary cell wall synthesis of Arabidopsis and the findings were confirmed in planta by bimolecular fluorescence complementation (BiFC) assay. Results indicated that although all CESA proteins can interact with each other, only CESA4 is able to form homodimers. A model is proposed for the secondary rosette structure. The RING-motif proved not to be essential for the interaction between the CESA proteins.

Structured summary

MINT-6951243: PIP2-1 (uniprotkb:P43286) physically interacts (MI:0218) with PIP2-1 (uniprotkb:P43286) by bimolecular fluorescence complementation (MI:0809)MINT-6950816: CESA4 (uniprotkb:Q84JA6) physically interacts (MI:0218) withCESA4 (uniprotkb:Q84JA6) by membrane bound complementation assay (MI:0230)MINT-6951056, MINT-6951071, MINT-6951088, MINT-6951103: CESA7 (uniprotkb:Q9SWW6) physically interacts (MI:0218) with CESA4 (uniprotkb:Q84JA6) by bimolecular fluorescence complementation (MI:0809)MINT-6950949, MINT-6950990: CESA4 (uniprotkb:Q84JA6) physically interacts (MI:0218) with CESA8 (uniprotkb:Q8LPK5) by membrane bound complementation assay (MI:0230)MINT-6950909, MINT-6951030: CESA4 (uniprotkb:Q8LPK5) physically interacts (MI:0218) with CESA7 (uniprotkb:Q9SWW6) by membrane bound complementation assay (MI:0230)MINT-6951042: CESA4 (uniprotkb:Q84JA6) physically interacts (MI:0218) with CESA4 (uniprotkb:Q84JA6) by bimolecular fluorescence complementation (MI:0809)MINT-6951004, MINT-6951016: CESA8 (uniprotkb:Q8LPK5) physically interacts (MI:0218) with CESA7 (uniprotkb:Q9SWW6) by membrane bound complementation assay (MI:0230)MINT-6951217, MINT-6951230: CESA4 (uniprotkb:Q84JA6) physically interacts (MI:0218) with CESA8 (uniprotkb:Q8LPK5) by bimolecular fluorescence complementation (MI:0809)MINT-6951120, MINT-6951140, MINT-6951156, MINT-6951170, MINT-6951185: CESA8 (uniprotkb:Q8LPK5) physically interacts (MI:0218) withCESA7 (uniprotkb:Q9SWW6) by bimolecular fluorescence complementation (MI:0809)MINT-6951199: CESA8 (uniprotkb:Q8LPK5) physically interacts (MI:0218) withCESA8 (uniprotkb:Q8LPK5) by bimolecular fluorescence complementation (MI:0809)     

FKBP25, a novel regulator of the p53 pathway, induces the degradation of MDM2 and activation of p53

The p53 tumour suppressor protein is tightly controlled by the E3 ubiquitin ligase, mouse double minute 2 (MDM2), but maintains MDM2 expression as part of a negative feedback loop. We have identified the immunophilin, 25 kDa FK506-binding protein (FKBP25), previously shown to be regulated by p53-mediated repression, as an MDM2-interacting partner. We show that FKBP25 stimulates auto-ubiquitylation and proteasomal degradation of MDM2, leading to the induction of p53. Depletion of FKBP25 by siRNA leads to increased levels of MDM2 and a corresponding reduction in p53 and p21 levels. These data are consistent with the idea that FKBP25 contributes to regulation of the p53-MDM2 negative feedback loop.

Structured summary

MINT-6823686:MDM2 (uniprotkb:Q00987) physically interacts (MI:0218) with FKBP25 (uniprotkb:Q00688) by anti bait coimmunoprecipitation (MI:0006)MINT-6823707, MINT-6823722:MDM2 (uniprotkb:Q00987) physically interacts (MI:0218) with FKBP25 (uniprotkb:Q62446) by pull down (MI:0096)MINT-6823775:P53 (uniprotkb:Q04637) physically interacts (MI:0218) with MDM2 (uniprotkb:Q00987) by anti bait coimmunoprecipitation (MI:0006)MINT-6823735, MINT-6823749:FKBP25 (uniprotkb:Q62446) binds (MI:0407) to MDM2 (uniprotkb:Q00987) by pull down (MI:0096)MINT-6823761:Ubiquitin (UNIPROTKB:62988)P physically interacts (MI:0218) with MDM2 (uniprotkb:Q00987) by pull down (MI:0096)MINT-6823669:MDM2 (uniprotkb:Q00987) physically interacts (MI:0218) with FKBP25 (uniprotkb:Q00688) by two hybrid (MI:0018)     

CAP (Cbl associated protein) regulates receptor-mediated endocytosis

CAP (c-Cbl associated protein)/ponsin belongs to a family of adaptor proteins implicated in cell adhesion and signaling. Here we show that CAP binds to and co-localizes with the essential endocytic factor dynamin. We demonstrate that CAP promotes the formation of dynamin-decorated tubule like structures, which are also coated with actin filaments. Accordingly, we found that the expression of CAP leads to the inhibition of dynamin-mediated endocytosis and increases EGFR stability. Thus, we suggest that CAP may coordinate the function of dynamin with the regulation of the actin cytoskeleton during endocytosis.

Structured summary:

MINT-6804322: CAP (uniprotkb:Q9BX66) physically interacts (MI:0218) with Cbl (uniprotkb:Q8K4S7) and dynamin 2 (uniprotkb:P39052) by pull down (MI:0096)MINT-6804285: CAP (uniprotkb:Q9BX66) physically interacts (MI:0218) with FAK (uniprotkb:O35346), vinculin (uniprotkb:P85972) and dynamin 2 (uniprotkb:P39052) by pull down (MI:0096)MINT-6804245, MINT-6804259, MINT-6804272: CAP (uniprotkb:Q9BX66) physically interacts (MI:0218) with dynamin 2 (uniprotkb:P39052) by pull down (MI:0096)MINT-6804344: CAP (uniprotkb:Q9BX66) physically interacts (MI:0218) with dynamin 2 (uniprotkb:P50570) by anti tag coimmunoprecipitation (MI:0007)MINT-6804371: dynamin 1 (uniprotkb:P21575) physically interacts (MI:0218) with CAP (uniprotkb:O35413) by anti bait coimmunoprecipitation (MI:0006)MINT-6804446, MINT-6804464: F-actin (uniprotkb:P60709), CAP (uniprotkb:Q9BX66) and dynamin 2 (uniprotkb:P50570) colocalize (MI:0403) by fluorescence microscopy (MI:0416)     

NMDA receptors interact with flotillin-1 and -2, lipid raft-associated proteins

N-methyl-d-aspartate receptors (NMDARs) mediate excitatory synaptic transmission in the brain. Here we demonstrate interactions between the NR2A and NR2B subunits of NMDARs with flotillin-1 (flot-1), a lipid raft-associated protein. When mapped, analogous regions in the far distal C-termini of NR2A and NR2B mediate binding to flot-1, and the prohibitin homology domain of flot-1 contains binding sites for NR2A and NR2B. Although NR2B can also directly bind to flot-2 via a separate region of its distal C-terminus, NMDARs were significantly more colocalized with flot-1 than flot-2 in cultured hippocampal neurons. Overall, this study defines a novel interaction between NMDARs and flotillins.

Structured summary

MINT-7013094: NR2A (uniprotkb:Q00959), NR2B (uniprotkb:Q00960) and Flot2 (uniprotkb:Q9Z2S9) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7013147: Flot1 (uniprotkb:Q9Z1E1) physically interacts (MI:0218) with NR2A (uniprotkb:Q00959) by anti bait coimmunoprecipitation (MI:0006)MINT-7013189: Flot1 (uniprotkb:Q9Z1E1) physically interacts (MI:0218) with Flot2 (uniprotkb:Q9Z2S9) by anti bait coimmunoprecipitation (MI:0006)MINT-7013033: NR2A (uniprotkb:Q00959) physically interacts (MI:0218) with Flot1 (uniprotkb:Q9Z1E1) by two hybrid (MI:0018)MINT-7013178: NR1 (uniprotkb:P35439) physically interacts (MI:0218) with Flot2 (uniprotkb:Q9Z2S9) by anti bait coimmunoprecipitation (MI:0006)MINT-7013197, MINT-7013210: NR2B (uniprotkb:Q00960) and NR2A (uniprotkb:Q00959) physically interact (MI:0218) with Flot2 (uniprotkb:Q9Z2S9) by anti bait coimmunoprecipitation (MI:0006)MINT-7013002: NR2B (uniprotkb:Q00960) physically interacts (MI:0218) with Flot1 (uniprotkb:O08917) by two hybrid (MI:0018)MINT-7013117: Flot1 (uniprotkb:Q9Z1E1), NR2B (uniprotkb:Q00960) and NR2A (uniprotkb:Q00959) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7013171: NR1 (uniprotkb:P35439) physically interacts (MI:0218) with Flot1 (uniprotkb:Q9Z1E1) by anti bait coimmunoprecipitation (MI:0006)MINT-7013017: NR2A (uniprotkb:Q00959) physically interacts (MI:0218) with Flot1 (uniprotkb:O08917) by two hybrid (MI:0018)MINT-7013054: NR2B (uniprotkb:Q00960) physically interacts (MI:0218) with Flot1 (uniprotkb:Q9Z1E1) by two hybrid (MI:0018)MINT-7013129: Flot1 (uniprotkb:Q9Z1E1) physically interacts (MI:0218) with NR2B (uniprotkb:Q00960) by anti bait coimmunoprecipitation (MI:0006)MINT-7013155: NR1 (uniprotkb:P35439) physically interacts (MI:0218) with NR2B (uniprotkb:Q00960) by anti bait coimmunoprecipitation (MI:0006)MINT-7013074: NR2B (uniprotkb:Q00960) physically interacts (MI:0218) with Flot2 (uniprotkb:Q9Z2S9) by two hybrid (MI:0018)MINT-7013162: NR1 (uniprotkb:P35439) physically interacts (MI:0218) with NR2A (uniprotkb:Q00959) by anti bait coimmunoprecipitation (MI:0006)     

Transcriptional activation of hypoxia-inducible factor-1α by HDAC4 and HDAC5 involves differential recruitment of p300 and FIH-1

The interplay between hypoxia-inducible factor-1α (HIF-1α) and histone deacetylase (HDACs) have been well studied; however, the mechanism of cross-talk is unclear. Here, we investigated the roles of HDAC4 and HDAC5 in the regulation of HIF-1α function and its associated mechanisms. HDAC4 and HDAC5 enhanced transactivation by HIF-1α without stabilizing HIF-1α. HDAC4 and HDAC5 physically associated with HIF-1α through the inhibitory domain (ID) that is the binding site for factor inhibiting HIF-1 (FIH-1). In the presence of these HDACs, binding of HIF-1α to FIH-1 decreased, whereas binding to p300 increased. These results indicate that HDAC4 and HDAC5 increase the transactivation function of HIF-1α by promoting dissociation of HIF-1α from FIH-1 and association with p300.

Structured summary:

MINT-6802187:HIF1 alpha (uniprotkb:Q16665) physically interacts (MI:0218) with FIH1 (uniprotkb:Q9NWT6) by anti bait coimmunoprecipitation (MI:0006)MINT-6802058:HIF1 alpha (uniprotkb:Q16665) physically interacts (MI:0218) with HDAC4 (uniprotkb:P56524) by pull down (MI:0096)MINT-6802021:HIF1 alpha (uniprotkb:Q61221) physically interacts (MI:0218) with HDAC4 (uniprotkb:P56524) by anti bait coimmunoprecipitation (MI:0006)MINT-6802036:HIF1 alpha (uniprotkb:Q61221) physically interacts (MI:0218) with HDAC5 (uniprotkb:Q9UQL6) by anti bait coimmunoprecipitation (MI:0006)MINT-6802102:HIF1 alpha (uniprotkb:Q16665) physically interacts (MI:0218) with HDAC5 (uniprotkb:Q9UQL6) by pull down (MI:0096)MINT-6802121, MINT-6802156:P300 (uniprotkb:Q09472) physically interacts (MI:0218) with HIF1 alpha (uniprotkb:Q16665) by anti bait coimmunoprecipitation (MI:0006)     

β2-Chimaerin binds to EphA receptors and regulates cell migration

Ephrins and Eph receptors have key roles in regulation of cell migration during development. We found that the RacGAP β2-chimaerin (chimerin) bound to EphA2 and EphA4 and inactivated Rac1 in response to ephrinA1 stimulation. EphA4 bound to β2-chimaerin through its kinase domain and promoted binding of Rac1 to β2-chimaerin. In addition, knockdown of endogenous β2-chimaerin blocked ephrinA1-induced suppression of cell migration. These results suggest that β2-chimaerin is activated by EphA receptors and mediates the EphA receptor-dependent regulation of cell migration.

Structured summary

MINT-7013428: EphA1 (uniprotkb:Q60750) physically interacts (MI:0218) with Chimaerin beta 2 (uniprotkb:Q80XD1-2) and EphA4 (uniprotkb:O08542) by anti tag coimmunoprecipitation (MI:0007)MINT-7013515: Chimaerin beta 2 (uniprotkb:Q80XD1-2) physically interacts (MI:0218) with Rac1 (uniprotkb:P63001) by anti tag coimmunoprecipitation (MI:0007)MINT-7013410: EphA1 (uniprotkb:Q60750) physically interacts (MI:0218) with Chimaerin beta 1 (uniprotkb:Q80XD1-1) and EphA4 (uniprotkb:O08542) by anti tag coimmunoprecipitation (MI:0007)MINT-7013503: Chimaerin beta 1 (uniprotkb:Q80XD1-1) physically interacts (MI:0218) with EphA4 (uniprotkb:O08542) by anti tag coimmunoprecipitation (MI:0007)MINT-7013472: Chimaerin beta 2 (uniprotkb:Q80XD1-2) physically interacts (MI:0218) with EphA2 (uniprotkb:O43921) by anti tag coimmunoprecipitation (MI:0007)MINT-7013450: EphA1 (uniprotkb:Q60750) physically interacts (MI:0218) with EphA2 (uniprotkb:O43921) and Chimaerin beta 2 (uniprotkb:P52757-1) by anti tag coimmunoprecipitation (MI:0007)MINT-7013491: Chimaerin beta 2 (uniprotkb:Q80XD1-2) physically interacts (MI:0218) with EphA4 (uniprotkb:O08542) by anti tag coimmunoprecipitation (MI:0007)     

Cereblon is expressed in the retina and binds to voltage-gated chloride channels

The function of the central nervous system depends on the correct regulation of ion channels by interacting proteins. Here, we identified cereblon as a new interactor of the voltage-gated chloride channel ClC-2. A distal region of the ClC-2 C-terminus interacts with the Lon domain of cereblon. Cereblon is expressed in several brain regions including the retina. There, we detected cereblon in nuclear and synaptic layers and localized the protein in the same subcellular region of bipolar cell bodies previously reported to express ClC-2. Our data suggest that cereblon might be associated with voltage-gated chloride channels in the central nervous system.

Structured summary

MINT-6823070: CIC-2 (uniprotkb:O54822) physically interacts (MI:0218) with CRBN (uniprotkb:Q0P564) by two hybrid (MI:0018) MINT-6823160, MINT-6823197: CIC-2 (uniprotkb:O54822) physically interacts (MI:0218) with CRBN (uniprotkb:Q56AP7) by pull down (MI:0096) MINT-6823105: CIC-2 (uniprotkb:O54822) physically interacts (MI:0218) with IK (uniprotkb:A4FUY8) by two hybrid (MI:0018)     

Interaction of the human somatostatin receptor 3 with the multiple PDZ domain protein MUPP1 enables somatostatin to control permeability of epithelial tight junctions

The presence of heterotrimeric G-proteins at epithelial tight junctions suggests that these cellular junctions are regulated by so far unknown G-protein coupled receptors. We identify here an interaction between the human somatostatin receptor 3 (hSSTR3) and the multiple PDZ protein MUPP1. MUPP1 is a tight junction scaffold protein in epithelial cells, and as a result of the interaction with MUPP1 the hSSTR3 is targeted to tight junctions. Interaction with MUPP1 enables the receptor to regulate transepithelial permeability in a pertussis toxin sensitive manner, suggesting that hSSTR3 can activate G-proteins locally at tight junctions.

Structured summary:

MINT-6800756, MINT-6800770: MUPP1 (uniprotkb:O75970) and hSSTR3 (uniprotkb:P32745) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-6800587:hSSTR3 (uniprotkb:P32745) physically interacts (MI:0218) with MUPP1 (uniprotkb:O55164) by pull down (MI:0096)MINT-6800562:hSSTR3 (uniprotkb:P32745) physically interacts (MI:0218) with MUPP1 (uniprotkb:O75970) by two hybrid (MI:0018)MINT-6800622:hSSTR3 (uniprotkb:P32745) physically interacts (MI:0218) with PIST (uniprotkb: Q9HD26), Hsp70 (uniprotkb:P08107), Maguk p55 (uniprotkb: Q8N3R9), MAGI3 (uniprotkb:Q5TCQ9), ZO-2 (uniprotkb:Q9UDY2), ZO-1 (uniprotkb:Q07157) and MUPP1 (uniprotkb:O55164) by pull down (MI:0096)MINT-6800607, MINT-6801122:hSSTR3 (uniprotkb:P32745) physically interacts (MI:0218) with MUPP1 (uniprotkb:O75970) by anti bait coimmunoprecipitation (MI:0006)     

Programmed cell death-2 isoform1 is ubiquitinated by parkin and increased in the substantia nigra of patients with autosomal recessive Parkinson’s disease

Mutations in parkin gene are responsible for autosomal recessive Parkinson’s disease (ARPD) and its loss-of-function is assumed to affect parkin ubiquitin ligase activity. Accumulation of its substrate may induce dopaminergic neurodegeneration in the substantia nigra (SN) of ARPD. Here, we show that parkin interacts with programmed cell death-2 isoform 1 (PDCD2-1) and promotes its ubiquitination. Furthermore, accumulation of PDCD2-1 was found in the SN of ARPD as well as in sporadic PD, suggesting that common failure of the ubiquitin-proteasome system is associated with neuronal death in both ARPD and sporadic PD.Structured summary:MINT-6805975, MINT-6806032, MINT-6806051, MINT-6806070:PDCD2 (uniprotkb:Q16342) physically interacts (MI:0218) with Parkin (uniprotkb:O60260) by anti tag coimmunoprecipitation (MI:0007)MINT-6805947:Parkin (uniprotkb:O60260) physically interacts (MI:0218) with PDCD2 (uniprotkb:Q16342) by two hybrid (MI:0018)MINT-6806000: PDCD2 (uniprotkb:Q16342) physically interacts (MI:0218) with ubiquitin (uniprotkb:P62988) by anti tag coimmunoprecipitation (MI:0007).     

Requirement for microtubule integrity in the SOCS1-mediated intracellular dynamics of HIV-1 Gag

Suppressor of cytokine signaling 1 (SOCS1) is a recently identified host factor that positively regulates the intracellular trafficking and stability of HIV-1 Gag. We here examine the molecular mechanism by which SOCS1 regulates intercellular Gag trafficking and virus particle production. We find that SOCS1 colocalizes with Gag along the microtubule network and promotes microtubule stability. SOCS1 also increases the amount of Gag associated with microtubules. Both nocodazole treatment and the expression of the microtubule-destabilizing protein, stathmin, inhibit the enhancement of HIV-1 particle production by SOCS1. SOCS1 facilitates Gag ubiquitination and the co-expression of a dominant-negative ubiquitin significantly inhibits the association of Gag with microtubules. We thus propose that the microtubule network plays a role in SOCS1-mediated HIV-1 Gag transport and virus particle formation.

Structured summary

MINT-7014185: Gag (uniprotkb:P05888) and SOCS1 (uniprotkb:O15524) colocalize (MI:0403) by cosedimentation (MI:0027)MINT-7014239: Cullin 2 (uniprotkb:Q13617) physically interacts (MI:0218) with RelA (uniprotkb:Q04206), RBX1 (uniprotkb:P62877), SOCS1 (uniprotkb:O15524), elongin B (uniprotkb:Q15369) and elongin C (uniprotkb:Q15370) by pull-down (MI:0096)MINT-7014046: gag (uniprotkb:P05888), SOCS1 (uniprotkb:O15524) and tubulin alpha (uniprotkb:Q13748) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7014269: tubulin alpha (uniprotkb:Q13748) physically interacts (MI:0218) with Gag (uniprotkb:P05888) by anti tag coimmunoprecipitation (MI:0007)MINT-7014036: tubulin alpha (uniprotkb:Q13748) and SOCS1 (uniprotkb:O15524) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7014201: Cullin 2 (uniprotkb:Q13617) physically interacts (MI:0218) with RBX1 (uniprotkb:P62877), SOCS1 (uniprotkb:O15524), elongin B (uniprotkb:Q15369) and elongin C (uniprotkb:Q15370) by pull-down (MI:0096)MINT-7014257: Gag (uniprotkb:P05888) physically interacts (MI:0218) with Ubiquitin (uniprotkb:P62988) by anti tag coimmunoprecipitation (MI:0007)MINT-7014221: Cullin 2 (uniprotkb:Q13617) physically interacts (MI:0218) with Gag (uniprotkb:P05888), elongin C (uniprotkb:Q15370), elongin B (uniprotkb:Q15369), SOCS1 (uniprotkb:O15524) and RBX1 (uniprotkb:P62877) by pull-down (MI:0096)     

Nuclear localization of the pre-mRNA associating protein THOC7 depends upon its direct interaction with Fms tyrosine kinase interacting protein (FMIP)

THOC7 and Fms-interacting protein (FMIP) are members of the THO complex that associate with the mRNA export apparatus. FMIP is a nucleocytoplasmic shuttling protein with a nuclear localization signal (NLS), whereas THOC7 does not contain a typical NLS motif. We show here that THOC7 (50-137, amino acid numbers) binds to the N-terminal portion (1-199) of FMIP directly. FMIP is detected mainly in the nucleus. In the absence of exogenous FMIP, THOC7 resides mainly in the cytoplasm, while in the presence of FMIP, THOC7 is transported into the nucleus with FMIP. Furthermore, THOC7 lacking the FMIP binding site does not co-localize with FMIP, indicating that THOC7/FMIP interaction is required for nuclear localization of THOC7.

Structured summary

MINT-6799962, MINT-6799973, MINT-6800005: THOC7 (uniprotkb:Q6I9Y2) physically interacts (MI:0218) with THOC5 (uniprotkb:Q13769) by pull down (MI:0096)MINT-6800108: FMIP (uniprotkb:Q13769) and THOC7 (uniprotkb: Q6I9Y2) co-localize (MI:0403) by fluorescence microscopy (MI:0416)MINT-6800052: FMIP (uniprotkb:Q13769) physically interacts (MI:0218) with THOC1 (uniprotkb: Q96FV9) by anti tag coimmunoprecipitation (MI:0007)MINT-6800022: THOC7 (uniprotkb:Q6I9Y2) physically interacts (MI:0218) with FMIP (uniprotkb:Q6DFL5) by pull down (MI:0096)MINT-6799989: THOC7 (uniprotkb:Q6I9Y2) binds (MI:0407) to FMIP (uniprotkb:Q13769) by pull down (MI:0096)MINT-6800071, MINT-6800089: FMIP (uniprotkb:Q13769) physically interacts (MI:0218) with THOC7 (uniprotkb:Q6I9Y2) and THOC1 (uniprotkb:Q96FV9) by anti tag coimmunoprecipitation (MI:0007)     

Phosphorylation of the eukaryotic initiation factor 3f by cyclin-dependent kinase 11 during apoptosis

eIF3f is a subunit of eukaryotic initiation factor 3 (eIF3). We previously showed that eIF3f is phosphorylated by cyclin dependent kinase 11 (CDK11^p46) which is an important effector in apoptosis. Here, we identified a second eIF3f phosphorylation site (Thr119) by CDK11^p46 during apoptosis. We demonstrated that eIF3f is directly phosphorylated by CDK11^p46 in vivo. Phosphorylation of eIF3f plays an important role in regulating its function in translation and apoptosis. Phosphorylation of eIF3f enhances the association of eIF3f with the core eIF3 subunits during apoptosis. Our data suggested that eIF3f may inhibit translation by increasing the binding to the eIF3 complex during apoptosis.

Structured summary

MINT-6948874: EIF3b (uniprotkb:P55884) physically interacts (MI:0218) with EIF3f (uniprotkb:O00303) by anti bait coimmunoprecipitation (MI:0006)MINT-6948891: EIF3b (uniprotkb:P55884) physically interacts (MI:0218) with EIF3c (uniprotkb:Q99613), EIF3a (uniprotkb:Q14152) and EIF3f (uniprotkb:O00303) by anti bait coimmunoprecipitation (MI:0006)MINT-6948836, MINT-6948849, MINT-6948862: CDK11p46 (uniprotkb:P21127) phosphorylates (MI:0217) EIF3f (uniprotkb:O00303) by protein kinase assay (MI:0424)     

BS69 negatively regulates the canonical NF-κB activation induced by Epstein-Barr virus-derived LMP1

Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) activates NF-κB signaling pathways through the two C-terminal regions, CTAR1 and CTAR2. BS69 has previously been shown to be involved in LMP1-induced c-Jun N-terminal kinase activation through CTAR2 by interacting with tumor necrosis factor (TNFR) receptor-associated factor 6. In the present study, our manipulation of BS69 expression clearly indicates that BS69 negatively regulates LMP1-mediated NF-κB activation and up-regulates IL-6 mRNA expression and IκB degradation. Our immunoprecipitation experiments suggest that BS69 decreases complex formation between LMP1 and TNFR-associated death domain protein (TRADD).

Structured summary

MINT-7032462: LMP1 (uniprotkb:P03230) physically interacts (MI:0218) with TRADD (uniprotkb:Q15628) by anti bait coimmunoprecipitation (MI:0006)MINT-7032451: BS69 (uniprotkb:Q15326) and LMP1 (uniprotkb:P03230) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7032478: LMP1 (uniprotkb:P03230) physically interacts (MI:0218) with BRAM1 (uniprotkb:Q15326) by anti bait coimmunoprecipitation (MI:0006)     

Interaction specificity of Arabidopsis 14-3-3 proteins with phototropin receptor kinases

Phototropin receptor kinases play an important role in optimising plant growth in response to blue light. Much is known regarding their photochemical reactivity, yet little progress has been made to identify downstream signalling components. Here, we isolated several interacting proteins for Arabidopsis phototropin 1 (phot1) by yeast two-hybrid screening. These include members of the NPH3/RPT2 (NRL) protein family, proteins associated with vesicle trafficking, and the 14-3-3 lambda (λ) isoform from Arabidopsis. 14-3-3λ and phot1 were found to colocalise and interact in vivo. Moreover, 14-3-3 binding to phot1 was limited to non-epsilon 14-3-3 isoforms and was dependent on key sites of receptor autophosphorylation. No 14-3-3 binding was detected for Arabidopsis phot2, suggesting that 14-3-3 proteins are specific to phot1 signalling.

Structured summary

MINT-7146953: PHOT1 (uniprotkb:O48963) physically interacts (MI:0915) with ARF7 (uniprotkb:Q9LFJ7) by two hybrid (MI:0018)MINT-7147335: PHOT1 (uniprotkb:O48963) physically interacts (MI:0914) with 14-3-3 phi (uniprotkb:P46077) by far Western blotting (MI:0047)MINT-7146854: PHOT1 (uniprotkb:O48963) physically interacts (MI:0915) with RPT2 (uniprotkb:Q682S0) by two hybrid (MI:0018)MINT-7147215: PHOT1 (uniprotkb:O48963) physically interacts (MI:0914) with 14-3-3 lambda (uniprotkb:P48349) by anti tag coimmunoprecipitation (MI:0007)MINT-7147044, MINT-7147185, MINT-7147200, MINT-7147413: PHOT1 (uniprotkb:O48963) physically interacts (MI:0914) with 14-3-3 lambda (uniprotkb:P48349) by far Western blotting (MI:0047)MINT-7146983: PHOT1 (uniprotkb:O48963) physically interacts (MI:0915) with 14-3-3 lambda (uniprotkb:P48349) by two hybrid (MI:0018)MINT-7146871: PHOT1 (uniprotkb:O48963) physically interacts (MI:0915) with NPH3-like (uniprotkb:Q9S9Q9) by two hybrid (MI:0018)MINT-7146905: PHOT1 (uniprotkb:O48963) physically interacts (MI:0915) with ARF2 (uniprotkb:Q9M1P5) by two hybrid (MI:0018)MINT-7147364: PHOT1 (uniprotkb:O48963) physically interacts (MI:0914) with 14-3-3 upsilon (uniprotkb:P42645) by far Western blotting (MI:0047)MINT-7147234: PHOT1 (uniprotkb:O48963) physically interacts (MI:0914) with 14-3-3 kappa (uniprotkb:P48348) by far Western blotting (MI:0047)     

Deletion of Swm2p selectively impairs trimethylation of snRNAs by trimethylguanosine synthase (Tgs1p)

The 5′ cap trimethylation of small nuclear (snRNAs) and several nucleolar RNAs (snoRNAs) by trimethylguanosine synthase 1 (Tgs1p) is required for efficient pre-mRNA splicing. The previously uncharacterised protein Swm2p interacted with Tgs1p in yeast two-hybrid screens. In the present study we show that Swm2p interacts with the N-terminus of Tgs1p and its deletion impairs pre-mRNA splicing and pre-rRNA processing. The trimethylation of spliceosomal snRNAs and the U3 snoRNA, but not other snoRNAs, was abolished in the absence of Swm2p, indicating that Swm2p is required for a substrate-specific activity of Tgs1p.

Structured summary

MINT-7949608: p53 (uniprotkb:P02340) physically interacts (MI:0915) with large T-antigen (uniprotkb:P03070) by two-hybrid (MI:0018)MINT-7949574: swm2 (uniprotkb:P40342) physically interacts (MI:0915) with swm2 (uniprotkb:P40342) by pull down (MI:0096)MINT-7949556: swm2 (uniprotkb:P40342) physically interacts (MI:0915) with TGS1 (uniprotkb:Q12052) by pull down (MI:0096)MINT-7949587: swm2 (uniprotkb:P40342) physically interacts (MI:0915) with tgs1 (uniprotkb:Q12052) by two-hybrid (MI:0018)MINT-7949641: nop1 (uniprotkb:P15646) colocalizes (MI:0403) with TGS1 (uniprotkb:Q12052) by fluorescence microscopy (MI:0416)MINT-7949627: swm2 (uniprotkb:P40342) and nop1 (uniprotkb:P15646) colocalize (MI:0403) by fluorescence microscopy (MI:0416)MINT-7949540: swm2 (uniprotkb:P40342) physically interacts (MI:0915) with TGS1 (uniprotkb:Q12052) by tandem affinity purification (MI:0676)     

Serine 62 is a phosphorylation site in folliculin, the Birt-Hogg-Dubé gene product

Recently, it was reported that the product of Birt-Hogg-Dubé syndrome gene (folliculin, FLCN) is directly phosphorylated by 5′-AMP-activated protein kinase (AMPK). In this study, we identified serine 62 (Ser62) as a phosphorylation site in FLCN and generated an anti-phospho-Ser62-FLCN antibody. Our analysis suggests that Ser62 phosphorylation is indirectly up-regulated by AMPK and that another residue is directly phosphorylated by AMPK. By binding with FLCN-interacting proteins (FNIP1 and FNIP2/FNIPL), Ser62 phosphorylation is increased. A phospho-mimic mutation at Ser62 enhanced the formation of the FLCN-AMPK complex. These results suggest that function(s) of FLCN-AMPK-FNIP complex is regulated by Ser62 phosphorylation.

Structured summary

MINT-7298145, MINT-7298166: Flcn (uniprotkb:Q76JQ2) physically interacts (MI:0915) with AMPK alpha 1 (uniprotkb:P54645) by anti tag coimmunoprecipitation (MI:0007)MINT-7298267: AMPK alpha 1 (uniprotkb:Q13131) phosphorylates (MI:0217) tsc2 (uniprotkb:P49816) by protein kinase assay (MI:0424)MINT-7298182: FNIP1 (uniprotkb:Q8TF40) physically interacts (MI:0915) with Flcn (uniprotkb:Q76JQ2) by anti tag coimmunoprecipitation (MI:0007)MINT-7298132: AMPK alpha 1 (uniprotkb:Q13131) phosphorylates (MI:0217) Flcn (uniprotkb:Q76JQ2) by protein kinase assay (MI:0424)MINT-7298229: FNIPL (uniprotkb:Q9P278) physically interacts (MI:0915) with Flcn (uniprotkb:Q76JQ2) by anti tag coimmunoprecipitation (MI:0007)