Biochemical Journal (BIOCHEM J )

Publisher: Biochemical Society (Great Britain), Portland Press

Description

The Biochemical Journal publishes over 7000 pages of high-quality scientific information every year. The papers are carefully selected by an international editorial board to cover all aspects of biochemistry, and cell and molecular biology. The journal features regular papers, authoritative reviews, research communications (short articles reporting novel and significant findings) and correspondence.

Impact factor 4.78

  • Hide impact factor history
     
    Impact factor
  • 5-year impact
    5.02
  • Cited half-life
    0.00
  • Immediacy index
    1.19
  • Eigenfactor
    0.08
  • Article influence
    1.94
  • Website
    Biochemical Journal website
  • Other titles
    Biochemical journal (London, England: 1984), Biochemical journal
  • ISSN
    0264-6021
  • OCLC
    10379627
  • Material type
    Periodical, Internet resource
  • Document type
    Journal / Magazine / Newspaper, Internet Resource

Publisher details

Portland Press

  • Pre-print
    • Author cannot archive a pre-print version
  • Post-print
    • Author cannot archive a post-print version
  • Restrictions
    • 6 months embargo
  • Conditions
    • On author's personal website or institutional repository
    • Accepted version may be placed on PubMed Central and Europe PMC after 12 months from publication
    • Must link to journal website
    • Published source must be acknowledged ('The final version of record is available at [insert Journals URL]')
    • Publisher's version/PDF cannot be used
    • Must link to journal website, e.g. with a DOI
  • Classification
    ​ white

Publications in this journal

  • [Show abstract] [Hide abstract]
    ABSTRACT: ESCRT-III mediated membrane invagination and scission is a critical step in MVB sorting of ubiquitinated membrane receptors and generally thought to be required for degradation of these receptors in lysosomes. The adaptor protein ALIX is critically involved in multiple ESCRT-III-mediated membrane remodeling processes in mammalian cells. However, ALIX knockdown does not inhibit degradation of activated EGFR in mammalian cell lines, leading to a widely held notion that ALIX is not critically involved in MVB sorting of ubiquitinated membrane receptors in mammalian cells. In this study, we demonstrate that despite its non-essential roles in degradation of activated EGFR, ALIX plays a critical role in MVB sorting and silencing of activated EGFR. EGF stimulation of mammalian cell lines induces ALIX interaction with ubiquitinated EGFR through the ALIX V domain and increases ALIX association with membrane-bound CHMP4 through the ALIX Bro1 domain. Under both continuous and pulse-chase EGF stimulation conditions, inhibition of ALIX interaction with membrane-bound CHMP4, inhibition of ALIX dimerization through the V domain or ALIX knockdown dramatically inhibits MVB sorting of activated EGFR and promotes sustained activation of ERK1/2. Under the continuous EGF stimulation conditions, these cell treatments also retard degradation of activated EGFR. These findings indicate that ALIX is critically involved in MVB sorting of ubiquitinated membrane receptors in mammalian cells.
    Biochemical Journal 12/2014;
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    ABSTRACT: High Ca2+ content in the Golgi apparatus (Go) is essential for protein processing and sorting. Additionally, Go can shape the cytosolic Ca2+ signals by releasing or sequestering Ca2+. We generated two new aequorin-based Ca2+ probes to specifically measure Ca2+ in the cis-/cis to medial Golgi (cGo) or the trans-Golgi (tGo). Ca2+ homeostasis in these compartments and in the ER has been studied and compared. Moreover, the relative size of each subcompartment was estimated from aequorin consumption. We find that the cGo accumulates Ca2+ to high concentrations (150-300 µM) through the sarco-endoplasmic reticulum Ca2+ ATPase (SERCA). The tGo, in turn, is divided into two subcompartments, tGo1 and tGo2. The subcompartment tGo1 contains 20% of the aequorin and has a high [Ca2+] inside; Ca2+ is accumulated in this subcompartment via the Secretory Pathway Ca2+-ATPase 1 (SPCA1) at very high affinity (K50 = 30 nM). The subcompartment tGo2 contains 80% of aequorin, has a lower [Ca2+] and no SPCA1 activity; Ca2+ uptake happens through SERCA and is slower than in tGo1. The two tGo subcompartments, tGo1 and tGo2, are diffusionally isolated. Inositol-trisphosphate mobilizes Ca2+ from cGo and tGo2, but not from tGo1, whereas caffeine releases Ca2+ from all the Golgi regions, and NAADP and cADPR from none.
    Biochemical Journal 12/2014;
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    ABSTRACT: A decade ago, a Motif at N-terminus with Eight-Cysteines or in short MANEC was defined as a new protein domain family. This domain is found exclusively in the N-terminus of >400 multi-domain type-1 transmembrane proteins from animals. Despite the large number of MANEC-containing proteins, only one has been characterized at the protein level: hepatocyte growth factor activator inhibitor-1 (HAI-1). HAI-1 is an essential protein, as knockout mice die in utero due to placental defects. HAI-1 is an inhibitor of matriptase, hepsin and hepatocyte growth factor activator, all serine proteases with important roles in epithelial development, cell growth and homeostasis. Dysregulation of these proteases has been causatively implicated in pathological conditions such as skin diseases and cancer. Detailed functional understanding of HAI-1 and other MANEC-containing proteins is hampered by the lack of structural information on MANEC. Although many MANEC sequences exist, sequence-based database searches fail to predict structural homology. Here we present the NMR solution structure of the MANEC domain from HAI-1, the first three-dimensional structure from the MANEC domain family. Unexpectedly, MANEC is a new subclass of the PAN/apple domain family, with its own unifying features, such as two additional disulfide bonds, two extended loop regions and additional α-helical elements. As shown for other PAN/apple domain-containing proteins, we propose a similar active role of the MANEC domain in intramolecular and intermolecular interactions. The structure provides a tool for the further elucidation of HAI-1 function as well as a reference for the study of other MANEC-containing proteins.
    Biochemical Journal 12/2014;
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    ABSTRACT: FoxO1, which is upregulated during early stages of diet-induced leptin resistance, directly interacts with STAT3 and prevents STAT3 from binding to SP1-POMC promoter complex, and thereby inhibits STAT3-mediated regulation of POMC transcription. FoxO1 also binds directly to the POMC promoter and negatively regulates its transcription. This study aims to understand the relative contribution of the two interactions in regulating POMC expression. We studied the structural requirement of FoxO1 for its interaction with STAT3 and POMC promoter, and tested the inhibitory action of FoxO1 mutants by using biochemical assays, molecular biology and computer modeling. FoxO1 mutant with deletion of residues Ala137-Leu160 failed to bind to STAT3 or inhibit STAT3-mediated POMC activation, although its binding to the POMC promoter was unaffected. Further analysis mapped Gly140-Leu160 to be critical for STAT3 binding. The identified region Gly140-Leu160 was conserved among mammalian FoxO1 proteins, and showed high degree of sequence identity with FoxO3, but not FoxO4. Consistently, FoxO3 could interact with STAT3 and inhibit POMC promoter activity, while FoxO4 could not bind to STAT3 or affect POMC promoter activity. We further identified that five residues (Gln145, Arg147, Lys148, Arg153, Arg154) in FoxO1 were necessary in FoxO1-STAT3 interaction, and mutation of these residues abolished its interaction with STAT3 and inhibition of POMC promoter activity. Finally, a FoxO1-STAT3 interaction interface model generated by computational docking simulations confirmed that the identified residues of FoxO1 were in close proximity to STAT3. These results show that FoxO1 inhibits STAT3-mediated leptin signaling through direct interaction with STAT3.
    Biochemical Journal 12/2014;
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    ABSTRACT: The sigma-1 receptor (S1R) is a membrane 223-amino acid protein that resides in the endoplasmic reticulum and the plasma membrane of some mammalian cells. The S1R is regulated by a various synthetic molecules including (+)-pentazocine, cocaine and haloperidol, and endogenous molecules such as sphingosine, dimethyltryptamine and dehydroepiandrosterone. Ligand regulated protein chaperone functions linked to oxidative stress and neurodegenerative disorders such as ALS and neuropathic pain have been attributed to the S1R. Several client proteins that interact with S1R have been identified including various types of ion channels and G-protein coupled receptors. When S1R constructs containing C-terminal monomeric GFP2 and YFP fusions were co-expressed in COS-7 cells and subjected to FRET spectrometry analysis, monomers, dimers, and higher oligomeric forms of S1R were identified under non-liganded conditions. In the presence of the prototypic S1R agonist, (+)-pentazocine, however, monomers and dimers were the prevailing forms of S1R. The prototypic antagonist, haloperidol, on the other hand, favored higher order S1R oligomers. These data, in sum, indicate that heterologously expressed S1Rs occur in vivo in COS-7 cells in multiple oligomeric forms and that S1R ligands alter these oligomeric structures. We suggest that the S1R oligomerization states may regulate its function(s).
    Biochemical Journal 12/2014;
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    ABSTRACT: Central to Alzheimer's Disease is the misfolding of amyloid-beta (Aβ) peptide, which generates an assorted population of amorphous aggregates, oligomers and fibres. Metal ion homeostasis is disrupted in the brains of sufferers of Alzheimer's disease and causes heightened Alzheimer's disease phenotype in animal models. Here we illustrate that substochiometric Cu2+ effects the misfolding pathway of Aβ(1-40), and the more toxic Aβ(1-42), in markedly different ways. Cu2+ accelerates Aβ(1-40) fibre formation, in contrast, for Aβ(1-42) substoichiometric levels of Cu2+ almost exclusively promotes the formation of oligomeric and protofibrillar assemblies. Indeed, mature Aβ(1-42) fibres are disassembled into oligomers when Cu2+ is added. These Cu2+ stabilised oligomers of Aβ(1-42) interact with the lipid-bilayer, disrupting the membrane and increasing permeability. Our investigation of Aβ(1-40)/Aβ(1-42) mixtures with Cu2+ revealed that Aβ(1-40) neither contributed nor perturbed formation of Aβ(1-42) oligomers, though Cu2+-Aβ(1-42) will frustrate Cu2+-Aβ(1-40) fibre growth. Small amounts of Cu2+ accentuates differences in the propensity of Aβ(1-40) and Aβ(1-42) to form synaptotoxic oligomers, providing an explanation for the connection between disrupted Cu2+ homeostasis and elevated Aβ(1-42) neurotoxicity in Alzheimer's disease.
    Biochemical Journal 12/2014;
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    ABSTRACT: Recent advances in G-quadruplex (G4) studies have confirmed that G4 structures exist in living cells and may have detrimental effects on various DNA transactions. How helicases resolve G4, however, has just begun to be studied and remains largely unknown. Here we use single-molecule fluorescence assays to probe Pif1-catalysed unfolding of G4 in a DNA construct resembling an ongoing synthesis of lagging-strand stalled by G4. Strikingly, Pif1 unfolds, and then halts at the ss/dsDNA junction, followed by rapid reformation of G4 and 'acrobatic' reinitiation of unfolding by the same monomer. Thus, Pif1 unfolds single G4 structures repetitively. Furthermore, it is found that Pif1 unfolds G4 sequentially in two large steps. Our study has revealed that, as a stable intermediate, G-triplex plays an essential role in this process. The repetitive unfolding activity may facilitate Pif1 disrupting the continuously-reforming obstructive G4 structures to rescue a stalled replication fork. The proposed mechanism for step-wise unfolding of G4 is probably applicable to other helicases that resolve G4 structures for maintaining genome stability.
    Biochemical Journal 12/2014;
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    ABSTRACT: Class II/III PI3Ks (phosphoinositide 3-kinases) produce the PtdIns(3)P lipid that is involved in intracellular vesicular trafficking. In contrast with class I PI3Ks, the potential signalling roles of class II/III PI3Ks are poorly understood. In a recent article in the Biochemical Journal, Bago and co-workers report that Vps34 (vacuolar protein sorting 34), the only class III PI3K, controls the activity of SGK3 (serum- and glucocorticoid-regulated protein kinase 3). Like other AGC kinases, the SGKs (SGK1, SGK2 and SGK3) are activated by dual phosphorylation. Unlike its cousins SGK1 and SGK2, SGK3 contains a PtdIns(3)P-binding domain, providing an additional element of regulation. The study by Bago et al. characterizes and makes extensive use of a Novartis Vps34 inhibitor (VPS34-IN1) that inhibits this PI3K isoform with nanomolar potency, without affecting other lipid kinases or more than 300 protein kinases. The authors show that this compound very rapidly reduced PtdIns(3)P levels at the endosome with concomitant loss of SGK3 phosphorylation. Co-inhibition of class I PI3Ks led to a further reduction in SGK3 activity, indicating that class I PI3Ks may also regulate SGK3 activity through an additional, currently unknown, mechanism. It remains to be assessed whether the novel PI3K-protein kinase connection established by this study is subject to acute cellular stimulation or is part of a constitutive housekeeping function. VPS34-IN1 will provide a useful tool to decipher the kinase-dependent functions of Vps34, with acute changes in SGK3 phosphorylation and subcellular localization being new biomarkers of Vps34 activity.
    Biochemical Journal 12/2014; 464(2):e7-e10.
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    ABSTRACT: In eukaryotes, amino acid deprivation leads to the accumulation of uncharged tRNAs that are detected by Gcn2, which in turn phosphorylates eIF2α, an essential process for overcoming starvation. In Saccharomyces cerevisiae, sensing amino acid shortages requires that Gcn2 binds directly to its effector protein Gcn1, and both must associate with the ribosome. Our hypothesis is that uncharged tRNAs occur in the ribosomal A-site, and that Gcn1 is directly involved in transfer of this starvation signal to Gcn2. Here we provide evidence that Gcn1 directly contacts the small ribosomal protein Rps10. Gcn1 residues 1060-1777 showed a yeast 2-hybrid interaction with Rps10A. In vitro, Rps10A or Rps10B co-precipitated Gcn1[1060-1777] in an RNA-independent manner. rps10AΔ or rps10BΔ strains showed reduced eIF2α phosphorylation under replete conditions and shortly after onset of starvation, suggesting that Gcn1-mediated Gcn2 activation was impaired. Overexpression of GST-tagged Rps10 reduced growth under amino acid starvation, and this was exacerbated by the Gcn1-M7A mutation known to impair Gcn1-ribosome interaction and Gcn2 activity. Under amino acid starvation, eEF3 overexpression known to weaken Gcn1 function on the ribosome, exacerbated the growth defect of rps10AΔ or rps10BΔ strains. Taken together, these data support the idea that Gcn1 contacts ribosome-bound Rps10 to efficiently mediate Gcn2 activation.
    Biochemical Journal 12/2014;
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    ABSTRACT: The proton-linked monocarboxylate transporters (MCTs) are required for lactic acid transport into all mammalian cells. Thus they play an essential role in tumour cells that are usually highly glycolytic and are promising targets for anti-cancer drugs. AR-C155858 is a potent MCT1 inhibitor (Ki ~ 2 nM) that also inhibits MCT2 when associated with basigin but not MCT4. Previous work revealed that AR-C155858 binding to MCT1 occurs from the intracellular side and involves transmembrane helices 7-10. Here we generate a molecular model of MCT4 based on our previous models of MCT1 and identify residues in the intracellular substrate binding cavity that differ significantly between MCT4 and MCT1 / MCT2 and so might account for differences in inhibitor binding. We tested their involvement using site-directed mutagenesis (SDM) of MCT1 to change residues individually or in combination to their MCT4 equivalent and determined inhibitor sensitivity following expression in Xenopus oocytes. Phe360 and Ser364 were identified as important for AR-C155858 binding with the F360Y/S364G mutant exhibiting > 100-fold reduction in inhibitor sensitivity. To refine the binding site further we used molecular dynamics simulations and additional SDM. This approach implicated six more residues whose involvement was confirmed by both transport studies and [3H]- AR-C155858 binding to oocyte membranes. Taken together our data imply that N147, R306 and S364 are important for directing AR-C155858 to its final binding site which involves interaction of the inhibitor with K38, D302 and F360 (residues that also play key roles in the translocation cycle) and also L274 and S278.
    Biochemical Journal 12/2014;
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    ABSTRACT: Glucocorticoids play an important role in the treatment of inflammation and immune disorders, despite side effects, which include metabolic derangements such as central adiposity. These studies examine the role of protein phosphatase 5 (Ppp5) in glucocorticoid receptor complexes, which mediate response to glucocorticoids. Mice homozygous for inactivated Ppp5 (Ppp5D274A/D274A) exhibit decreased adipose tissue surrounding the gonads and kidneys compared with wild type mice. Adipocyte size is smaller, more preadipocytes/stromal cells are present in their gonadal fat tissue and differentiation of preadipocytes to adipocytes is retarded. Glucocorticoid levels are raised and the glucocorticoid receptor is hyperphosphorylated in adipose tissue of Ppp5D274A/D274A mice at Ser212 and Ser220 (orthologous to human Ser203 and Ser211) in the absence of glucocorticoids. Preadipocyte cultures from Ppp5D274A/D274A mice show decreased down regulation of Delta-like protein-1/Preadipocyte factor-1, hyperphosphorylation of ERK2 and increased concentration of SOX9, changes in a pathway essential for preadipocyte differentiation, which leads to decreased concentrations of the transcription factors CEBPβ and CEBPα necessary for the later stages of adipogenesis. The data indicate that Ppp5 plays a crucial role in modifying glucocorticoid receptor-mediated initiation of adipose tissue differentiation, suggesting that inhibition of Ppp5 may potentially be beneficial to prevent obesity during glucocorticoid treatment.
    Biochemical Journal 12/2014;
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    ABSTRACT: Vibrio cholerae cytolysin (VCC) kills target eukaryotic cells by forming transmembrane oligomeric β-barrel pores. Once irreversibly converted into the transmembrane oligomeric form, VCC acquires unusual structural stability, and loses cytotoxic property. It is therefore possible that upon exerting its cytotoxic activity, oligomeric form of VCC retained in the disintegrated membrane fractions of the lysed cells would survive within the host cellular milieu for sufficiently prolonged duration, without causing any further cytotoxicity. Under such circumstances, VCC oligomers may potentially be recognized by the host immune cells. Based on such hypothesis, in the present study we have explored the interaction of the transmembrane oligomeric form of VCC with the monocytes and macrophage cells of the innate immune system. Our study shows that the VCC oligomers assembled in the liposome membranes elicit potent pro-inflammatory responses in the monocytes and macrophages, via stimulation of the TLR2/TLR6-dependent signalling cascades that involve MyD88/IRAK1/TRAF6. VCC oligomer-mediated pro-inflammatory responses critically depend on the activation of the transcription factor NF-kB. Pro-inflammatory responses induced by the VCC oligomers also require activation of the MAPK family member JNK, which presumably acts via stimulation of the transcription factor AP-1. Notably, role of the MAPK p38 could not be documented in the process.
    Biochemical Journal 11/2014;
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    ABSTRACT: The protein mammalian target of rapamycin (mTOR) is a serine/threonine kinase regulating a number of biochemical pathways controlling cell growth. mTOR exists in two complexes termed mTORC1 and mTORC2. Raptor is associated with mTORC1 and is essential for its function. Ablation of Raptor in skeletal muscle results in several phenotypic changes including decreased life expectancy, increased glycogen deposits and alterations of the twitch kinetics of slow fibres. Here we show that in muscle specific raptor knock-out the bulk of glycogen phosphorylase is mainly associated in its cAMP-non stimulated form with sarcoplasmic reticulum membranes. In addition, 3[H]-ryanodine and 3[H]-PN200-110 equilibrium binding show a ryanodine to dihydropyridine receptors ratio of 0.79 and 1.35 for wild type and raptor knock-out skeletal muscle membranes, respectively. Peak amplitude and time to peak of the global calcium transients evoked by supramaximal field stimulation were not different between wild type and raptor knock-out. However, the increase of the voltage sensor-uncoupled RyRs leads to an increase of both frequency and mass of elementary calcium release events (ECRE) induced by hyper-osmotic shock in FDB fibres from raptor knock-out. This study shows that the protein composition and function of the molecular machinery involved in skeletal muscle excitation-contraction coupling is affected by mTORC1 signaling.
    Biochemical Journal 11/2014;
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    ABSTRACT: Plants and bacteria synthesize the essential human micronutrient riboflavin (vitamin B2) via the same multistep pathway. The early intermediates of this pathway are notoriously reactive, and may be over-produced in vivo because riboflavin biosynthesis enzymes lack feedback controls. Here we demon-str-ate disposal of riboflavin intermediates by COG3236 (DUF1768), a protein of previously unknown funct-ion that is fused to two different riboflavin pathway enzymes in plants and bacteria (RIBR and RibA, respectively). We present cheminformatic, biochemical, genetic, and genomic evidence to show that: (i) plant and bacterial COG3236 proteins cleave the N-glycosidic bond of the first two intermed-iates of riboflavin biosynthesis, yielding relatively innocuous products; (ii) certain COG3236 proteins are in a multienzyme riboflavin biosynthesis complex that gives them privileged access to riboflavin intermediates; and (iii) COG3236 action in Arabidopsis thaliana and Escherichia coli helps maintain flavin levels. COG3236 proteins thus illustrate two emerging principles in chemical biology: directed overflow metabolism, in which excess flux is diverted out of a pathway, and the pre-emption of damage from reactive metabolites.
    Biochemical Journal 11/2014;
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    ABSTRACT: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide and chronic hepatitis B virus (HBV) infection is the most common risk factor for HCC. The HBV proteins can induce oncogenic effects or synergy effects with the hyperproliferative response on transformation into HCC. Cyclic-AMP-responsive-element-binding protein H (CREBH) activated by ER stress is liver-specific expressed ER-resident transmembrane bZIP transcription factors. Here, we addressed the roles of CREBH activated by ER stress on HBV-induced hepatic cell proliferation. We confirmed CREBH activation by ER stress and showed CREBH activation by HBx-induced ER stress. CREBH activated by HBx increased the expressions of AP-1 target genes through c-Jun induction. Under pathological conditions such as liver damage or liver regeneration, activated CREBH may play important roles on hepatic inflammation and cell proliferation as insulin receptor with dual-function under the conditions. We showed that CREBH activated by HBx interacted with HBx protein, leading to synergic effect on the expressions of AP-1 target genes and proliferation of heptocellular carcinoma cells and mouse primary hepatocytes. In conclusion, in HBV-infected hepatic cells or chronic hepatitis B patients, CREBH may induce proliferation of hepatic cells in cooperation with HBx, resulting in HCC.
    Biochemical Journal 11/2014;
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    ABSTRACT: In insects, molting and metamorphosis are strictly regulated by ecdysteroids. Ecdysteroid synthesis is positively or negatively controlled by several neuropeptides. The prothoracicostatic peptide BmPTSP, isolated from the larval brain of Bombyx mori, has been demonstrated to inhibit ecdysteroid synthesis in the prothoracic glands. More recently, the newly recognized B. mori receptor for Drosophila melanogaster sex peptide has been identified as a receptor for BmPTSP. However, details on the signaling pathways and physiological functions of this receptor have remained elusive. Here, we report the functional characterization of the BmPTSPR/SPR using both mammalian and insect cells. Synthetic DmSP shows the potential to inhibit forskolin or AKH-induced CRE-driven luciferase activity in a manner comparable to synthetic BmPTSP1. However, DmSP displayed a much lower activity in triggering Ca2+ mobilization and internalization than did BmPTSP1. Additionally, FAM fluorophore-labeled DmSP and BmPTSP3 were found to bind specifically to BmPTSPR/SPR. The binding of FAM-DmSP was displaced by unlabeled DmSP, but not by unlabeled BmPTSP1 and, vice versa, the binding of FAM-BmPTSP3 was blocked by unlabeled BmPTSP3, but not by unlabeled DmSP. Moreover, internalization assays demonstrated that BmPTSP1, but not DmSP, evoked recruitment of the Bombyx nonvisual arrestin, Kurtz, to the activated BmPTSPR/SPR in the plasma membrane. This was followed by induction of internalization. This suggests that BmPTSP1 is likely an endogenous ligand specific for BmPTSPR/SPR. We therefore designate this receptor BmPTSPR. In contrast, DmSP is an allosteric agonist that is biased towards Gαi/o-dependent cAMP production and away from Ca2+ mobilization and arrestin recruitment.
    Biochemical Journal 11/2014;