scholarly journals Lipid-derived electrophiles induce covalent modification and aggregation of Cu,Zn-superoxide dismutase in a hydrophobicity-dependent manner

2019 ◽  
Author(s):  
Lucas S. Dantas ◽  
Lucas G. Viviani ◽  
Alex Inague ◽  
Erika Piccirillo ◽  
Leandro de Rezende ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Critical Factor ◽  
Covalent Modification ◽  
Protein Sequencing ◽  
Size Exclusion ◽  
Dependent Manner ◽  
Loss Of Function ◽  
Dimer Interface ◽  
Electrostatic Loop

ABSTRACTLipid peroxidation generates a huge number of reactive electrophilic aldehyde products. These reactive aldehydes can modify macromolecules such as proteins, resulting in loss of function and/or aggregation. The accumulation of Cu,Zn-superoxide dismutase (SOD1) aggregates is associated with familial cases of amyotrophic lateral sclerosis (ALS). Recent studies have shown that lipid and its oxidized derivatives may play a role in this process. Here we aimed to compare and characterize the ability of lipid-derived electrophiles with different hydrophobicities to induce SOD1 modification and aggregation in vitro. SOD1 was incubated with 4-hydroxy-2-hexenal (HHE), 4-hydroxy- 2-nonenal (HNE), 2-hexen-1-al (HEX), 2,4-nonadienal (NON), 2,4-decadienal (DEC) or secosterol aldehydes (Seco-A or Seco-B) at 37°C for 24 h. Size exclusion chromatography analysis showed that hydrophobic aldehydes smarkedly enhances apo- SOD1 aggregation. More importantly, aggregation level was positively correlated to calculated aldehyde hydrophobicities (LogP). Protein sequencing by LC-MS/MS showed that aldehydes covalently modifies SOD1 at aggregation prone regions. For instance, specific lysine residues located mainly nearby the dimer interface (K3, K9) and at the electrostatic loop (K122, K128, K136) were ubiquitously modified by all aldehydes. The α,β-unsaturated aldehydes also promoted modifications on histidine and cysteine residues, with H120 and C6 being the most commonly modified residues. Overall, our data suggest that electrophile’s hydrophobicity is a critical factor that strongly influences protein aggregation propensity.Graphical abstractHighlights- Aldehyde hydrophobicity is positively correlated to SOD1 aggregation;- Lys residues located nearby the SOD1 dimer interface and electrostatic loop are ubiquitously modified by all aldehydes;- Hydrophobic aldehydes increase the lipophilic potential surface of the region where they bind;

scholarly journals Stem cell factor and interleukin-7 synergize to enhance early myelopoiesis in vitro

Blood ◽  
1994 ◽  
Vol 84 (5) ◽  
pp. 1450-1456 ◽  
Author(s):  
C Fahlman ◽  
HK Blomhoff ◽  
OP Veiby ◽  
IK McNiece ◽  
SE Jacobsen
Keyword(s):  
Stem Cell ◽  
B Cells ◽  
Stem Cell Factor ◽  
Critical Factor ◽  
Dependent Manner ◽  
Murine Bone Marrow ◽  
Proliferative Effect ◽  
Interleukin 7 ◽  
Potent Growth

Abstract Interleukin-7 (IL-7) has been shown to be a critical factor in murine lymphoid development. It stimulates pre-B cells to divide in the absence of stroma cells and it is an important growth regulator of immature and mature T cells. IL-7 has been shown to synergize with stem cell factor (SCF) to provide a potent growth stimulus for pre-B cells. However, the combined effects of IL-7 and SCF on murine primitive hematopoietic cells in vitro have not been established. In the present study, the effects of recombinant rat (rr) SCF and recombinant human (rh) IL-7 on primitive murine bone marrow progenitors (Lin-Sca1+) were investigated in single-cell cloning experiments. rhIL-7 alone had no proliferative effect on Lin-Sca1+ cells, but in a dose-dependent manner directly enhanced rrSCF-induced colony formation, with an average increase in colony numbers of 2.7-fold. Interestingly, the cells formed in response to SCF and IL-7 were predominantly mature granulocytes. Thus, SCF and IL-7 synergize to stimulate early myelopoiesis in vitro.

GFI1 Is a Tumor Suppressor in Myeloid Progenitors

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2942-2942
Author(s):  
Aditya Chaubey ◽  
Shane Hormon ◽  
Chinavenmeni S. Velu ◽  
Tristan Bourdeau ◽  
Jinfang Zhu ◽  
...  
Keyword(s):  
Dna Sequences ◽  
Myeloid Leukemia ◽  
Dependent Manner ◽  
Loss Of Function ◽  
Increased Risk ◽  
Myeloid Leukemias ◽  
Dose Dependent ◽  
Anterior Posterior

Abstract In severe congenital neutropenia (SCN) patients and mice with Growth factor independent-1 (Gfi1) loss of function, arrested progenitors are suspended in a hyperproliferative state while terminal granulpoiesis is blocked. SCN patients are at increased risk for the development of acute myeloid leukemia. We demonstrate that Gfi1 directly targets HoxA9, Pbx1 and Meis1 during normal myelopoiesis. Gfi1−/− progenitors exhibit elevated levels of HoxA9, Pbx1 and Meis1, exaggerated HoxA9-Pbx1-Meis1 activity, and increased persistence in vivo and in vitro. Limiting HoxA9 alleles corrects, in a dose dependent manner, in vivo and in vitro phenotypes observed with loss of Gfi1. Moreover, in a manner conserved in Drosophila anterior/posterior patterning, we demonstrate that these factors can compete for occupancy of DNA sequences encoding composite Gfi1-HoxA9-Pbx1-Meis1 binding sites. Finally, the expression of Gfi1 and HoxA9 are inverse and stratify human myeloid leukemias, suggesting a role for HoxA9- Gfi1 antagonism in human AML. In agreement with this, a myeloproliferative disorder progresses into a rapid, lethal and transplantable myeloid leukemia in a Gfi1−/− setting. We conclude that the lifespan and oncogenic transformation of hematopoietic progenitor cells is regulated through a conserved competition between Gfi1 and HoxA9-Pbx1-Meis1.

scholarly journals Processing of progranulin into granulins involves multiple lysosomal proteases and is affected in frontotemporal lobar degeneration

2020 ◽  
Author(s):  
Swetha Mohan ◽  
Paul J. Sampognaro ◽  
Andrea R. Argouarch ◽  
Jason C. Maynard ◽  
Anand Patwardhan ◽  
...  
Keyword(s):  
Frontotemporal Lobar Degeneration ◽  
Cathepsin L ◽  
Dependent Manner ◽  
Loss Of Function ◽  
New Strategy ◽  
Lysosomal Proteases ◽  
Ph Dependent ◽  
Increased Activity

Abstract Background: Progranulin loss-of-function mutations are linked to frontotemporal lobar degeneration with TDP-43 positive inclusions (FTLD-TDP-Pgrn). Progranulin (PGRN) is an intracellular and secreted pro-protein that is proteolytically cleaved into individual granulin peptides, which are increasingly thought to contribute to FTLD-TDP-Pgrn disease pathophysiology. Intracellular PGRN is processed into granulins in the endo-lysosomal compartments. Therefore, to better understand the conversion of intracellular PGRN into granulins, we systematically tested the ability of different classes of endo-lysosomal proteases at a range of pH setpoints.Results: In vitro cleavage assays identified multiple enzymes that can process human PGRN into multi- and single-granulin fragments in a pH-dependent manner. We confirmed the role of cathepsin B and cathepsin L in PGRN processing and showed that these and several previously unidentified lysosomal proteases (cathepsins E, G, K, S and V) are able to process PGRN in variable, pH-dependent manners. In addition, we have demonstrated a new role for asparagine endopeptidase (AEP) in processing PGRN, with AEP having the unique ability to liberate granulin F from the pro-protein. Brain tissue from individuals with FTLD-TDP-Pgrn show increased PGRN processing to granulin F, correlating with increased activity of AEP, in a region-specific manner. Conclusions: This study demonstrates that multiple lysosomal proteases may work in concert to liberate granulins and implicates both AEP and granulin F in the neurobiology of FTLD-TDP-Pgrn. Modulating progranulin cleavage may represent a new strategy to modulate PGRN and granulin levels in disease.

scholarly journals EPH/EPHRIN SIGNALING CONTROLS PROGENITOR IDENTITIES IN THE VENTRAL SPINAL CORD

2016 ◽  
Author(s):  
Julien Laussu ◽  
Christophe Audouard ◽  
Anthony Kischel ◽  
Poincyane Assis-Nascimento ◽  
Nathalie Escalas ◽  
...  
Keyword(s):  
Spinal Cord ◽  
Sonic Hedgehog ◽  
Cell Communication ◽  
Dependent Manner ◽  
Eph Receptors ◽  
Loss Of Function ◽  
Summary Statement ◽  
Dose Dependent ◽  
Ventral Spinal Cord

SUMMARY STATEMENTThis article by Laussu et al. describes a role for Eph:ephrin signaling in controlling the identity of neural progenitors in the ventral spinal cord.Early specification of progenitors of the ventral spinal cord involves the morphogen Sonic Hedgehog which induces distinct progenitor identities in a dose-dependent manner. Following these initial patterning events, progenitor identities have to be maintained in order to generate appropriate numbers of progeny. Here we provide evidence that communication via Eph:ephrin signaling is required to maintain progenitor identities in the ventral spinal cord. We show that ephrinB2 and ephrinB3 are expressed in restricted progenitor domains in the ventral spinal cord while several Eph receptors are more broadly expressed. Further, we provide evidence that expression of Efnb3 and EphA4 is controlled by Shh. Genetic loss-of-function analyses indicate that expression of ephrinB2 and ephrinB3 is required to control progenitor identities and in vitro experiments reveal that activation of Eph forward signaling in spinal progenitors up-regulates the expression of the identity transcription factor Nkx2.2. Altogether our results indicate that cell-to-cell communication is necessary to control progenitor identity in the ventral spinal cord.

scholarly journals Capsule carbohydrate structure determines virulence in Acinetobacter baumannii

2021 ◽  
Vol 17 (2) ◽  
pp. e1009291
Author(s):  
Yuli Talyansky ◽  
Travis B. Nielsen ◽  
Jun Yan ◽  
Ulrike Carlino-Macdonald ◽  
Gisela Di Venanzio ◽  
...  
Keyword(s):  
Acinetobacter Baumannii ◽  
Bacterial Pathogen ◽  
Specific Gene ◽  
Dependent Manner ◽  
Bacterial Burden ◽  
Carbohydrate Structure ◽  
Loss Of Function ◽  
Capsule Assembly

Acinetobacter baumannii is a highly antibiotic-resistant bacterial pathogen for which novel therapeutic approaches are needed. Unfortunately, the drivers of virulence in A. baumannii remain uncertain. By comparing genomes among a panel of A. baumannii strains we identified a specific gene variation in the capsule locus that correlated with altered virulence. While less virulent strains possessed the intact gene gtr6, a hypervirulent clinical isolate contained a spontaneous transposon insertion in the same gene, resulting in the loss of a branchpoint in capsular carbohydrate structure. By constructing isogenic gtr6 mutants, we confirmed that gtr6-disrupted strains were protected from phagocytosis in vitro and displayed higher bacterial burden and lethality in vivo. Gtr6+ strains were phagocytized more readily and caused lower bacterial burden and no clinical illness in vivo. We found that the CR3 receptor mediated phagocytosis of gtr6+, but not gtr6-, strains in a complement-dependent manner. Furthermore, hypovirulent gtr6+ strains demonstrated increased virulence in vivo when CR3 function was abrogated. In summary, loss-of-function in a single capsule assembly gene dramatically altered virulence by inhibiting complement deposition and recognition by phagocytes across multiple A. baumannii strains. Thus, capsular structure can determine virulence among A. baumannii strains by altering bacterial interactions with host complement-mediated opsonophagocytosis.

scholarly journals Novel Antioxidant, Anti-α-Amylase, Anti-Inflammatory and Antinociceptive Water-Soluble Polysaccharides from the Aerial Part of Nitraria retusa

Foods ◽  
2019 ◽  
Vol 9 (1) ◽  
pp. 28 ◽  
Author(s):  
Ilhem Rjeibi ◽  
Faiez Hentati ◽  
Anouar Feriani ◽  
Najla Hfaiedh ◽  
Cédric Delattre ◽  
...  
Keyword(s):  
Impact Analysis ◽  
Water Soluble ◽  
Size Exclusion ◽  
Gas Chromatography Mass Spectrometry ◽  
Dependent Manner ◽  
Mice Model ◽  
Biological Potential ◽  
Nitraria Retusa ◽  
Anti Inflammatory

In this paper, water-soluble polysaccharides (named as NRLP) were extracted from Nitraria retusa leaves. The main structural features of NRLP were determined by High-pressure size exclusion chromatography, Fourier transform infrared and Gas Chromatography/Mass Spectrometry-Electronic Impact analysis. The in vitro and in vivo biological potential of NRLP were evaluated by measuring its antioxidant (•OH and DPPH• scavenging, total antioxidant capacity), anti-α-amylase as well as anti-inflammatory and antinociceptive activities in a mice model. NRLP was composed of Rha (33.7%), Gal (18.1%), GalA (15.0%), Glc (13.3%), Ara (13.3%), Xyl (3.8%), and GlcA (2.8%) and showed a Molecular Weight (Mw) of 23.0 kDa and a polydispersity index (PDI) of 1.66. The investigations highlighted a significant antioxidant activity (IC50 = 2.4–2.6 mg/mL) and an inhibition activity against α-amylase (IC50 = 4.55 mg/mL) in a dose-dependent manner. Further, NRLP revealed interesting anti-edematous effects and antinociceptive activities (both > 70%). These results open up new pharmacological prospects for the water-soluble polysaccharides extracted from Nitraria retusa leaves.

scholarly journals Processing of progranulin into granulins involves multiple lysosomal proteases and is affected in frontotemporal lobar degeneration

2020 ◽  
Author(s):  
Swetha Mohan ◽  
Paul J. Sampognaro ◽  
Andrea R. Argouarch ◽  
Jason C. Maynard ◽  
Anand Patwardhan ◽  
...  
Keyword(s):  
Frontotemporal Lobar Degeneration ◽  
Cathepsin L ◽  
Dependent Manner ◽  
Loss Of Function ◽  
Specific Manner ◽  
Lysosomal Proteases ◽  
Ph Dependent ◽  
Increased Activity

Abstract Background - Progranulin loss-of-function mutations are linked to frontotemporal lobar degeneration with TDP-43 positive inclusions (FTLD-TDP-Pgrn). Progranulin (PGRN) is an intracellular and secreted pro-protein that is proteolytically cleaved into individual granulin peptides, which are increasingly thought to contribute to FTLD-TDP-Pgrn disease pathophysiology. Intracellular PGRN is processed into granulins in the endo-lysosomal compartments. Therefore, to better understand the conversion of intracellular PGRN into granulins, we systematically tested the ability of different classes of endo-lysosomal proteases to process PGRN at a range of pH setpoints. Results - In vitro cleavage assays identified multiple enzymes that can process human PGRN into multi- and single-granulin fragments in a pH-dependent manner. We confirmed the role of cathepsin B and cathepsin L in PGRN processing and showed that these and several previously unidentified lysosomal proteases (cathepsins E, G, K, S and V) are able to process PGRN in distinctive, pH-dependent manners. In addition, we have demonstrated a new role for asparagine endopeptidase (AEP) in processing PGRN, with AEP having the unique ability to liberate granulin F from the pro-protein. Brain tissue from individuals with FTLD-TDP-Pgrn show increased PGRN processing to granulin F and an increased activity of AEP, in a region-specific manner. Conclusions - This study demonstrates that multiple lysosomal proteases may work in concert to liberate multi-granulin fragments and granulins. It also implicates both AEP and granulin F in the neurobiology of FTLD-TDP-Pgrn. Modulating progranulin cleavage and granulin production may represent therapeutic strategies for FTLD-Pgrn and other progranulin-related diseases.

scholarly journals Processing of progranulin into granulins involves multiple lysosomal proteases and is affected in frontotemporal lobar degeneration

2020 ◽  
Author(s):  
Swetha Mohan ◽  
Paul J. Sampognaro ◽  
Andrea R. Argouarch ◽  
Jason C. Maynard ◽  
Anand Patwardhan ◽  
...  
Keyword(s):  
Frontotemporal Lobar Degeneration ◽  
Cathepsin L ◽  
Dependent Manner ◽  
Loss Of Function ◽  
New Strategy ◽  
Lysosomal Proteases ◽  
Ph Dependent ◽  
Increased Activity

Abstract Background - Progranulin loss-of-function mutations are linked to frontotemporal lobar degeneration with TDP-43 positive inclusions (FTLD-TDP-Pgrn). Progranulin (PGRN) is an intracellular and secreted pro-protein that is proteolytically cleaved into individual granulin peptides, which are increasingly thought to contribute to FTLD-TDP-Pgrn disease pathophysiology. Intracellular PGRN is processed into granulins in the endo-lysosomal compartments. Therefore, to better understand the conversion of intracellular PGRN into granulins, we systematically tested the ability of different classes of endo-lysosomal proteases at a range of pH setpoints. Results - In vitro cleavage assays identified multiple enzymes that can process human PGRN into multi- and single-granulin fragments in a pH-dependent manner. We confirmed the role of cathepsin B and cathepsin L in PGRN processing and showed that these and several previously unidentified lysosomal proteases (cathepsins E, G, K, S and V) are able to process PGRN in variable, pH-dependent manners. In addition, we have demonstrated a new role for asparagine endopeptidase (AEP) in processing PGRN, with AEP having the unique ability to liberate granulin F from the pro-protein. Brain tissue from individuals with FTLD-TDP-Pgrn show increased PGRN processing to granulin F, correlating with increased activity of AEP, in a region-specific manner. Conclusions - This study demonstrates that multiple lysosomal proteases may work in concert to liberate granulins and implicates both AEP and granulin F in the neurobiology of FTLD-TDP-Pgrn. Modulating progranulin cleavage may represent a new strategy to modulate PGRN and granulin levels in disease.

Pelle kinase is activated by autophosphorylation during Toll signaling in Drosophila

Development ◽  
2002 ◽  
Vol 129 (8) ◽  
pp. 1925-1933 ◽  
Author(s):  
Baohe Shen ◽  
James L. Manley
Keyword(s):  
Dependent Manner ◽  
Loss Of Function ◽  
Concentration Dependent Manner ◽  
Downstream Target ◽  
Toll Signaling ◽  
Early Embryos ◽  
High Concentrations ◽  
L2 Cells

The Drosophila Pelle kinase plays a key role in the evolutionarily conserved Toll signaling pathway, but the mechanism responsible for its activation has been unknown. We present in vivo and in vitro evidence establishing an important role for concentration-dependent autophosphorylation in the signaling process. We first show that Pelle phosphorylation can be detected transiently in early embryos, concomitant with activation of signaling. Importantly, Pelle phosphorylation is enhanced in a gain-of-function Toll mutant (Toll10b), but decreased by loss-of-function Toll alleles. Next we found that Pelle is phosphorylated in transfected Schneider L2 cells in a concentration-dependent manner such that significant modification is observed only at high Pelle concentrations, which coincide with levels required for phosphorylation and activation of the downstream target, Dorsal. Pelle phosphorylation is also enhanced in L2 cells co-expressing Toll10b, and is dependent on Pelle kinase activity. In vitro kinase assays revealed that recombinant, autophosphorylated Pelle is far more active than unphosphorylated Pelle. Importantly, unphosphorylated Pelle becomes autophosphorylated, and activated, by incubation at high concentrations. We discuss these results in the context of Toll-like receptor mediated signaling in both flies and mammals.

TAMI-28. IDENTIFICATION OF A NEUROLIGIN-3 BINDING PARTNER IN HIGH-GRADE GLIOMAS AND NORMAL PROGENITORS

2021 ◽  
Vol 23 (Supplement_6) ◽  
pp. vi204-vi204
Author(s):  
Shawn Gillespie ◽  
Yoon Seok Kim ◽  
Anna Geraghty ◽  
Michael Quezada ◽  
James Reed ◽  
...  
Keyword(s):  
Neuronal Activity ◽  
Glioma Cells ◽  
Diffuse Intrinsic Pontine Glioma ◽  
Size Exclusion ◽  
Dependent Manner ◽  
High Grade ◽  
Gamma Secretase ◽  
Binding Partner ◽  
High Grade Gliomas

Abstract High-grade gliomas, including diffuse intrinsic pontine glioma, are lethal cancers whose progression is strongly regulated by neuronal activity. One way in which gliomas detect neuronal activity is via interaction with the ectodomain of post-synaptic adhesion protein neuroligin-3 (NLGN3), which is shed from neurons and oligodendrocyte precursors (OPCs) by the ADAM10 sheddase in an activity dependent manner. NLGN3 signaling drives glioma growth, but the cognate binding partner of shed NLGN3 (sNLGN3) on glioma cells is unknown. Here, we employed a proximity labeling technique to identify chondroitin sulfate proteoglycan 4 (CSPG4) as a putative binding partner of sNLGN3 in gliomas. We then confirmed complexing between recombinant proteins with size exclusion chromatography and are determining kinetics and affinity by surface plasmon resonance. When looking for evidence of binding in cells, we were surprised to find that sNLGN3 triggers regulated intramembrane proteolysis (RIP) of CSPG4, leaving no trace of the interaction at the membrane. sNLGN3 binding first induces ADAM10-mediated cleavage and release of the CSPG4 ectodomain, followed by gamma secretase-mediated release of the intracellular domain and downstream signaling in OPCs and gliomas. Pre-treatment of glioma cells or OPCs with an ADAM10 inhibitor entirely blocks sNLGN3-induced CSPG4 shedding. Acute depletion of CSPG4 via CRISPR gene editing renders glioma cells insensitive to the growth-promoting effects of sNLGN3 in vitro. Furthermore, we find that tamoxifen-induced deletion of NLGN3 from murine OPCs reduces the total number of OPCs, suggesting that this signaling axis promotes maintenance of OPC stemness in an autocrine fashion. Indeed, gamma secretase inhibition accelerates OPC differentiation in vitro, pointing towards a fundamental role for sNLGN3-CSPG4 signaling in OPCs and high-grade gliomas. Altogether, our results form a critical missing link in understanding how glioma cells detect a key neuronal activity-regulated signal, suggest intriguing links to OPC biology and identify a therapeutic target to disrupt neuron-glioma interactions.

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