scholarly journals Progranulin deficiency leads to reduced glucocerebrosidase activity

2019 ◽  
Author(s):  
Xiaolai Zhou ◽  
Daniel H. Paushter ◽  
Mitchell D. Pagan ◽  
Dongsung Kim ◽  
Raquel L. Lieberman ◽  
...  
Keyword(s):  
Lysosomal Enzyme ◽  
Frontotemporal Lobar Degeneration ◽  
Neuronal Ceroid Lipofuscinosis ◽  
Complete Loss ◽  
Lysosomal Hydrolase ◽  
Gene Encoding ◽  
Exact Function ◽  
Pathological Mechanism ◽  
Dose Dependent ◽  
Deficient Mice

AbstractMutation in the GRN gene, encoding the progranulin (PGRN) protein, shows a dose-dependent disease correlation, wherein haploinsufficiency results in frontotemporal lobar degeneration (FTLD) and complete loss results in neuronal ceroid lipofuscinosis (NCL). Although the exact function of PGRN is unknown, it has been increasingly implicated in lysosomal physiology. Here we report that PGRN interacts with the lysosomal enzyme, glucocerebrosidase (GBA), and is essential for proper GBA activity. GBA activity is significantly reduced in tissue lysates from PGRN-deficient mice. This is further evidence that reduced lysosomal hydrolase activity may be a pathological mechanism in cases of GRN-related FTLD and NCL.

scholarly journals UBQLN2 Promotes the Production of Type I Interferon via the TBK1-IRF3 Pathway

Cells ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 1205
Author(s):  
Tianhong Chen ◽  
Wenjuan Zhang ◽  
Bo Huang ◽  
Xuan Chen ◽  
Cao Huang
Keyword(s):  
Inflammatory Cytokines ◽  
Type I Interferon ◽  
Complete Loss ◽  
Functional Assay ◽  
Type I ◽  
Dependent Manner ◽  
Frontotemporal Degeneration ◽  
Dose Dependent ◽  
Lateral Sclerosis ◽  
Pro Inflammatory Cytokines

Mutations of Ubiquilin 2 (UBQLN2) or TANK-binding kinase 1 (TBK1) are associated with amyotrophic lateral sclerosis and frontotemporal degeneration (ALS/FTD). However, the mechanisms whereby UBQLN2 or TBK1 mutations lead to ALS and FTD remain unclear. Here, we explored the effect of UBQLN2 on TBK1 in HEK-293T cells or in CRISPR–Cas9-mediated IRF3 and IRF7 knockout (KO) cells. We found an interaction between TBK1 and UBQLN2, which was affected by ALS/FTD-linked mutations in TBK1 or UBQLN2. Co-expression of UBQLN2 with TBK1 elevated the protein level of TBK1 as well as the phosphorylation of TBK1 and IRF3 in a UBQLN2 dose-dependent manner, and this phosphorylation was reduced by mutant UBQLN2. In addition, the cellular production of IFN1 and related pro-inflammatory cytokines was substantially elevated when UBQLN2 and TBK1 were co-expressed, which was also decreased by mutant UBQLN2. Functional assay revealed that mutant UBQLN2 significantly reduced the binding affinity of TBK1 for its partners, including IRF3, (SQSTM1)/p62 and optineurin (OPTN). Moreover, complete loss of IRF3 abolished the induction of IFN1 and related pro-inflammatory cytokines enhanced by UBQLN2 in HEK-293T cells, whereas no significant change in IRF7 knockout cells was observed. Thus, our findings suggest that UBQLN2 promotes IRF3 phosphorylation via TBK1, leading to enhanced IFN1 induction, and also imply that the dysregulated TBK1-IRF3 pathway may play a role in UBQLN2-related neurodegeneration.

scholarly journals Streptococcus pneumoniae Autolysis Prevents Phagocytosis and Production of Phagocyte-Activating Cytokines

2009 ◽  
Vol 77 (9) ◽  
pp. 3826-3837 ◽  
Author(s):  
Anna Martner ◽  
Susann Skovbjerg ◽  
James C. Paton ◽  
Agnes E. Wold
Keyword(s):  
Streptococcus Pneumoniae ◽  
Mononuclear Cells ◽  
Human Peripheral Blood ◽  
Interleukin 12 ◽  
Dependent Manner ◽  
Peripheral Blood Mononuclear ◽  
Exact Function ◽  
Ifn Γ ◽  
Dose Dependent

ABSTRACT Streptococcus pneumoniae is a major pathogen in humans. The pathogenicity of this organism is related to its many virulence factors, the most important of which is the thick pneumococcal capsule that minimizes phagocytosis. Another virulence-associated trait is the tendency of this bacterium to undergo autolysis in stationary phase through activation of the cell wall-bound amidase LytA, which breaks down peptidoglycan. The exact function of autolysis in pneumococcal pathogenesis is, however, unclear. Here, we show the selective and specific inefficiency of wild-type S. pneumoniae for inducing production of phagocyte-activating cytokines in human peripheral blood mononuclear cells (PBMC). Indeed, clinical pneumococcal strains induced production of 30-fold less tumor necrosis factor (TNF), 15-fold less gamma interferon (IFN-γ), and only negligible amounts of interleukin-12 (IL-12) compared with other closely related Streptococcus species, whereas the levels of induction of IL-6, IL-8, and IL-10 production were similar. If pneumococcal LytA was inactivated by mutation or by culture in a medium containing excess choline, the pneumococci induced production of significantly more TNF, IFN-γ, and IL-12 in PBMC, whereas the production of IL-6, IL-8, and IL-10 was unaffected. Further, adding autolyzed pneumococci to intact bacteria inhibited production of TNF, IFN-γ, and IL-12 in a dose-dependent manner but did not inhibit production of IL-6, IL-8, and IL-10 in response to the intact bacteria. Fragments from autolyzed bacteria inhibited phagocytosis of intact bacteria and reduced the in vitro elimination of pneumococci from human blood. Our results suggest that fragments generated by autolysis of bacteria with reduced viability interfere with phagocyte-mediated elimination of live pneumococci.

scholarly journals Thrombasthenic mice generated by replacement of the integrin αIIb gene: demonstration that transcriptional activation of this megakaryocytic locus precedes lineage commitment

Blood ◽  
2000 ◽  
Vol 96 (4) ◽  
pp. 1399-1408 ◽  
Author(s):  
Diana Tronik-Le Roux ◽  
Valérie Roullot ◽  
Christel Poujol ◽  
Thierry Kortulewski ◽  
Paquita Nurden ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Fibrin Clot ◽  
Lineage Commitment ◽  
Gene Encoding ◽  
Α Subunit ◽  
Knock Out ◽  
Toxic Gene ◽  
Specific Locus ◽  
Deficient Mice

Abstract To analyze the transcriptional activity of the gene encoding the α subunit of the platelet integrin αIIbβ3during the hematopoietic differentiation, mice were produced in which the herpes virus thymidine kinase (tk) was introduced in this megakaryocytic specific locus using homologous recombination technology. This provided a convenient manner in which to induce the eradication of particular hematopoietic cells expressing the targeted gene. Results of progenitor cell cultures and long-term bone marrow (BM) assays showed that the growth of a subset of stem cells was reduced in the presence of the antiherpetic drug ganciclovir, demonstrating that the activation of the toxic gene occurs before the commitment to the megakaryocytic lineage. Furthermore theknock-in of the tk gene into the αIIb locus resulted in the knock-out of the αIIb gene in homozygous mice. Cultures of BM cells of these animals, combined with ultrastructural analysis, established that the αIIbglycoprotein is dispensable for lineage commitment and megakaryocytic maturation. Platelets collected from αIIb-deficient mice failed to bind fibrinogen, to aggregate, and to retract a fibrin clot. Moreover, platelet α-granules did not contain fibrinogen. Consistent with these characteristics, the mice displayed bleeding disorders similar to those in humans with Glanzmann thrombasthenia.

scholarly journals Lipopolysaccharide biosynthesis-related genes are required for colony pigmentation of Porphyromonas gingivalis

Microbiology ◽  
2009 ◽  
Vol 155 (4) ◽  
pp. 1282-1293 ◽  
Author(s):  
Keiko Sato ◽  
Nobuo Kido ◽  
Yukitaka Murakami ◽  
Charles I. Hoover ◽  
Koji Nakayama ◽  
...  
Keyword(s):  
Cell Surface ◽  
Porphyromonas Gingivalis ◽  
Dependent Manner ◽  
Wild Type ◽  
Gene Encoding ◽  
Lipopolysaccharide Biosynthesis ◽  
Ladder Pattern ◽  
Surface Polysaccharides ◽  
Culture Supernatants ◽  
Dose Dependent

The periodontopathic bacterium Porphyromonas gingivalis forms pigmented colonies when incubated on blood agar plates as a result of accumulation of μ-oxo haem dimer on the cell surface. Gingipain–adhesin complexes are responsible for production of μ-oxo haem dimer from haemoglobin. Non-pigmented mutants (Tn6-5, Tn7-1, Tn7-3 and Tn10-4) were isolated from P. gingivalis by Tn4351 transposon mutagenesis [Hoover & Yoshimura (1994), FEMS Microbiol Lett 124, 43–48]. In this study, we found that the Tn6-5, Tn7-1 and Tn7-3 mutants carried Tn4351 DNA in a gene homologous to the ugdA gene encoding UDP-glucose 6-dehydrogenase, a gene encoding a putative group 1 family glycosyltransferase and a gene homologous to the rfa gene encoding ADP heptose-LPS heptosyltransferase, respectively. The Tn10-4 mutant carried Tn4351 DNA at the same position as that for Tn7-1. Gingipain activities associated with cells of the Tn7-3 mutant (rfa) were very weak, whereas gingipain activities were detected in the culture supernatants. Immunoblot and mass spectrometry analyses also revealed that gingipains, including their precursor forms, were present in the culture supernatants. A lipopolysaccharide (LPS) fraction of the rfa deletion mutant did not show the ladder pattern that was usually seen for the LPS of the wild-type P. gingivalis. A recombinant chimera gingipain was able to bind to an LPS fraction of the wild-type P. gingivalis in a dose-dependent manner. These results suggest that the rfa gene product is associated with biosynthesis of LPS and/or cell-surface polysaccharides that can function as an anchorage for gingipain–adhesin complexes.

scholarly journals Spontaneous lysosomal enzyme secretion by a murine macrophage-like cell line

1980 ◽  
Vol 190 (3) ◽  
pp. 847-850 ◽  
Author(s):  
W Jessup ◽  
R T Dean
Keyword(s):  
Cell Line ◽  
Lysosomal Enzyme ◽  
Peritoneal Macrophages ◽  
Enzyme Secretion ◽  
Murine Macrophage ◽  
Lysosomal Hydrolase

Lysosomal enzyme secretion by the murine macrophage-like cell line, P388D1, was compared with that of normal peritoneal macrophages. Unlike macrophages, lysosomal hydrolase secretion by P388D1 cells occurred spontaneously in vitro and was not further stimulated by the presentation of inflammatory agents such as zymosan and asbestos.

PU.1 Plays a Critical Role in Maintenance of Mature Dendritic Cell Pool

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3554-3554
Author(s):  
Tadafumi Iino ◽  
Yong Chong ◽  
Shin-ichi Mizuno ◽  
Kyoko Ito ◽  
Daniel G. Tenen ◽  
...  
Keyword(s):  
Critical Role ◽  
Underlying Mechanism ◽  
Complete Loss ◽  
Lymphoid Cells ◽  
Lymphoid Tissues ◽  
Immune Disorders ◽  
Normal Numbers ◽  
Hematopoietic Stem ◽  
Cell Pool ◽  
Deficient Mice

Abstract PU.1, a hematopoietic transcription factor, is absolutely required for development of myelo-lymphoid cells from hematopoietic stem cells (HSC). PU.1-deficient mice fail to develop common myeloid progenitors (CMPs) or common lymphoid progenitors (CLPs), resulting in complete loss of dendritic cells (DC) in addition to mature myeloid and lymphoid cells. In this study, by disrupting PU.1 specifically at the mature DC stage, we here show that PU.1 is necessary for maintenance of mature DC pool. By crossing PU.1 floxed/floxed mice with a mouse line harboring the Cre transgene driven by the CD11c-BAC, we disrupted PU.1 at the CD11c+ DC stage. In these mice, development of DC precursors such as Lin−c-KitloFLT3+MCSFR+ DC progenitors, FLT3+ CLP and FLT3+CMP were not affected. The number of CD11c+B220− DCs, however, significantly reduced in all lymphoid tissues including the thymus, the spleen, the lymph node and the skin, down to <30%, <10%, <10% and <5% of DC numbers in control mice, respectively. In contrast, mice possessed normal numbers of granulocytes/monocytes, B cells, and naïve, effector or regulatory T cells. These mice have not developed any significant hematological or immune disorders at least until 6 months after birth. These results clearly show that PU.1 is required not only for DC development but also for maintenance of the peripheral DC pool. We are currently trying to elucidate the underlying mechanism for PU.1 to maintain mature DC numbers in peripheral organs.

Platelet endothelial cell adhesion molecule-1 is a negative regulator of platelet-collagen interactions

Blood ◽  
2001 ◽  
Vol 98 (5) ◽  
pp. 1456-1463 ◽  
Author(s):  
Karen L. Jones ◽  
Sascha C. Hughan ◽  
Sacha M. Dopheide ◽  
Richard W. Farndale ◽  
Shaun P. Jackson ◽  
...  
Keyword(s):  
Cell Adhesion Molecule ◽  
Collagen Matrix ◽  
Negative Regulator ◽  
Dependent Manner ◽  
Wild Type ◽  
Platelet Endothelial Cell Adhesion ◽  
Cell Adhesion Molecule 1 ◽  
Dose Dependent ◽  
Deficient Mice ◽  
Endothelial Cell Adhesion

The functional importance of platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) in platelets is unclear. Because PECAM-1 represents a newly assigned immunoglobulin–ITIM superfamily member expressed on the surface of platelets, it was hypothesized that it may play an important regulatory role in modulating ITAM-bearing receptors such as collagen (GP)VI receptor and FcγRIIA. To examine the functional role of PECAM-1 in regulating platelet-collagen interactions, 2 different approaches were applied using recombinant human PECAM-1–immunoglobulin chimeras and platelets derived from PECAM-1–deficient mice. Stimulation of platelets by collagen-, (GP)VI-selective agonist, collagen-related peptide (CRP)–, and PECAM-1–immunoglobulin chimera induced tyrosine phosphorylation of PECAM-1 in a time- and dose-dependent manner. Activation of PECAM-1 directly through the addition of soluble wild-type PECAM-1–immunoglobulin chimera, but not mutant K89A PECAM-1–immunoglobulin chimera that prevents homophilic binding, was found to inhibit collagen- and CRP-induced platelet aggregation. PECAM-1–deficient platelets displayed enhanced platelet aggregation and secretion responses on stimulation with collagen and CRP, though the response to thrombin was unaffected. Under conditions of flow, human platelet thrombus formation on a collagen matrix was reduced in a dose-dependent manner by human PECAM-1–immunoglobulin chimera. Platelets derived from PECAM-1–deficient mice form larger thrombi when perfused over a collagen matrix under flow at a shear rate of 1800 seconds−1 compared to wild-type mice. Collectively, these results indicate that PECAM-1 serves as a physiological negative regulator of platelet-collagen interactions that may function to negatively limit growth of platelet thrombi on collagen surfaces.

scholarly journals Impaired Activity of the Extraneuronal Monoamine Transporter System Known as Uptake-2 inOrct3/Slc22a3-Deficient Mice

2001 ◽  
Vol 21 (13) ◽  
pp. 4188-4196 ◽  
Author(s):  
Ronald Zwart ◽  
Sandra Verhaagh ◽  
Marije Buitelaar ◽  
Corrie Popp-Snijders ◽  
Denise P. Barlow
Keyword(s):  
Principal Component ◽  
Mutant Mice ◽  
In Vivo Model ◽  
Gene Encoding ◽  
Homozygous Mutant ◽  
Site Of Action ◽  
Extraneuronal Monoamine Transporter ◽  
Noradrenaline And Adrenaline ◽  
Deficient Mice

ABSTRACT Two uptake systems that control the extracellular concentrations of released monoamine neurotransmitters such as noradrenaline and adrenaline have been described. Uptake-1 is present at presynaptic nerve endings, whereas uptake-2 is extraneuronal and has been identified in myocardium and vascular and nonvascular smooth muscle cells. The gene encoding the uptake-2 transporter has recently been identified in humans (EMT), rats (OCT3), and mice (Orct3/Slc22a3). To generate an in vivo model for uptake-2, we have inactivated the mouseOrct3 gene. Homozygous mutant mice are viable and fertile with no obvious physiological defect and also show no significant imbalance of noradrenaline or dopamine. However,Orct3-null mice show an impaired uptake-2 activity as measured by accumulation of intravenously administered [3H]MPP+ (1-methyl-4-phenylpyridinium). A 72% reduction in MPP+ levels was measured in hearts of both male and female Orct3 mutant mice. No significant differences between wild-type and mutant mice were found in any other adult organ or in plasma. When [3H]MPP+ was injected into pregnant females, a threefold-reduced MPP+accumulation was observed in homozygous mutant embryos but not in their placentas or amniotic fluid. These data show that Orct3is the principal component for uptake-2 function in the adult heart and identify the placenta as a novel site of action of uptake-2 that acts at the fetoplacental interface.

scholarly journals A core function for p120-catenin in cadherin turnover

2003 ◽  
Vol 163 (3) ◽  
pp. 525-534 ◽  
Author(s):  
Michael A. Davis ◽  
Renee C. Ireton ◽  
Albert B. Reynolds
Keyword(s):  
Cell Adhesion ◽  
Cell Surface ◽  
Complete Loss ◽  
Vascular Endothelial ◽  
P120 Catenin ◽  
Epithelial Cadherin ◽  
Sirna Expression ◽  
Core Function ◽  
Dose Dependent ◽  
E Cadherin

p120-catenin stabilizes epithelial cadherin (E-cadherin) in SW48 cells, but the mechanism has not been established. Here, we show that p120 acts at the cell surface to control cadherin turnover, thereby regulating cadherin levels. p120 knockdown by siRNA expression resulted in dose-dependent elimination of epithelial, placental, neuronal, and vascular endothelial cadherins, and complete loss of cell–cell adhesion. ARVCF and δ-catenin were functionally redundant, suggesting that proper cadherin-dependent adhesion requires the presence of at least one p120 family member. The data reveal a core function of p120 in cadherin complexes, and strongly predict a dose-dependent loss of E-cadherin in tumors that partially or completely down-regulate p120.

Congenital Neuronal Ceroid Lipofuscinosis with a Novel CTSD Gene Mutation: A Rare Cause of Neonatal-Onset Neurodegenerative Disorder

2017 ◽  
Vol 49 (02) ◽  
pp. 150-153 ◽  
Author(s):  
K. Varvagiannis ◽  
S. Hanquinet ◽  
M. Billieux ◽  
R. De Luca ◽  
P. Rimensberger ◽  
...  
Keyword(s):  
Neurodegenerative Disorder ◽  
Age Of Onset ◽  
Neuronal Ceroid Lipofuscinosis ◽  
Molecular Genetic ◽  
Bioinformatic Analysis ◽  
Neuronal Ceroid Lipofuscinoses ◽  
Gene Encoding ◽  
Functional Studies ◽  
Ceroid Lipofuscinosis ◽  
Similar Phenotype

AbstractNeuronal ceroid lipofuscinoses represent a heterogeneous group of early onset neurodegenerative disorders that are characterized by progressive cognitive and motor function decline, visual loss, and epilepsy. The age of onset has been historically used for the phenotypic classification of this group of disorders, but their molecular genetic delineation has now enabled a better characterization, demonstrating significant genetic heterogeneity even among individuals with a similar phenotype. The rare Congenital Neuronal Ceroid Lipofuscinosis (CLN10) caused by mutations in the CTSD gene encoding for cathepsin D is associated with a dramatic presentation with onset before or around birth. We report on a female born to consanguineous parents who presented at birth with severe neonatal encephalopathy with massive cerebral and cerebellar shrinking on magnetic resonance imaging. Whole exome sequencing with targeted bioinformatic analysis of a panel of genes associated with prenatal/perinatal onset of neurodegenerative disease was performed and revealed the presence of a novel homozygous in-frame deletion in CTSD. Additional functional studies further confirmed the pathogenic character of this variant and established the diagnosis of CLN10 in the patient.

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