scholarly journals The Nitric Oxide Donor, S-Nitrosoglutathione, Rescues Peroxisome Number and Activity Defects in PEX1G843D Mild Zellweger Syndrome Fibroblasts

2021 ◽  
Vol 9 ◽  
Author(s):  
Yidi Liu ◽  
Ceileigh M. Weaver ◽  
Yarina Sen ◽  
Gary Eitzen ◽  
Andrew J. Simmonds ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Zellweger Syndrome ◽  
Nitric Oxide Donor ◽  
Metabolic Dysfunction ◽  
Peroxisome Biogenesis Disorders ◽  
Genes Encoding ◽  
Strong Candidate ◽  
And Function

Peroxisome biogenesis disorders (PBDs) are a group of metabolic developmental diseases caused by mutations in one or more genes encoding peroxisomal proteins. Zellweger syndrome spectrum (PBD-ZSS) results from metabolic dysfunction caused by damaged or non-functional peroxisomes and manifests as a multi-organ syndrome with significant morbidity and mortality for which there is no current drug therapy. Mild PBD-ZSS patients can exhibit a more progressive disease course and could benefit from the identification of drugs to improve the quality of life and extend the lifespan of affected individuals. Our study used a high-throughput screen of FDA-approved compounds to identify compounds that improve peroxisome function and biogenesis in human fibroblast cells carrying the mild PBD-ZSS variant, PEX1G843D. Our screen identified the nitrogen oxide donor, S-nitrosoglutathione (GSNO), as a potential therapeutic for this mild form of PBD-ZSS. Further biochemical characterization showed that GSNO enhances both peroxisome number and function in PEX1G843D mutant fibroblasts and leads to increased survival and longer lifespan in an in vivo humanized Drosophila model carrying the PEX1G843D mutation. GSNO is therefore a strong candidate to be translated to clinical trials as a potential therapeutic for mild PBD-ZSS.

Phenotype–genotype relationships in peroxisome biogenesis disorders of PEX1-defective complementation group 1 are defined by Pex1p–Pex6p interaction

2001 ◽  
Vol 357 (2) ◽  
pp. 417-426 ◽  
Author(s):  
Shigehiko TAMURA ◽  
Naomi MATSUMOTO ◽  
Atsushi IMAMURA ◽  
Nobuyuki SHIMOZAWA ◽  
Yasuyuki SUZUKI ◽  
...  
Keyword(s):  
Zellweger Syndrome ◽  
Complementation Group ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Peroxisome Biogenesis ◽  
Causative Gene ◽  
Aaa Atpase ◽  
Peroxisome Biogenesis Disorders ◽  
The Stability

The peroxisome biogenesis disorders (PBDs), including Zellweger syndrome (ZS), neonatal adrenoleucodystrophy (NALD) and infantile Refsum disease (IRD), are fatal autosomal recessive diseases caused by impaired peroxisome biogenesis, of which 12 genotypes have been reported. ZS patients manifest the severest clinical and biochemical abnormalities, whereas those with NALD and IRD show less severity and the mildest features respectively. We have reported previously that temperature-sensitive peroxisome assembly is responsible for the mildness of the clinical features of IRD. PEX1 is the causative gene for PBDs of complementation group E (CG-E, CG1 in the U.S.A. and Europe), the PBDs of highest incidence, encoding the peroxin Pex1p of the AAA ATPase family. It has been also reported that Pex1p and Pex6p interact with each other. In the present study we investigated phenotype–genotype relationships of CG1 PBDs. Pex1p from IRD such as Pex1p with the most frequently identified mutation at G843D was largely degraded in vivo at 37°C, whereas a normal level of Pex1p was detectable at the permissive temperature. In contrast, PEX1 proteins derived from ZS patients, including proteins with a mutation at L664P or the deletion of residues 634–690, were stably present at both temperatures. Pex1p-G843D interacted with Pex6p at approx. 50% of the level of normal Pex1p, whereas Pex1p from ZS patients mostly showing non-temperature-sensitive peroxisome biogenesis hardly bound to Pex6p. Taking these results together, we consider it most likely that the stability of Pex1p reflects temperature-sensitive peroxisome assembly in IRD fibroblasts. Failure in Pex1p–Pex6p interaction gives rise to more severe abnormalities, such as those manifested by patients with ZS.

scholarly journals Reconstitution of the Human tRNA Splicing Endonuclease Complex: insight into the regulation of pre-tRNA cleavage

2019 ◽  
Author(s):  
Cassandra K. Hayne ◽  
Casey A. Schmidt ◽  
A. Gregory Matera ◽  
Robin E. Stanley
Keyword(s):  
Animal Cells ◽  
Trna Splicing ◽  
Polynucleotide Kinase ◽  
Genes Encoding ◽  
Intron Removal ◽  
And Function ◽  
Insight Into ◽  
Negative Modulator

ABSTRACTThe splicing of tRNA introns is a critical step in pre-tRNA maturation. In archaea and eukaryotes, tRNA intron removal is catalyzed by the tRNA splicing endonuclease (TSEN) complex. Eukaryotic TSEN is comprised of four core subunits (TSEN54, TSEN2, TSEN34, and TSEN15). The human TSEN complex additionally co-purifies with the polynucleotide kinase CLP1; however, CLP1’s role in tRNA splicing remains unclear. Mutations in genes encoding all four TSEN subunits, as well as CLP1, are known to cause neurodegenerative disorders, yet the mechanisms underlying the pathogenesis of these disorders are unknown. Here, we developed a recombinant system that produces active TSEN complex. Co-expression of all four TSEN subunits is required for efficient formation and function of the complex. We show that human CLP1 associates with the active TSEN complex, but is not required for tRNA intron cleavage in vitro. Moreover, RNAi knockdown of the Drosophila CLP1 orthologue, cbc, promotes biogenesis of mature tRNAs and circularized tRNA introns (tricRNAs) in vivo. Collectively, these and other findings suggest that CLP1/cbc plays a regulatory role in tRNA splicing by serving as a negative modulator of the direct tRNA ligation pathway in animal cells.

scholarly journals Reconstitution of the human tRNA splicing endonuclease complex: insight into the regulation of pre-tRNA cleavage

2020 ◽  
Vol 48 (14) ◽  
pp. 7609-7622 ◽  
Author(s):  
Cassandra K Hayne ◽  
Casey A Schmidt ◽  
Maira I Haque ◽  
A Gregory Matera ◽  
Robin E Stanley
Keyword(s):  
Animal Cells ◽  
Trna Splicing ◽  
Polynucleotide Kinase ◽  
Genes Encoding ◽  
Intron Removal ◽  
And Function ◽  
Insight Into ◽  
Negative Modulator

Abstract The splicing of tRNA introns is a critical step in pre-tRNA maturation. In archaea and eukaryotes, tRNA intron removal is catalyzed by the tRNA splicing endonuclease (TSEN) complex. Eukaryotic TSEN is comprised of four core subunits (TSEN54, TSEN2, TSEN34 and TSEN15). The human TSEN complex additionally co-purifies with the polynucleotide kinase CLP1; however, CLP1’s role in tRNA splicing remains unclear. Mutations in genes encoding all four TSEN subunits, as well as CLP1, are known to cause neurodegenerative disorders, yet the mechanisms underlying the pathogenesis of these disorders are unknown. Here, we developed a recombinant system that produces active TSEN complex. Co-expression of all four TSEN subunits is required for efficient formation and function of the complex. We show that human CLP1 associates with the active TSEN complex, but is not required for tRNA intron cleavage in vitro. Moreover, RNAi knockdown of the Drosophila CLP1 orthologue, cbc, promotes biogenesis of mature tRNAs and circularized tRNA introns (tricRNAs) in vivo. Collectively, these and other findings suggest that CLP1/cbc plays a regulatory role in tRNA splicing by serving as a negative modulator of the direct tRNA ligation pathway in animal cells.

scholarly journals Copper binding by a unique family of metalloproteins is dependent on kynurenine formation

2021 ◽  
Vol 118 (23) ◽  
pp. e2100680118
Author(s):  
Anastasia C. Manesis ◽  
Richard J. Jodts ◽  
Brian M. Hoffman ◽  
Amy C. Rosenzweig
Keyword(s):  
Biochemical Characterization ◽  
Protein A ◽  
Copper Ion ◽  
Copper Homeostasis ◽  
Metabolic Enzyme ◽  
Copper Binding ◽  
Methane Oxidizing Bacteria ◽  
And Function

Some methane-oxidizing bacteria use the ribosomally synthesized, posttranslationally modified natural product methanobactin (Mbn) to acquire copper for their primary metabolic enzyme, particulate methane monooxygenase. The operons encoding the machinery to biosynthesize and transport Mbns typically include genes for two proteins, MbnH and MbnP, which are also found as a pair in other genomic contexts related to copper homeostasis. While the MbnH protein, a member of the bacterial diheme cytochrome c peroxidase (bCcP)/MauG superfamily, has been characterized, the structure and function of MbnP, the relationship between the two proteins, and their role in copper homeostasis remain unclear. Biochemical characterization of MbnP from the methanotroph Methylosinus trichosporium OB3b now reveals that MbnP binds a single copper ion, present in the +1 oxidation state, with high affinity. Copper binding to MbnP in vivo is dependent on oxidation of the first tryptophan in a conserved WxW motif to a kynurenine, a transformation that occurs through an interaction of MbnH with MbnP. The 2.04-Å-resolution crystal structure of MbnP reveals a unique fold and an unusual copper-binding site involving a histidine, a methionine, a solvent ligand, and the kynurenine. Although the kynurenine residue may not serve as a CuI primary-sphere ligand, being positioned ∼2.9 Å away from the CuI ion, its presence is required for copper binding. Genomic neighborhood analysis indicates that MbnP proteins, and by extension kynurenine-containing copper sites, are widespread and may play diverse roles in microbial copper homeostasis.

scholarly journals Extracellular matrix regulates morphogenesis and function of ciliated sensory organs in Caenorhabditis elegans

2018 ◽  
Author(s):  
Deanna M. De Vore ◽  
Karla M. Knobel ◽  
Ken C.Q. Nguyen ◽  
David H. Hall ◽  
Maureen M. Barr
Keyword(s):  
Extracellular Matrix ◽  
Dendritic Morphology ◽  
C Elegans ◽  
Genes Encoding ◽  
Health And Disease ◽  
Autosomal Dominant Polycystic Kidney ◽  
Signaling Organelles ◽  
And Function

ABSTRACTCilia and extracellular vesicles (EVs) are signaling organelles that play important roles in human health and disease. In C. elegans and mammals, the Autosomal Dominant Polycystic Kidney Disease (ADPKD) gene products polycystin-1 and polycystin-2 localize to both cilia and EVs, act in the same genetic pathway, and function in a sensory capacity, suggesting ancient conservation. Hence, the nematode offers an excellent system in which to address central questions regarding the biology of cilia, EVs, and the polycystins. We discovered an unexpected role of the mec-1, mec-5, and mec-9 genes encoding extracellular matrix (ECM) components. We determined that these ECM encoding genes regulate polycystin localization and function, ciliary EV release, cilia length, dendritic morphology, and neuron-glia interactions. Abnormal ECM and fibrosis are observed in ciliopathies such as ADPKD, nephronophthisis, and Bardet-Biedl Syndrome. Our studies reveal multifaceted roles for ECM proteins in the ciliated nervous system of the worm and provide a powerful new in vivo model to study the relationship between ECM, the polycystins, and ciliopathies.

scholarly journals Subpopulation-specific differences in skeletal muscle mitochondria in humans with obesity: insights from studies employing acute nutritional and exercise stimuli

2020 ◽  
Vol 318 (4) ◽  
pp. E538-E553
Author(s):  
Nisreen Wahwah ◽  
Katon A. Kras ◽  
Lori R. Roust ◽  
Christos S. Katsanos
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Function ◽  
Subcellular Location ◽  
Metabolic Dysfunction ◽  
Muscle Mitochondria ◽  
Skeletal Muscle Mitochondria ◽  
Experimental Approaches ◽  
And Function

Mitochondria from skeletal muscle of humans with obesity often display alterations with respect to their morphology, proteome, biogenesis, and function. These changes in muscle mitochondria are considered to contribute to metabolic abnormalities observed in humans with obesity. Most of the evidence describing alterations in muscle mitochondria in humans with obesity, however, lacks reference to a specific subcellular location. This is despite data over the years showing differences in the morphology and function of subsarcolemmal (found near the plasma membrane) and intermyofibrillar (nested between the myofibrils) mitochondria in skeletal muscle. Recent studies reveal that impairments in mitochondrial function in obesity with respect to the subcellular location of the mitochondria in muscle are more readily evident following exposure of the skeletal muscle to physiological stimuli. In this review, we highlight the need to understand skeletal muscle mitochondria metabolism in obesity in a subpopulation-specific manner and in the presence of physiological stimuli that modify mitochondrial function in vivo. Experimental approaches employed under these conditions will allow for more precise characterization of impairments in skeletal muscle mitochondria and their implications in inducing metabolic dysfunction in human obesity.

scholarly journals The emergence of protein arginine methyltransferases in skeletal muscle and metabolic disease

2019 ◽  
Vol 317 (6) ◽  
pp. E1070-E1080 ◽  
Author(s):  
Tiffany L. vanLieshout ◽  
Vladimir Ljubicic
Keyword(s):  
Skeletal Muscle ◽  
Cell Function ◽  
In Vivo Studies ◽  
Whole Body ◽  
Metabolic Dysfunction ◽  
Protein Arginine Methyltransferases ◽  
Arginine Residues ◽  
Whole Body Metabolism ◽  
And Function

Protein arginine methyltransferases (PRMTs) are a family of enzymes that catalyze the methylation of arginine residues on target proteins and thus alter the stability, localization, or activity of the substrate. In doing so, PRMTs mediate a variety of intracellular functions that are essential for survival. Additionally, PRMT dysregulation is involved in a number of the most prevalent health disorders, including cancer and neurodegenerative and cardiovascular diseases, as well as in the aging process. Investigations of PRMT biology in skeletal muscle cells began in 2002, and since then these enzymes have emerged as regulators of skeletal muscle phenotype determination, maintenance, and remodeling. Specifically, more recent in vivo studies have revealed that PRMTs impact multiple aspects of skeletal muscle biology, including satellite cell function and phenotypic plasticity in response to exercise and disuse. Skeletal muscle plays critically important roles in regulating whole body metabolism, and recent investigations have also begun elucidating PRMT expression and function under conditions of metabolic dysfunction. The goals of this review are to 1) summarize the literature on PRMT biology in skeletal muscle with a particular emphasis on the in vivo evidence and 2) survey PRMTs in metabolic disorders, namely, obesity and type 2 diabetes mellitus. We also identify notable knowledge gaps therein and present opportunities to further expand our understanding of these enzymes so critical to health and disease.

scholarly journals Identification and characterization of genes encoding the nuclear envelope LINC complex in the monocot species Zea mays

2018 ◽  
Author(s):  
Hardeep K. Gumber ◽  
Joseph F. McKenna ◽  
Amado L. Estrada ◽  
Andrea F. Tolmie ◽  
Katja Graumann ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Nuclear Periphery ◽  
Coiled Coil ◽  
Gene Families ◽  
Grass Species ◽  
Linc Complex ◽  
Genes Encoding ◽  
Model Grass ◽  
And Function

ABSTRACTThe LINC (Linker of Nucleoskeleton to Cytoskeleton) complex is an essential multi-protein structure spanning the nuclear envelope. It connects the cytoplasm to the nucleoplasm, functions to maintain nuclear shape and architecture, and regulates chromosome dynamics during cell division. Knowledge of LINC complex composition and function in the plant kingdom is primarily limited to Arabidopsis, but critically missing from the evolutionarily distant monocots which include grasses, the most important agronomic crops worldwide. To fill this knowledge gap, we identified and characterized 22 maize genes, including a new grass-specific KASH gene family. Using bioinformatic, biochemical, and cell biological approaches, we provide evidence that representative KASH candidates localize to the nuclear periphery and interact with ZmSUN2 in vivo. FRAP experiments using domain-deletion constructs verified that this SUN-KASH interaction was dependent on the SUN but not the coiled-coil domain of ZmSUN2. A summary working model is proposed for the entire maize LINC complex encoded by conserved and divergent gene families. These findings expand our knowledge of the plant nuclear envelope in a model grass species, with implications for both basic and applied cellular research.SUMMARY STATEMENTGenes encoding maize candidates for the core LINC and associated complex proteins have been comprehensively identified with functional validation by one or more assays for several of the KASH genes.

scholarly journals R-MC46 monoclonal antibody stimulates adhesion and phagocytosis by rat macrophages

2004 ◽  
Vol 61 (6) ◽  
pp. 581-588
Author(s):  
Sonja Gasic ◽  
Dragana Vucevic ◽  
Petar Popovic ◽  
Sasa Vasilijic ◽  
Miodrag Colic
Keyword(s):  
Molecular Weight ◽  
Monoclonal Antibody ◽  
Western Blot ◽  
Biochemical Characterization ◽  
Antigen Expression ◽  
Peritoneal Macrophages ◽  
Low Molecular Weight ◽  
And Function ◽  
Stimulation Of

Background. In our previous experiments it was shown that R-MC46 monoclonal antibody (mAb), produced at our Institute, stimulated homotypic aggregation of rat granulocytes and production of proinflammatory cytokines. The aim of this study was to examine antigen expression and function, recognized by R-MC46 mAb on macrophages. Methods. The expression of R-MC46 antigen on thymic and peritoneal macrophages was investigated using immunocytochemistry and flow cytometry methods. Its biochemical characterization was performed by Western blot. The ability of R-MC46 mAb to modulate adhesion and phagocytosis by macrophages was studied by using co-culture experiments with autologous thymocytes. Results. R-MC46 mAb stained thymic macrophages more strongly than peritoneal macrophages. After in vivo treatment of peritoneal macrophages with Pristane, a significant up-regulation of the R-MC46 antigen expression was observed. Western blot analysis showed that the mAb recognized a low molecular weight antigen of about 5.5 kDa. R-MC46 mAb significantly enhanced binding and phagocytosis of thymocytes by both thymic and peritoneal macrophages. These processes were completely blocked by WT.3 (anti-CD18) mAb. The stimulation of binding thymocyte to macrophages was higher with the use of thymic macrophages,while the phagocytosis of these cells was higher in the presence of peritoneal macrophages. Conclusion. R-MC46 mAb recognized a new molecule expressed by rat macrophages. The antigen is most probably involved in ?2 integrin-mediated adhesion and phagocytosis, as well as proinflammatory functions of macrophages.

Osseointegration interface of calcified bone and endosseous implants

1992 ◽  
Vol 50 (2) ◽  
pp. 1096-1097
Author(s):  
K.E. Krizan ◽  
J.E. Laffoon ◽  
M.J. Buckley
Keyword(s):  
Structure And Function ◽  
Titanium Implants ◽  
En Bloc ◽  
Endosseous Implants ◽  
Ultrastructural Studies ◽  
The Past ◽  
Cacodylate Buffer ◽  
One Year ◽  
And Function

With increase use of tissue-integrated prostheses in recent years it is a goal to understand what is happening at the interface between haversion bone and bulk metal. This study uses electron microscopy (EM) techniques to establish parameters for osseointegration (structure and function between bone and nonload-carrying implants) in an animal model. In the past the interface has been evaluated extensively with light microscopy methods. Today researchers are using the EM for ultrastructural studies of the bone tissue and implant responses to an in vivo environment. Under general anesthesia nine adult mongrel dogs received three Brånemark (Nobelpharma) 3.75 × 7 mm titanium implants surgical placed in their left zygomatic arch. After a one year healing period the animals were injected with a routine bone marker (oxytetracycline), euthanized and perfused via aortic cannulation with 3% glutaraldehyde in 0.1M cacodylate buffer pH 7.2. Implants were retrieved en bloc, harvest radiographs made (Fig. 1), and routinely embedded in plastic. Tissue and implants were cut into 300 micron thick wafers, longitudinally to the implant with an Isomet saw and diamond wafering blade [Beuhler] until the center of the implant was reached.

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