Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 20 de 57
Filter
1.
Mar Environ Res ; 193: 106297, 2024 Jan.
Article in English | MEDLINE | ID: mdl-38096713

ABSTRACT

Protoporphyrin IX (PPIX), a key precursor for the synthesis of chlorophyll and heme, is fundamental to photosynthetic eukaryotic cells and participates in light absorption, energy transduction, and numerous other cellular metabolic activities. Along with the application of genetic and biochemical techniques over the past few years, our understanding of the formation of PPIX has been largely advanced, especially regarding possible metabolic pathways. However, the ecological role and function of PPIX in natural ecosystems remains unclear. We have previously established a method for quantifying PPIX in marine ecosystems. Here, our results provide evidence that PPIX is not only subtly linked to nutrient uptake but also triggers phytoplankton productivity. PPIX and its derivatives are dynamic spatiotemporally in direct response to increased nutrient availability. Using 16 S rRNA gene amplicon sequencing, PPIX was revealed to interact strongly with many microorganisms, indicating that PPIX serves as a critical metabolite in maintaining microbial metabolism and community development. In summary, we observed that PPIX is linearly related to nutrient availability and microbial diversity. The levels of microbial PPIX reflect ecological health, and the availability of PPIX and nutrients jointly affect microbial community composition.


Subject(s)
Ecosystem , Protoporphyrins , Protoporphyrins/genetics , Protoporphyrins/metabolism , Heme/metabolism
2.
Br Poult Sci ; 63(3): 421-425, 2022 Jun.
Article in English | MEDLINE | ID: mdl-34585996

ABSTRACT

1. The goal of this study was to investigate the colour diversity of egg shells and expression of related genes in the uterus of chickens that produce eggs of different colours.2. Four colour types of Changshun blue-shell chickens, producing dark or light blue, greenish-brown and brown shelled eggs, were selected. The eggshell pigment concentration and colour values in each group were examined. The relative gene expression of solute carrier organic anion transporter family member 1C1 (SLCO1C1), ferrochelatase (FECH), haem oxygenase 1 (HO-1), ovotransferrin (OF) and biliverdin reductase A (BLVRA) in eggshell gland were measured.3. The Δb, ΔE and protoporphyrin in brown and greenish-brown groups were significantly higher in the blue egg group (P < 0.01), whereas ΔL was significantly lower than that in the blue eggs (P < 0.01). There was no significant difference in biliverdin concentration between the brown and blue groups.4. The Δa values, in descending order, were 8.27 ± 2.76 in the brown, -3.79 ± 2.39 in the greenish-brown and -7.29 ± 2.27 in the blue groups, respectively. The relative expression of HO-1 in the greenish-brown and light blue groups was significantly higher than in the dark blue and brown groups. The relative expression of FECH in the light blue group was significantly lower than that in the dark blue, greenish-brown or brown group (P < 0.01). The relative expression of HO-1 and BLVRA genes in the dark blue group was significantly higher than that in the light blue, greenish-brown and the brown group (P < 0.01).5. The Δa might provide a better index than protoporphyrin and biliverdin contents for eggshell colour breeding. Overall, HO-1 as well as BLVRA were important candidate genes for the selection of dark blue eggs.


Subject(s)
Chickens , Egg Shell , Animals , Biliverdine/genetics , Biliverdine/metabolism , Chickens/genetics , Chickens/metabolism , Female , Gene Expression , Ovum , Pigmentation/genetics , Protoporphyrins/genetics , Protoporphyrins/metabolism , Uterus/metabolism
3.
J Nippon Med Sch ; 87(6): 310-317, 2021 Jan 08.
Article in English | MEDLINE | ID: mdl-32238732

ABSTRACT

BACKGROUND: 5-Aminolevulinic Acid (5-ALA) photodiagnosis (PD) is an effective method to detect residual tumors during glioma surgery. However, fluorescence strength differs in malignant gliomas, and false-negative fluorescence may result in tumor residue. We investigated the effect of ultrasound on the intracellular level of protoporphyrin IX (PpIX) and expression level of ATP-binding cassette transporter 2 (ABCG2), which is thought to act as a membrane efflux pump of PpIX from cytosol. METHODS: The malignant glioma cell lines SNB19, U87MG, and T98G were used for in vitro experiments. Cultured cells underwent ultrasound irradiation (1 MHz, 3 W/cm2, duty cycle 10%) after administration of 5-ALA, and morphological changes in tumor cells were observed. PpIX levels and ABCG2 expression were evaluated. RESULTS: The glioma tumor cells showed transient morphological changes and detachment from the culture dish; however, most cells survived and reverted to their original morphology within 6 hours. PpIX expression levels increased in glioma cells after ultrasound irradiation, and the increase was earlier and greater than that for 5-ALA alone. ABCG2 expressions increased after 5-ALA administration but were lower in ultrasound-irradiated glioma cells. CONCLUSIONS: Ultrasound irradiation of malignant gliomas contributes to stronger 5-ALA-induced fluorescence by elevating intracellular PpIX levels. Suppression of ABCG2 expression by ultrasound may contribute to PpIX accumulation in glioma cells.


Subject(s)
ATP Binding Cassette Transporter, Subfamily G, Member 2/genetics , ATP Binding Cassette Transporter, Subfamily G, Member 2/metabolism , Fluorescence , Gene Expression , Glioma/diagnosis , Glioma/genetics , RNA, Messenger/genetics , RNA, Messenger/metabolism , Ultrasonography , Aminolevulinic Acid , Cell Line, Tumor , Glioma/pathology , Humans , Protoporphyrins/genetics , Protoporphyrins/metabolism
4.
Biochem J ; 477(23): 4635-4654, 2020 12 11.
Article in English | MEDLINE | ID: mdl-33211085

ABSTRACT

During bacteriochlorophyll a biosynthesis, the oxygen-independent conversion of Mg-protoporphyrin IX monomethyl ester (Mg-PME) to protochlorophyllide (Pchlide) is catalyzed by the anaerobic Mg-PME cyclase termed BchE. Bioinformatics analyses in combination with pigment studies of cobalamin-requiring Rhodobacter capsulatus mutants indicated an unusual radical S-adenosylmethionine (SAM) and cobalamin-dependent BchE catalysis. However, in vitro biosynthesis of the isocyclic ring moiety of bacteriochlorophyll using purified recombinant BchE has never been demonstrated. We established a spectroscopic in vitro activity assay which was subsequently validated by HPLC analyses and H218O isotope label transfer onto the carbonyl-group (C-131-oxo) of the isocyclic ring of Pchlide. The reaction product was further converted to chlorophyllide in the presence of light-dependent Pchlide reductase. BchE activity was stimulated by increasing concentrations of NADPH or SAM, and inhibited by S-adenosylhomocysteine. Subcellular fractionation experiments revealed that membrane-localized BchE requires an additional, heat-sensitive cytosolic component for activity. BchE catalysis was not sustained in chimeric experiments when a cytosolic extract from E. coli was used as a substitute. Size-fractionation of the soluble R. capsulatus fraction indicated that enzymatic activity relies on a specific component with an estimated molecular mass between 3 and 10 kDa. A structure guided site-directed mutagenesis approach was performed on the basis of a three-dimensional homology model of BchE. A newly established in vivo complementation assay was used to investigate 24 BchE mutant proteins. Potential ligands of the [4Fe-4S] cluster (Cys204, Cys208, Cys211), of SAM (Phe210, Glu308 and Lys320) and of the proposed cobalamin cofactor (Asp248, Glu249, Leu29, Thr71, Val97) were identified.


Subject(s)
Bacterial Proteins , Bacteriochlorophylls , Oxygenases , Protoporphyrins , Rhodobacter capsulatus , S-Adenosylmethionine , Bacterial Proteins/chemistry , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Bacteriochlorophylls/biosynthesis , Bacteriochlorophylls/chemistry , Bacteriochlorophylls/genetics , Oxygenases/chemistry , Oxygenases/genetics , Oxygenases/metabolism , Protoporphyrins/biosynthesis , Protoporphyrins/chemistry , Protoporphyrins/genetics , Rhodobacter capsulatus/chemistry , Rhodobacter capsulatus/genetics , Rhodobacter capsulatus/metabolism , S-Adenosylmethionine/chemistry , S-Adenosylmethionine/metabolism
5.
Biochim Biophys Acta Mol Cell Res ; 1867(11): 118817, 2020 11.
Article in English | MEDLINE | ID: mdl-32777371

ABSTRACT

Iron acquisition is challenging in most environments. As an alternative to elemental iron, organisms can take up iron-protoporphyrin IX, or heme. Heme can be found in decaying organic matter and is particularly prevalent in animal hosts. Fungi have evolved at least three distinct endocytosis-mediated heme uptake systems, which have been studied in detail in the organisms Candida albicans, Cryptococcus neoformans and Schizosaccharomyces pombe. Here we summarize the known molecular details of these three uptake systems that enable parasitic and saprophytic fungi to take advantage of external heme as either cellular iron or heme sources.


Subject(s)
Endocytosis/genetics , Heme/metabolism , Iron/metabolism , Protoporphyrins/metabolism , Candida albicans/metabolism , Cryptococcus neoformans/metabolism , Heme/genetics , Protoporphyrins/genetics , Schizosaccharomyces/metabolism , Signal Transduction/genetics
6.
Sci Rep ; 10(1): 3774, 2020 02 28.
Article in English | MEDLINE | ID: mdl-32111964

ABSTRACT

Hydrogen has the potential to play an important role in decarbonising our energy systems. Crucial to achieving this is the ability to produce clean sources of hydrogen using renewable energy sources. Currently platinum is commonly used as a hydrogen evolution catalyst, however, the scarcity and expense of platinum is driving the need to develop non-platinum-based catalysts. Here we report a protein-based hydrogen evolution catalyst based on a recombinant silk protein from honeybees and a metal macrocycle, cobalt protoporphyrin (CoPPIX). We enhanced the hydrogen evolution activity three fold compared to the unmodified silk protein by varying the coordinating ligands to the metal centre. Finally, to demonstrate the use of our biological catalyst, we built a proton exchange membrane (PEM) water electrolysis cell using CoPPIX-silk as the hydrogen evolution catalyst that is able to produce hydrogen with a 98% Faradaic efficiency. This represents an exciting advance towards allowing protein-based catalysts to be used in electrolysis cells.


Subject(s)
Bees/chemistry , Hydrogen/chemistry , Insect Proteins/chemistry , Metalloproteins/chemistry , Protoporphyrins/chemistry , Silk/chemistry , Animals , Bees/genetics , Catalysis , Insect Proteins/genetics , Metalloproteins/genetics , Protein Engineering , Protoporphyrins/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Silk/genetics
7.
Photodermatol Photoimmunol Photomed ; 36(1): 29-33, 2020 Jan.
Article in English | MEDLINE | ID: mdl-31374130

ABSTRACT

BACKGROUND: Erythropoietic protoporphyria (EPP) is a semi-dominantly inherited porphyria presenting with photosensitivity during early childhood. Acquired EPP has been reported; however, data regarding this rare disorder are scarce. PURPOSE: To evaluate the characteristics of acquired EPP. METHODS: A comprehensive search of PubMed, Google Scholar, ScienceDirect, and clinicaltrials.gov databases was performed by three reviewers. Studies describing patients with acquired EPP were included. Additionally, we present an index case of a 26-year-old patient who acquired clinically and biochemically typical EPP in association with myelodysplastic syndrome (MDS). RESULTS: We included 20 case reports describing 20 patients. Most (80%) patients were male of mean age 58 ± 13 years. In all patients, acquired EPP was associated with hematological disease, most commonly MDS (85%) followed by myeloproliferative disease (10%). In 86% of cases, hematological disease led to abnormality or somatic mutation in chromosome 18q (the locus of the ferrochelatase gene). The mean erythrocyte protoporphyrin IX concentration was very high (4286 µg/dL). Most (90%) patients presented with photosensitivity, 20% experienced blistering, and 25% presented with hepatic insufficiency, both uncommon in EPP. In 55% of patients, hematological disease was diagnosed after occurrence of cutaneous symptoms. Beta-carotene led to partial control of symptoms in 5 patients and resolution in another patient. Azacitidine treatment of MDS led to resolution of cutaneous symptoms in three patients. CONCLUSION: We present the distinct features of acquired EPP and highlight that any patient presenting with new-onset photosensitivity, irrespective of age should be evaluated for porphyria.


Subject(s)
Azacitidine/therapeutic use , Myelodysplastic Syndromes , Photosensitivity Disorders , Protoporphyria, Erythropoietic , beta Carotene/therapeutic use , Adult , Aged , Chromosomes, Human, Pair 18/genetics , Chromosomes, Human, Pair 18/metabolism , Erythrocytes/metabolism , Female , Ferrochelatase/genetics , Ferrochelatase/metabolism , Genetic Loci , Humans , Male , Middle Aged , Mutation , Myelodysplastic Syndromes/drug therapy , Myelodysplastic Syndromes/genetics , Myelodysplastic Syndromes/metabolism , Photosensitivity Disorders/chemically induced , Photosensitivity Disorders/drug therapy , Photosensitivity Disorders/genetics , Photosensitivity Disorders/metabolism , Protoporphyria, Erythropoietic/chemically induced , Protoporphyria, Erythropoietic/drug therapy , Protoporphyria, Erythropoietic/genetics , Protoporphyria, Erythropoietic/metabolism , Protoporphyrins/genetics , Protoporphyrins/metabolism
8.
Sci Rep ; 9(1): 20103, 2019 12 27.
Article in English | MEDLINE | ID: mdl-31882813

ABSTRACT

Neurological diseases have a close relationship to excessive reactive oxygen species (ROS). Neuroglobin (Ngb), an intrinsic protective factor, protected cells from hypoxic/ischemic injury. In the present, we reported a novel neuroprotective manganese porphyrin reconstituted metal protein, Mn-TAT PTD-Ngb, consisting of a HIV Tat protein transduction domain sequence (TAT PTD) attached to the N-terminal of apo-Ngb. Mn-TAT PTD-Ngb had a stronger ROS scavenging ability than that of TAT PTD-Ngb, and reduced intracellular ROS production and restored the function of the mitochondria and inhibited the mitochondria-dependent apoptosis. Besides, Mn-TAT PTD-Ngb activated the phosphoinositide-3 kinase (PI3K)/Akt signaling pathway, which up-regulated the expression of nuclear factor E2-related factor 2 (Nrf2), Heme oxygenase-1 (HO-1), superoxide dismutase (SOD), catalase (CAT). The results showed that the redox chemistry of Mn-TAT PTD-Ngb and redox regulation of multiple signaling pathways attenuated the oxidative injury.


Subject(s)
Oxidation-Reduction , Oxidative Stress , Protoporphyrins/metabolism , Reactive Oxygen Species/metabolism , Recombinant Fusion Proteins/metabolism , Signal Transduction , tat Gene Products, Human Immunodeficiency Virus/metabolism , Apoptosis , Humans , Hydrogen Peroxide/metabolism , Mitochondria/metabolism , Models, Biological , Protoporphyrins/genetics , Recombinant Fusion Proteins/genetics , tat Gene Products, Human Immunodeficiency Virus/genetics
9.
Sci Adv ; 5(9): eaaw6127, 2019 09.
Article in English | MEDLINE | ID: mdl-31555729

ABSTRACT

Erythropoietic protoporphyria (EPP) is an inherited disease caused by loss-of-function mutations of ferrochelatase, an enzyme in the heme biosynthesis pathway that converts protoporphyrin IX (PPIX) into heme. PPIX accumulation in patients with EPP leads to phototoxicity and hepatotoxicity, and there is no cure. Here, we demonstrated that the PPIX efflux transporter ABCG2 (also called BCRP) determines EPP-associated phototoxicity and hepatotoxicity. We found that ABCG2 deficiency decreases PPIX distribution to the skin and therefore prevents EPP-associated phototoxicity. We also found that ABCG2 deficiency protects against EPP-associated hepatotoxicity by modulating PPIX distribution, metabolism, and excretion. In summary, our work has uncovered an essential role of ABCG2 in the pathophysiology of EPP, which suggests the potential for novel strategies in the development of therapy for EPP.


Subject(s)
ATP Binding Cassette Transporter, Subfamily G, Member 2 , Protoporphyria, Erythropoietic , Protoporphyrins , Skin , ATP Binding Cassette Transporter, Subfamily G, Member 2/genetics , ATP Binding Cassette Transporter, Subfamily G, Member 2/metabolism , Animals , Ferrochelatase/genetics , Ferrochelatase/metabolism , Liver/metabolism , Liver/pathology , Mice , Mice, Mutant Strains , Protoporphyria, Erythropoietic/genetics , Protoporphyria, Erythropoietic/metabolism , Protoporphyria, Erythropoietic/pathology , Protoporphyria, Erythropoietic/physiopathology , Protoporphyrins/genetics , Protoporphyrins/metabolism , Skin/metabolism , Skin/pathology
10.
Mol Genet Metab ; 128(3): 391-395, 2019 11.
Article in English | MEDLINE | ID: mdl-30391163

ABSTRACT

Accumulation of protoporphyrin IX (PPIX) and Zn-PPIX, are the clinical hallmarks of protoporphyria. Phenotypic expression of protoporphyria is due to decreased activity of ferrochelatase (FECH) or to increased activity of aminolevulinic acid synthase (ALAS) in red blood cells. Other genetic defects have been shown to contribute to disease severity including loss of function mutations in the mitochondrial AAA-ATPase, CLPX and mutations in the Iron-responsive element binding protein 2 (IRP2), in mice. It is clear that multiple paths lead to a common phenotype of excess plasma PPIX that causes a phototoxic reaction on sun exposed areas. In this study we examined the association between mitochondrial iron acquisition and utilization with activity of FECH. Our data show that there is a metabolic link between the activity FECH and levels of MFRN1 mRNA. We examined the correlation between FECH activity and MFRN1 mRNA in cell lines established from patients with the classical protoporphyria, porphyria due to defects in ALAS2 mutations. Our data confirm MFRN1 message levels positively correlated with FECH enzymatic activity in all cell types.


Subject(s)
Cation Transport Proteins/genetics , Ferrochelatase/metabolism , Lymphocytes/enzymology , Mitochondria/physiology , Mitochondrial Proteins/genetics , Protoporphyria, Erythropoietic/enzymology , Protoporphyria, Erythropoietic/genetics , 5-Aminolevulinate Synthetase/genetics , Cell Line, Transformed , Ferrochelatase/analysis , Humans , Phenotype , Protoporphyrins/genetics , Protoporphyrins/metabolism , RNA, Messenger
11.
Arch Biochem Biophys ; 644: 37-46, 2018 04 15.
Article in English | MEDLINE | ID: mdl-29481781

ABSTRACT

Protoporphyrin ferrochelatase catalyzes the insertion of Fe2+ into protoporphyrin IX to form heme. To determine whether a conserved, active site π-helix contributes to the translocation of the metal ion substrate to the ferrochelatase-bound porphyrin substrate, the invariant π-helix glutamates were replaced with amino acids with non-negatively charged side chains, and the kinetic mechanisms of the generated variants were examined. Analysis of yeast wild-type ferrochelatase-, E314Q- and E318Q-catalyzed reactions, under multi- and single-turnover conditions, demonstrated that the mutations of the π-helix glutamates hindered both protoporphyrin metalation and release of the metalated porphyrin, by slowing each step by approximately 30-50%. Protoporphyrin metalation occurred with an apparent pKa of 7.3 ±â€¯0.1, which was assigned to binding of Fe2+ by deprotonated Glu-314 and Glu-314-assisted Fe2+ insertion into the porphyrin ring. We propose that unwinding of the π-helix concomitant with the adoption of a protein open conformation positions the deprotonated Glu-314 to bind Fe2+ from the surface of the enzyme. Transition to the closed conformation, with π-helix winding, brings Glu-314-bound Fe2+ to the active site for incorporation into protoporphyrin.


Subject(s)
Bacillus subtilis/enzymology , Bacterial Proteins/chemistry , Ferrochelatase/chemistry , Iron/chemistry , Protoporphyrins/chemistry , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae/enzymology , Animals , Bacillus subtilis/genetics , Bacterial Proteins/genetics , Ferrochelatase/genetics , Glutamic Acid/chemistry , Glutamic Acid/genetics , Humans , Mice , Mutation , Protein Structure, Secondary , Protoporphyrins/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae Proteins/genetics
12.
Sci Adv ; 4(1): eaaq1407, 2018 01.
Article in English | MEDLINE | ID: mdl-29387799

ABSTRACT

Chlorophylls are essential cofactors for photosynthesis, which sustains global food chains and oxygen production. Billions of tons of chlorophylls are synthesized annually, yet full understanding of chlorophyll biosynthesis has been hindered by the lack of characterization of the Mg-protoporphyrin IX monomethyl ester oxidative cyclase step, which confers the distinctive green color of these pigments. We demonstrate cyclase activity using heterologously expressed enzyme. Next, we assemble a genetic module that encodes the complete chlorophyll biosynthetic pathway and show that it functions in Escherichia coli. Expression of 12 genes converts endogenous protoporphyrin IX into chlorophyll a, turning E. coli cells green. Our results delineate a minimum set of enzymes required to make chlorophyll and establish a platform for engineering photosynthesis in a heterotrophic model organism.


Subject(s)
Biosynthetic Pathways , Escherichia coli , Metabolic Engineering , Protoporphyrins , Escherichia coli/enzymology , Escherichia coli/genetics , Protoporphyrins/biosynthesis , Protoporphyrins/genetics , Recombinant Proteins/biosynthesis , Recombinant Proteins/genetics
13.
Funct Integr Genomics ; 18(2): 175-194, 2018 Mar.
Article in English | MEDLINE | ID: mdl-29270875

ABSTRACT

Optimizing the antenna size by reducing the chlorophyll (Chl) content is an effective strategy to improve solar energy conversion efficiencies in dense crop monocultures. To elucidate the physiological and molecular mechanisms that regulate Chl biosynthesis and understand the effects of lower Chl content on the photosynthetic process, a light-intensity-dependent yellow-green wheat mutant (Jimai5265yg) was characterized to determine its morphological, histological, physiological, and transcriptional differences with wild type. In addition to lower Chl content with a higher Chl a/b ratio, Jimai5265yg has spherical chloroplasts with few plastoglobule. It is counterintuitive that the photochemical quantum yield of both photosystem I and photosystem II and the following CO2 assimilation rate significantly increased, but the value of nonphotochemical quenching decreased, indicating a reduction of the photoprotective capacity of this yellow-green mutant. Analysis of intermediate pools and the expression of genes in the Chl synthesis pathway indicated that Mg-protoporphyrin IX (Mg-Proto IX) synthesis was partially blocked due to the imbalanced expression of Mg-chelatase subunits. Interestingly, the expression of photosynthesis-associated nuclear genes (PhANGs) was upregulated, resembling gun mutants which have defects in the Mg-Proto IX-mediated plastid-to-nucleus signaling pathway. A genetic analysis indicated that the yellow-green phenotype was controlled by two nuclear recessive genes located on chromosomes 4AL and 4BL. Jimai5265yg is a novel chlorina mutant which could be used for understanding photosynthesis improvement mechanisms.


Subject(s)
Mutation , Photosynthesis/genetics , Transcriptome , Triticum/genetics , Chlorophyll/biosynthesis , Chlorophyll/genetics , Lyases/genetics , Lyases/metabolism , Plant Proteins/genetics , Plant Proteins/metabolism , Protoporphyrins/biosynthesis , Protoporphyrins/genetics , Triticum/metabolism , Up-Regulation
14.
Proc Natl Acad Sci U S A ; 114(38): E8045-E8052, 2017 09 19.
Article in English | MEDLINE | ID: mdl-28874591

ABSTRACT

Loss-of-function mutations in genes for heme biosynthetic enzymes can give rise to congenital porphyrias, eight forms of which have been described. The genetic penetrance of the porphyrias is clinically variable, underscoring the role of additional causative, contributing, and modifier genes. We previously discovered that the mitochondrial AAA+ unfoldase ClpX promotes heme biosynthesis by activation of δ-aminolevulinate synthase (ALAS), which catalyzes the first step of heme synthesis. CLPX has also been reported to mediate heme-induced turnover of ALAS. Here we report a dominant mutation in the ATPase active site of human CLPX, p.Gly298Asp, that results in pathological accumulation of the heme biosynthesis intermediate protoporphyrin IX (PPIX). Amassing of PPIX in erythroid cells promotes erythropoietic protoporphyria (EPP) in the affected family. The mutation in CLPX inactivates its ATPase activity, resulting in coassembly of mutant and WT protomers to form an enzyme with reduced activity. The presence of low-activity CLPX increases the posttranslational stability of ALAS, causing increased ALAS protein and ALA levels, leading to abnormal accumulation of PPIX. Our results thus identify an additional molecular mechanism underlying the development of EPP and further our understanding of the multiple mechanisms by which CLPX controls heme metabolism.


Subject(s)
5-Aminolevulinate Synthetase/metabolism , Endopeptidase Clp , Mutation, Missense , Porphyria, Erythropoietic , Protoporphyrins/biosynthesis , 5-Aminolevulinate Synthetase/genetics , Adolescent , Amino Acid Substitution , Endopeptidase Clp/genetics , Endopeptidase Clp/metabolism , Enzyme Stability/genetics , Female , Humans , Male , Porphyria, Erythropoietic/genetics , Porphyria, Erythropoietic/metabolism , Porphyria, Erythropoietic/pathology , Protoporphyrins/genetics
15.
Sci Rep ; 7(1): 3174, 2017 06 09.
Article in English | MEDLINE | ID: mdl-28600527

ABSTRACT

Waveguide based optofluidic resonator features high precision and high sensitivity in real-time fluorescent analysis. We present a novel optofluidic resonator following the hollow-core metal-cladding waveguide structure, which is then used to record the real-time binding process of Fe2+ and Fe3+ with protoporphyrin IX (PpIX) in PBS solution, respectively. The central fluorescent wavelength of compound with Fe2+ is in good accordance with that of the normal hemoglobin, whilst the peaks of the Fe3+ compound match the hemoglobin specimen from sickle-cell disease (SCD) patients. Similar statement holds when we monitor the real-time oxidation processes of these products by injecting oxygen into the optofluidic chip. These observations lead to the speculation that the SCD is caused by replacing the Fe2+ in hemoglobin with Fe3+, which may be insightful in the discovery of new clinical routes to cure this disease.


Subject(s)
Anemia, Sickle Cell/blood , Ferric Compounds/blood , Protoporphyrins/blood , Anemia, Sickle Cell/genetics , Anemia, Sickle Cell/pathology , Female , Hemoglobins/genetics , Hemoglobins/metabolism , Humans , Male , Protein Binding , Protoporphyrins/genetics
16.
PLoS One ; 12(1): e0171118, 2017.
Article in English | MEDLINE | ID: mdl-28129387

ABSTRACT

In photosynthesis, the pigments chlorophyll a/b absorb light energy to convert to chemical energy in chloroplasts. Though most enzymes of chlorophyll biosynthesis from glutamyl-tRNA to chlorophyll a/b have been identified, the exact composition and regulation of the multimeric enzyme Mg-protoporphyrin IX monomethyl ester cyclase (MPEC) is largely unknown. In this study, we isolated a rice pale-green leaf mutant m167 with yellow-green leaf phenotype across the whole lifespan. Chlorophyll content decreases 43-51% and the granal stacks of chloroplasts becomes thinner in m167. Chlorophyll fluorescence parameters, including Fv/Fm (the maximum quantum efficiency of PSII) and quantum yield of PSII (Y(II)), were lower in m167 than those in wild type plants (WT), and photosynthesis rate decreases 40% in leaves of m167 mutant compared with WT plants, which lead to yield reduction in m167. Genetic analysis revealed that yellow-green leaf phenotype of m167 is controlled by a single recessive genetic locus. By positional cloning, a single mutated locus, G286A (Alanine 96 to Threonine in protein), was found in the coding sequence of LOC_Os01g17170 (Rice Copper Response Defect 1, OsCRD1), encoding a putative subunit of MPEC. Expression profile analysis demonstrated that OsCRD1 is mainly expressed in green tissues of rice. Sequence alignment analysis of CRD1 indicated that Alanine 96 is very conserved in all green plants and photosynthetic bacteria. OsCRD1 protein mainly locates in chloroplast and the point mutation A96T in OsCRD1 does not change its location. Therefore, Alanine96 of OsCRD1 might be fundamental for MPEC activity, mutation of which leads to deficiency in chlorophyll biosynthesis and chloroplast development and decreases photosynthetic capacity in rice.


Subject(s)
Chlorophyll/genetics , Oryza/genetics , Photosynthesis/genetics , Plant Proteins/genetics , Chlorophyll/biosynthesis , Chloroplasts/enzymology , Chloroplasts/genetics , Chromosomes, Plant/genetics , Gene Expression Regulation, Plant/genetics , Oryza/growth & development , Phenotype , Plant Leaves/genetics , Plant Leaves/metabolism , Point Mutation/genetics , Protoporphyrins/genetics , Thylakoids/enzymology , Thylakoids/genetics
17.
Free Radic Biol Med ; 102: 111-121, 2017 01.
Article in English | MEDLINE | ID: mdl-27884704

ABSTRACT

The bystander effects of anti-cancer ionizing radiation have been widely studied, but far less is known about such effects in the case of non-ionizing photodynamic therapy (PDT). In the present study, we tested the hypothesis that photodynamically-stressed prostate cancer PC3 cells can elicit nitric oxide (NO)-mediated pro-growth/migration responses in non-stressed bystander cells. A novel approach was used whereby both cell populations existed on a culture dish, but made no physical contact with one other. Visible light irradiation of target cells sensitized with 5-aminolevulinic acid-induced protoporphyrin IX resulted in a striking upregulation of inducible nitric oxide synthase (iNOS) along with NO, the level of which increased after irradiation. Slower and less pronounced iNOS/NO upregulation was also observed in bystander cells. Activation of transcription factor NF-κB was implicated in iNOS induction in both targeted and bystander cells. Like surviving targeted cells, bystanders exhibited a significant increase in growth and migration rate, both responses being strongly attenuated by an iNOS inhibitor (1400W), a NO scavenger (cPTIO), or iNOS knockdown. Incubating bystander cells with conditioned medium from targeted cells failed to stimulate growth/migration, ruling out involvement of relatively long-lived stimulants. The following post-irradiation changes in pro-survival/pro-growth proteins were observed in bystander cells: upregulation of COX-2 and activation of protein kinases Akt and ERK1/2, NO again playing a key role. This is the first reported evidence for NO-enhanced bystander aggressiveness in the context of PDT. In the clinical setting, such effects could be averted through pharmacologic use of iNOS inhibitors as PDT adjuvants.


Subject(s)
Nitric Oxide Synthase Type II/genetics , Nitric Oxide/metabolism , Photochemotherapy , Prostatic Neoplasms/genetics , Amidines/administration & dosage , Apoptosis/drug effects , Apoptosis/radiation effects , Benzoates/administration & dosage , Benzylamines/administration & dosage , Bystander Effect/radiation effects , Cell Line, Tumor , Cell Movement/drug effects , Cell Movement/radiation effects , Cell Proliferation/drug effects , Cell Proliferation/radiation effects , Cyclooxygenase 2/genetics , Cyclooxygenase 2/metabolism , Enzyme Inhibitors/administration & dosage , Gene Expression Regulation, Neoplastic/radiation effects , Gene Knockdown Techniques , Humans , Imidazoles/administration & dosage , Light , Male , NF-kappa B/genetics , NF-kappa B/metabolism , Nitric Oxide/genetics , Nitric Oxide Synthase Type II/metabolism , Prostatic Neoplasms/pathology , Protoporphyrins/genetics , Protoporphyrins/metabolism
18.
Redox Biol ; 8: 333-40, 2016 08.
Article in English | MEDLINE | ID: mdl-26966892

ABSTRACT

Radioadaptive response (RAR) is an important phenomenon induced by low dose radiation. However, the molecular mechanism of RAR is obscure. In this study, we focused on the possible role of heme oxygenase 1 (HO-1) in RAR. Consistent with previous studies, priming dose of X-ray radiation (1-10cGy) induced significant RAR in normal human skin fibroblasts (AG 1522 cells). Transcription and translation of HO-1 was up-regulated more than two fold by a priming dose of radiation (5cGy). Zinc protoporphyrin Ⅸ, a specific competitive inhibitor of HO-1, efficiently inhibited RAR whereas hemin, an inducer of HO-1, could mimic priming dose of X-rays to induce RAR. Knocking down of HO-1 by transfection of HO-1 siRNA significantly attenuated RAR. Furthermore, the expression of HO-1 gene was modulated by the nuclear factor (erythroid-derived 2)-like 2 (Nrf2), which translocated from cytoplasm to nucleus after priming dose radiation and enhance the antioxidant level of cells.


Subject(s)
Antioxidants/metabolism , Fibroblasts/metabolism , Heme Oxygenase-1/genetics , NF-E2-Related Factor 2/genetics , Cell Line , Cell Nucleolus/metabolism , Cell Nucleolus/radiation effects , Cytoplasm/radiation effects , Dose-Response Relationship, Radiation , Fibroblasts/radiation effects , Gene Expression Regulation/radiation effects , Heme Oxygenase-1/antagonists & inhibitors , Hemin/genetics , Hemin/metabolism , Humans , NF-E2-Related Factor 2/metabolism , Protoporphyrins/genetics , Protoporphyrins/metabolism , RNA, Small Interfering/genetics , X-Rays
19.
FASEB J ; 30(5): 1798-810, 2016 05.
Article in English | MEDLINE | ID: mdl-26839379

ABSTRACT

Protoporphyria is a metabolic disease that causes excess production of protoporphyrin IX (PP-IX), the final biosynthetic precursor to heme. Hepatic PP-IX accumulation may lead to end-stage liver disease. We tested the hypothesis that systemic administration of porphyrin precursors to zebrafish larvae results in protoporphyrin accumulation and a reproducible nongenetic porphyria model. Retro-orbital infusion of PP-IX or the iron chelator deferoxamine mesylate (DFO), with the first committed heme precursor α-aminolevulinic acid (ALA), generates high levels of PP-IX in zebrafish larvae. Exogenously infused or endogenously produced PP-IX accumulates preferentially in the liver of zebrafish larvae and peaks 1 to 3 d after infusion. Similar to patients with protoporphyria, PP-IX is excreted through the biliary system. Porphyrin accumulation in zebrafish liver causes multiorganelle protein aggregation as determined by mass spectrometry and immunoblotting. Endoplasmic reticulum stress and induction of autophagy were noted in zebrafish larvae and corroborated in 2 mouse models of protoporphyria. Furthermore, electron microscopy of zebrafish livers from larvae administered ALA + DFO showed hepatocyte autophagosomes, nuclear membrane ruffling, and porphyrin-containing vacuoles with endoplasmic reticulum distortion. In conclusion, systemic administration of the heme precursors PP-IX or ALA + DFO into zebrafish larvae provides a new model of acute protoporphyria with consequent hepatocyte protein aggregation and proteotoxic multiorganelle alterations and stress.-Elenbaas, J. S., Maitra, D., Liu, Y., Lentz, S. I., Nelson, B., Hoenerhoff, M. J., Shavit, J. A., Omary, M. B. A precursor-inducible zebrafish model of acute protoporphyria with hepatic protein aggregation and multiorganelle stress.


Subject(s)
Disease Models, Animal , Protein Aggregation, Pathological/pathology , Protoporphyria, Erythropoietic/genetics , Protoporphyria, Erythropoietic/pathology , Stress, Physiological , Zebrafish , Aminolevulinic Acid/pharmacology , Animals , Deferoxamine/pharmacology , Genetic Predisposition to Disease , Larva/metabolism , Liver/metabolism , Liver/pathology , Mice , Photosensitizing Agents/pharmacology , Protoporphyrins/genetics , Protoporphyrins/metabolism , Siderophores/pharmacology
20.
J Pharmacol Exp Ther ; 356(2): 267-75, 2016 Feb.
Article in English | MEDLINE | ID: mdl-26588930

ABSTRACT

Protoporphyrin IX (PPIX) is ubiquitously present in all living cells in small amounts as a precursor of heme. PPIX has some biologic functions of its own, and PPIX-based strategies have been used for cancer diagnosis and treatment (the good). PPIX serves as the substrate for ferrochelatase, the final enzyme in heme biosynthesis, and its homeostasis is tightly regulated during heme synthesis. Accumulation of PPIX in human porphyrias can cause skin photosensitivity, biliary stones, hepatobiliary damage, and even liver failure (the bad and the ugly). In this work, we review the mechanisms that are associated with the broad aspects of PPIX. Because PPIX is a hydrophobic molecule, its disposition is by hepatic rather than renal excretion. Large amounts of PPIX are toxic to the liver and can cause cholestatic liver injury. Application of PPIX in cancer diagnosis and treatment is based on its photodynamic effects.


Subject(s)
Protoporphyrins/metabolism , Animals , Cholestasis/genetics , Cholestasis/metabolism , Cholestasis/pathology , Humans , Liver/metabolism , Liver/pathology , Photosensitivity Disorders/genetics , Photosensitivity Disorders/metabolism , Photosensitivity Disorders/pathology , Protoporphyrins/genetics
SELECTION OF CITATIONS
SEARCH DETAIL
...