scholarly journals Role of Phosphate Transport System Component PstB1 in Phosphate Internalization by Nostoc punctiforme

2016 ◽  
Vol 82 (21) ◽  
pp. 6344-6356 ◽  
Author(s):  
L. Hudek ◽  
D. Premachandra ◽  
W. A. J. Webster ◽  
L. Bräu
Keyword(s):  
Transport System ◽  
Phosphate Uptake ◽  
Phosphate Transport ◽  
Mrna Levels ◽  
Nostoc Punctiforme ◽  
Content Type ◽  
Phosphate Accumulation ◽  
Phosphate Availability ◽  
In Cells

ABSTRACTIn bacteria, limited phosphate availability promotes the synthesis of active uptake systems, such as the Pst phosphate transport system. To understand the mechanisms that facilitate phosphate accumulation in the cyanobacteriumNostoc punctiforme, phosphate transport systems were identified, revealing a redundancy of Pst phosphate uptake systems that exists across three distinct operons. Four separate PstB system components were identified.pstB1was determined to be a suitable target for creating phenotypic mutations that could result in the accumulation of excessive levels of phosphate through its overexpression or in a reduction of the capacity to accumulate phosphate through its deletion. Using quantitative real-time PCR (qPCR), it was determined thatpstB1mRNA levels increased significantly over 64 h in cells cultured in 0 mM added phosphate and decreased significantly in cells exposed to high (12.8 mM) phosphate concentrations compared to the level in cells cultured under normal (0.8 mM) conditions. Possible compensation for the loss of PstB1 was observed whenpstB2,pstB3, andpstB4mRNA levels increased, particularly in cells starved of phosphate. The overexpression ofpstB1increased phosphate uptake byN. punctiformeand was shown to functionally complement the loss of PstB inE. coliPstB knockout (PstB−) mutants. The knockout ofpstB1inN. punctiformedid not have a significant effect on cellular phosphate accumulation or growth for the most part, which is attributed to the compensation for the loss of PstB1 by alterations in thepstB2,pstB3, andpstB4mRNA levels. This study provides novelin vivoevidence that PstB1 plays a functional role in phosphate uptake inN. punctiforme.IMPORTANCECyanobacteria have been evolving over 3.5 billion years and have become highly adept at growing under limiting nutrient levels. Phosphate is crucial for the survival and prosperity of all organisms. In bacteria, limited phosphate availability promotes the synthesis of active uptake systems. The Pst phosphate transport system is one such system, responsible for the internalization of phosphate when cells are in phosphate-limited environments. Our investigations reveal the presence of multiple Pst phosphate uptake systems that exist across three distinct operons inNostoc punctiformeand functionally characterize the role of the gene product PstB1 as being crucial for the maintenance of phosphate accumulation. We demonstrate that the genespstB2,pstB3, andpstB4show alterations in expression to compensate for the deletion ofpstB1. The overall outcomes of this work provide insights as to the complex transport mechanisms that exist in cyanobacteria likeN. punctiforme, allowing them to thrive in low-phosphate environments.

Phosphate uptake by renal membrane vesicles of rabbits adapted to high and low phosphorus diets

1983 ◽  
Vol 245 (2) ◽  
pp. F175-F180 ◽  
Author(s):  
L. Cheng ◽  
C. T. Liang ◽  
B. Sacktor
Keyword(s):  
Transport System ◽  
Phosphate Uptake ◽  
Membrane Vesicles ◽  
Phosphate Transport ◽  
Uptake System ◽  
Low Phosphorus ◽  
Dietary Phosphate ◽  
Preferred Species ◽  
Uptake Medium ◽  
Phosphate Carrier

Renal adaptation to changes in phosphate intake was studied by comparing phosphate uptake by proximal tubule brush border membrane vesicles from rabbits on a relatively high or low phosphorus diet. The low phosphorus diet increased Na+ gradient-dependent phosphate uptake. Uptake in the absence of Na+ and in the presence of Na+, but no gradient, was not significantly affected. The phosphorus diet did not alter Na+ gradient-dependent D-glucose and L-proline uptake. The low phosphorus diet increased Vmax; affinity for phosphate was not appreciably changed. At all concentrations of extravesicular Na+, phosphate uptake was higher in membrane vesicles from animals fed the low phosphorus diet; the kinetics of the phosphate uptake system, with respect to Na+, was also altered by the change in dietary phosphate. These findings suggest that adaptation involves an alteration in the rate of translocation of the Na+-phosphate carrier when energized by a Na+ gradient driving force rather than a change in the number of Na+-phosphate carrier sites. With membrane vesicles from rabbits fed a low phosphorus diet, phosphate uptake increased several-fold when the pH of the uptake medium was raised, whereas with membrane vesicles from animals fed a high phosphorus diet the enhancement of uptake with alkalinization was relatively small. Irrespective of the diet, divalent phosphate was the probable preferred species for transport. Dietary adaptation was associated, however, with an alteration in the pH dependency of the transport system per se. These findings provide evidence that the adaptation of the kidney phosphate transport system to dietary phosphate load involves an intrinsic change in the Na+-phosphate carrier.

scholarly journals Silencing the Honey Bee (Apis mellifera) Naked Cuticle Gene (nkd) Improves Host Immune Function and Reduces Nosema ceranae Infections

2016 ◽  
Vol 82 (22) ◽  
pp. 6779-6787 ◽  
Author(s):  
Wenfeng Li ◽  
Jay D. Evans ◽  
Qiang Huang ◽  
Cristina Rodríguez-García ◽  
Jie Liu ◽  
...  
Keyword(s):  
Apis Mellifera ◽  
Immune Function ◽  
Honey Bee ◽  
Honey Bees ◽  
Nosema Ceranae ◽  
Negative Regulator ◽  
Mrna Levels ◽  
Content Type ◽  
Bee Disease

ABSTRACTNosema ceranaeis a new and emerging microsporidian parasite of European honey bees,Apis mellifera, that has been implicated in colony losses worldwide. RNA interference (RNAi), a posttranscriptional gene silencing mechanism, has emerged as a potent and specific strategy for controlling infections of parasites and pathogens in honey bees. While previous studies have focused on the silencing of parasite/pathogen virulence factors, we explore here the possibility of silencing a host factor as a mechanism for reducing parasite load. Specifically, we used an RNAi strategy to reduce the expression of a honey bee gene,naked cuticle(nkd), which is a negative regulator of host immune function. Our studies found thatnkdmRNA levels in adult bees were upregulated byN. ceranaeinfection (and thus, the parasite may use this mechanism to suppress host immune function) and that ingestion of double-stranded RNA (dsRNA) specific tonkdefficiently silenced its expression. Furthermore, we found that RNAi-mediated knockdown ofnkdtranscripts inNosema-infected bees resulted in upregulation of the expression of several immune genes (Abaecin,Apidaecin,Defensin-1, andPGRP-S2), reduction ofNosemaspore loads, and extension of honey bee life span. The results of our studies clearly indicate that silencing the hostnkdgene can activate honey bee immune responses, suppress the reproduction ofN. ceranae, and improve the overall health of honey bees. This study represents a novel host-derived therapeutic for honey bee disease treatment that merits further exploration.IMPORTANCEGiven the critical role of honey bees in the pollination of agricultural crops, it is urgent to develop strategies to prevent the colony decline induced by the infection of parasites/pathogens. Targeting parasites and pathogens directly by RNAi has been proven to be useful for controlling infections in honey bees, but little is known about the disease impacts of RNAi silencing of host factors. Here, we demonstrate that knocking down the honey bee immune repressor-encodingnkdgene can suppress the reproduction ofN. ceranaeand improve the overall health of honey bees, which highlights the potential role of host-derived and RNAi-based therapeutics in controlling the infections in honey bees. The information obtained from this study will have positive implications for honey bee disease management practices.

scholarly journals An RNA guanine quadruplex regulated pathway to TRAIL-sensitization by DDX21

2019 ◽  
Author(s):  
Ewan K.S. McRae ◽  
Steven J. Dupas ◽  
Evan P. Booy ◽  
Ramanaguru S. Piragasam ◽  
Richard P. Fahlman ◽  
...  
Keyword(s):  
Mrna Levels ◽  
Wild Type ◽  
Biologically Relevant ◽  
Protein Levels ◽  
Guanine Quadruplex ◽  
Repressive Effect ◽  
G Quadruplex ◽  
Different Levels ◽  
In Cells

AbstractDDX21 is a newly discovered RNA G-quadruplex (rG4) binding protein with no known biological rG4 targets. In this study we identified 26 proteins that are expressed at significantly different levels in cells expressing wild type DDX21 relative to an rG4 binding deficient DDX21 (M4). From this list we validate MAGED2 as a protein that is regulated by DDX21 through rG4 in its 5’UTR. MAGED2 protein levels, but not mRNA levels, are reduced by half in cells expressing only DDX21 M4. MAGED2 has a repressive effect on TRAIL-R2 expression that is relieved under these conditions, resulting in elevated TRAIL-R2 mRNA and protein in cells expressing only DDX21 M4, and rendering previously resistant cells sensitive to TRAIL mediated apoptosis. Our work identifies the role of DDX21 in regulation at the translational level through biologically relevant rG4 and shows that MAGED2 protein levels are regulated, at least in part, by a rG4 forming potential in their 5’UTRs.

scholarly journals The Staphylococcus aureus Chaperone PrsA Is a New Auxiliary Factor of Oxacillin Resistance Affecting Penicillin-Binding Protein 2A

2015 ◽  
Vol 60 (3) ◽  
pp. 1656-1666 ◽  
Author(s):  
Ambre Jousselin ◽  
Caroline Manzano ◽  
Alexandra Biette ◽  
Patricia Reed ◽  
Mariana G. Pinho ◽  
...  
Keyword(s):  
Binding Protein ◽  
Mrna Levels ◽  
Penicillin Binding Protein ◽  
Content Type ◽  
Oxacillin Resistance ◽  
Protein Membrane ◽  
Membrane Bound ◽  
Additional Level ◽  
Auxiliary Factor

Expression of the methicillin-resistantS. aureus(MRSA) phenotype results from the expression of the extra penicillin-binding protein 2A (PBP2A), which is encoded bymecAand acquired horizontally on part of the SCCmeccassette. PBP2A can catalyzedd-transpeptidation of peptidoglycan (PG) because of its low affinity for β-lactam antibiotics and can functionally cooperate with the PBP2 transglycosylase in the biosynthesis of PG. Here, we focus upon the role of the membrane-bound PrsA foldase protein as a regulator of β-lactam resistance expression. Deletion ofprsAaltered oxacillin resistance in three different SCCmecbackgrounds and, more importantly, caused a decrease in PBP2A membrane amounts without affectingmecAmRNA levels. The N- and C-terminal domains of PrsA were found to be critical features for PBP2A protein membrane levels and oxacillin resistance. We propose that PrsA has a role in posttranscriptional maturation of PBP2A, possibly in the export and/or folding of newly synthesized PBP2A. This additional level of control in the expression of themecA-dependent MRSA phenotype constitutes an opportunity to expand the strategies to design anti-infective agents.

Differential regulation of the expression of Na+/H+ exchanger isoform NHE3 by PKC-α in Caco-2 cells

2001 ◽  
Vol 281 (5) ◽  
pp. C1551-C1558 ◽  
Author(s):  
W. A. Alrefai ◽  
B. Scaglione-Sewell ◽  
S. Tyagi ◽  
L. Wartman ◽  
T. A. Brasitus ◽  
...  
Keyword(s):  
Mrna Level ◽  
Cell Types ◽  
Differential Regulation ◽  
Mrna Levels ◽  
Kinase C ◽  
External Signals ◽  
Nhe Isoforms ◽  
In Cells

Na+/H+ exchange (NHE) activity has been shown to be regulated by various external signals and protein kinases in many tissues and cell types. A family of six NHE isoforms has been identified. Three isoforms, NHE1, NHE2, and NHE3, have been shown to be expressed in the human intestine. The present studies were designed to study regulation of these human NHE isoforms by the α-isoform of protein kinase C (PKC) in the Caco-2 cell line. The mRNA levels of the NHE isoforms in Caco-2 cells were initially measured by a semiquantitative RT-PCR technique in response to PKC downregulation by long-term exposure to 1 μM 12- O-tetradecanoylphorbol-13-acetate (TPA) for 24 h. PKC downregulation resulted in an ∼60% increase in the mRNA level for NHE3, but not for NHE1 or NHE2. Utilizing dichlorobenzimidazole riboside, an agent to block the synthesis of new mRNA, we demonstrated that the increase in the NHE3 mRNA in response to downregulation of PKC was predominantly due to an increase in the rate of transcription, rather than a decrease in the NHE3 mRNA stability. Consistent with the mRNA results, our data showed that amiloride-sensitive22Na+ uptake was increased after incubation of Caco-2 cells with 1 μM TPA for 24 h. To elucidate the role of PKC-α, an isoform downregulated by TPA, the relative abundance of NHE isoform mRNA levels and the apical NHE activity were assessed in Caco-2 cells over- and underexpressing PKC-α. Our results demonstrated that NHE3, but not NHE1 or NHE2, mRNA was downregulated by PKC-α and that apical NHE activity was higher in cells underexpressing PKC-α and lower in cells overexpressing PKC-α than in control cells. In conclusion, these data demonstrate a differential regulation of NHE3, but not NHE2 or NHE1, expression by PKC in Caco-2 cells, and this regulation appears to be predominantly due to PKC-α.

scholarly journals Phosphate Transport and Sensing in Saccharomyces cerevisiae

Genetics ◽  
2001 ◽  
Vol 159 (4) ◽  
pp. 1491-1499 ◽  
Author(s):  
Dennis D Wykoff ◽  
Erin K O'Shea
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Inorganic Phosphate ◽  
Phosphate Uptake ◽  
Phosphate Starvation ◽  
Phosphate Transport ◽  
Glucose Sensors ◽  
Phosphate Transporters ◽  
Synthetic Lethal ◽  
High Phosphate

Abstract Cellular metabolism depends on the appropriate concentration of intracellular inorganic phosphate; however, little is known about how phosphate concentrations are sensed. The similarity of Pho84p, a high-affinity phosphate transporter in Saccharomyces cerevisiae, to the glucose sensors Snf3p and Rgt2p has led to the hypothesis that Pho84p is an inorganic phosphate sensor. Furthermore, pho84Δ strains have defects in phosphate signaling; they constitutively express PHO5, a phosphate starvation-inducible gene. We began these studies to determine the role of phosphate transporters in signaling phosphate starvation. Previous experiments demonstrated a defect in phosphate uptake in phosphate-starved pho84Δ cells; however, the pho84Δ strain expresses PHO5 constitutively when grown in phosphate-replete media. We determined that pho84Δ cells have a significant defect in phosphate uptake even when grown in high phosphate media. Overexpression of unrelated phosphate transporters or a glycerophosphoinositol transporter in the pho84Δ strain suppresses the PHO5 constitutive phenotype. These data suggest that PHO84 is not required for sensing phosphate. We further characterized putative phosphate transporters, identifying two new phosphate transporters, PHO90 and PHO91. A synthetic lethal phenotype was observed when five phosphate transporters were inactivated, and the contribution of each transporter to uptake in high phosphate conditions was determined. Finally, a PHO84-dependent compensation response was identified; the abundance of Pho84p at the plasma membrane increases in cells that are defective in other phosphate transporters.

scholarly journals The Oxygenase CAO-1 of Neurospora crassa Is a Resveratrol Cleavage Enzyme

2013 ◽  
Vol 12 (9) ◽  
pp. 1305-1314 ◽  
Author(s):  
Violeta Díaz-Sánchez ◽  
Alejandro F. Estrada ◽  
M. Carmen Limón ◽  
Salim Al-Babili ◽  
Javier Avalos
Keyword(s):  
Neurospora Crassa ◽  
Sequence Similarity ◽  
Hydroxyl Groups ◽  
Mrna Levels ◽  
Pronounced Increase ◽  
Content Type ◽  
Carotenoid Metabolism ◽  
Cleavage Enzyme ◽  
Β Carotene

ABSTRACT The genome of the ascomycete Neurospora crassa encodes CAO-1 and CAO-2, two members of the carotenoid cleavage oxygenase family that target double bonds in different substrates. Previous studies demonstrated the role of CAO-2 in cleaving the C 40 carotene torulene, a key step in the synthesis of the C 35 apocarotenoid pigment neurosporaxanthin. In this work, we investigated the activity of CAO-1, assuming that it may provide retinal, the chromophore of the NOP-1 rhodopsin, by cleaving β-carotene. For this purpose, we tested CAO-1 activity with carotenoid substrates that were, however, not converted. In contrast and consistent with its sequence similarity to family members that act on stilbenes, CAO-1 cleaved the interphenyl Cα-Cβ double bond of resveratrol and its derivative piceatannol. CAO-1 did not convert five other similar stilbenes, indicating a requirement for a minimal number of unmodified hydroxyl groups in the stilbene background. Confirming its biological function in converting stilbenes, adding resveratrol led to a pronounced increase in cao-1 mRNA levels, while light, a key regulator of carotenoid metabolism, did not alter them. Targeted Δ cao-1 mutants were not impaired by the presence of resveratrol, a phytoalexin active against different fungi, which did not significantly affect the growth and development of wild-type Neurospora . However, under partial sorbose toxicity, the Δ cao-1 colonies exhibited faster radial growth than control strains in the presence of resveratrol, suggesting a moderate toxic effect of resveratrol cleavage products.

scholarly journals Growth-Independent Regulation of CLN3mRNA Levels by Nutrients in Saccharomyces cerevisiae

1998 ◽  
Vol 180 (2) ◽  
pp. 225-230 ◽  
Author(s):  
Fereshteh Parviz ◽  
Warren Heideman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Transcriptional Control ◽  
Protein A ◽  
Nitrogen Sources ◽  
Mrna Levels ◽  
Rich Medium ◽  
Phase Growth ◽  
Protein Levels ◽  
In Cells

ABSTRACT Saccharomyces cerevisiae cells regulate progress through the G1 phase of the cell cycle in response to nutrients, moving quickly through G1 in rich medium and slowly in poor medium. Recent work has shown that the levels of Cln3 protein, a G1 cyclin, are low in cells growing in poor medium and high in cells growing rapidly in rich medium, consistent with the previously recognized role of Cln3 in promoting passage through Start. Cln3 protein levels appear to be regulated both transcriptionally and posttranscriptionally. We have worked to define the nutrient signals that regulate CLN3 mRNA levels. We find that CLN3 mRNA levels are high during log-phase growth in glucose medium, low in postdiauxic cells growing on ethanol, and slightly lower still in cells in stationary phase. CLN3mRNA levels are induced by glucose in a process that involves transcriptional control, requires metabolism of the glucose, and is independent of the Ras-cyclic AMP pathway. CLN3 mRNA levels are also positively regulated by nitrogen sources, but phosphorus and sulfur limitation do not affect CLN3 message levels.

scholarly journals The Ubiquitin Ligase Ubr11 Is Essential for Oligopeptide Utilization in the Fission Yeast Schizosaccharomyces pombe

2012 ◽  
Vol 11 (3) ◽  
pp. 302-310 ◽  
Author(s):  
Kenji Kitamura ◽  
Mai Nakase ◽  
Hideki Tohda ◽  
Kaoru Takegawa
Keyword(s):  
Schizosaccharomyces Pombe ◽  
Ubiquitin Ligase ◽  
Peptide Transporter ◽  
Mrna Levels ◽  
Content Type ◽  
Peptide Transporters ◽  
Naturally Occurring ◽  
Actual Mechanism ◽  
Transcriptional Corepressors

ABSTRACT Uptake of extracellular oligopeptides in yeast is mediated mainly by specific transporters of the peptide transporter (PTR) and oligopeptide transporter (OPT) families. Here, we investigated the role of potential peptide transporters in the yeast Schizosaccharomyces pombe . Utilization of naturally occurring dipeptides required only Ptr2/SPBC13A2.04c and none of the other 3 OPT proteins (Isp4, Pgt1, and Opt3), whereas only Isp4 was indispensable for tetrapeptide utilization. Both Ptr2 and Isp4 localized to the cell surface, but under rich nutrient conditions Isp4 localized in the Golgi apparatus through the function of the ubiquitin ligase Pub1. Furthermore, the ubiquitin ligase Ubr11 played a significant role in oligopeptide utilization. The mRNA levels of both the ptr2 and isp4 genes were significantly reduced in ubr11 Δ cells, and the dipeptide utilization defect in the ubr11 Δ mutant was rescued by the forced expression of Ptr2. Consistent with its role in transcriptional regulation of peptide transporter genes, the Ubr11 protein was accumulated in the nucleus. Unlike the situation in Saccharomyces cerevisiae , the oligopeptide utilization defect in the S. pombe ubr11 Δ mutant was not rescued by inactivation of the Tup11/12 transcriptional corepressors, suggesting that the requirement for the Ubr ubiquitin ligase in the upregulation of peptide transporter mRNA levels is conserved in both yeasts; however, the actual mechanism underlying the control appears to be different. We also found that the peptidomimetic proteasome inhibitor MG132 was still operative in a strain lacking all known PTR and OPT peptide transporters. Therefore, irrespective of its peptide-like structure, MG132 is carried into cells independently of the representative peptide transporters.

Sign in / Sign up

Export Citation Format

Share Document