Browse
Submit Data
Databases
API
Help

Dataset Information

15 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Sodium bicarbonate transporter NBCe1 regulates proliferation and viability of human prostate cancer cells LNCaP and PC3.

ABSTRACT: Studies on cultured cancer cells or cell lines have revealed multiple acid extrusion mechanisms and their involvement in cancer cell growth and progression. In the present study, the role of the sodium bicarbonate transporters (NBCs) in prostate cancer cell proliferation and viability was examined. qPCR revealed heterogeneous expression of five NBC isoforms in human prostate cancer cell lines LNCaP, PC3, 22RV1, C4-2, DU145, and the prostate cell line RWPE-1. In fluorescence pH measurement of LNCaP cells, which predominantly express NBCe1, Na⁺ and HCO3^--mediated acid extrusion was identified by bath ion replacement and sensitivity to the NBC inhibitor S0859. NBCe1 knockdown using siRNA oligonucleotides decreased the number of viable cells, and pharmacological inhibition with S0859 (50 µM) resulted in a similar decrease. NBCe1 knockdown and inhibition also increased cell death, but this effect was small and slow. In PC3 cells, which express all NBC isoforms, NBCe1 knockdown decreased viable cell number and increased cell death. The effects of NBCe1 knockdown were comparable to those by S0859, indicating that NBCe1 among NBCs primarily contributes to PC3 cell proliferation and viability. S0859 inhibition also decreased the formation of cell spheres in 3D cultures. Immunohistochemistry of human prostate cancer tissue microarrays revealed NBCe1 localization to the glandular epithelial cells in prostate tissue and robust expression in acinar and duct adenocarcinoma. In conclusion, our study demonstrates that NBCe1 regulates acid extrusion in prostate cancer cells and inhibiting or abolishing this transporter decreases cancer cell proliferation.

SUBMITTER: Li JM

PROVIDER: S-EPMC8144930 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Antibodies against the cardiac sodium/bicarbonate co-transporter (NBCe1) as pharmacological tools.

Project description:Background and purposeNa(+) /HCO(3) (-) co-transport (NBC) regulates intracellular pH (pH(i) ) in the heart. We have studied the electrogenic NBC isoform NBCe1 by examining the effect of functional antibodies to this protein.Experimental approachWe generated two antibodies against putative extracellular loop domains 3 (a-L3) and 4 (a-L4) of NBCe1 which recognized NBCe1 on immunoblots and immunostaining experiments. pH(i) was monitored using epi-fluorescence measurements in cat ventricular myocytes. Transport activity of total NBC and of NBCe1 in isolation were evaluated after an ammonium ion-induced acidosis (expressed as H(+) flux, J(H) , in mmol·L(-1) ?min(-1) at pH(i) 6.8) and during membrane depolarization with high extracellular potassium (potassium pulse, expressed as ?pH(i) ) respectively.Key resultsThe potassium pulse produced a pH(i) increase of 0.18 ± 0.006 (n= 5), which was reduced by the a-L3 antibody (0.016 ± 0.019). The a-L-3 also decreased J(H) by 50%. Surprisingly, during the potassium pulse, a-L4 induced a higher pH(i) increase than control,(0.25 ± 0.018) whereas the recovery of pH(i) from acidosis was faster (J(H) was almost double the control value). In perforated-patch experiments, a-L3 prolonged and a-L4 shortened action potential duration, consistent with blockade and stimulation of NBCe1-carried anionic current respectively.Conclusions and implicationsBoth antibodies recognized NBCe1, but they had opposing effects on the function of this transporter, as the a-L3 was inhibitory and the a-L4 was excitatory. These antibodies could be valuable in studies on the pathophysiology of NBCe1 in cardiac tissue, opening a path for their potential clinical use.

| S-EPMC3246661 | biostudies-literature

Mutation of Aspartate 555 of the Sodium/Bicarbonate Transporter SLC4A4/NBCe1 Induces Chloride Transport.

Project description:To understand the mechanism for ion transport through the sodium/bicarbonate transporter SLC4A4 (NBCe1), we examined amino acid residues, within transmembrane domains, that are conserved among electrogenic Na/HCO(3) transporters but are substituted with residues at the corresponding site of all electroneutral Na/HCO(3) transporters. Point mutants were constructed and expressed in Xenopus oocytes to assess function using two-electrode voltage clamp. Among the mutants, D555E (charge-conserved substitution of the aspartate at position 555 with a glutamate) produced decreasing HCO(3)(-) currents at more positive membrane voltages. Immunohistochemistry showed D555E protein expression in oocyte membranes. D555E induced Na/HCO(3)-dependent pH recovery from a CO(2)-induced acidification. Current-voltage relationships revealed that D555E produced an outwardly rectifying current in the nominally CO(2)/HCO(3)(-)-free solution that was abolished by Cl(-) removal from the bath. In the presence of CO(2)/HCO(3)(-), however, the outward current produced by D555E decreased only slightly after Cl(-) removal. Starting from a Cl(-)-free condition, D555E produced dose-dependent outward currents in response to a series of chloride additions. The D555E-mediated chloride current decreased by 70% in the presence of CO(2)/HCO(3)(-). The substitution of Asp(555) with an asparagine also produced a Cl(-) current. Anion selectivity experiments revealed that D555E was broadly permissive to other anions including NO(3)(-). Fluorescence measurements of chloride transport were done with human embryonic kidney HEK 293 cells expressing NBCe1 and D555E. A marked increase in chloride transport was detected in cells expressing D555E. We conclude that Asp(555) plays a role in HCO(3)(-) selectivity.

| S-EPMC2708891 | biostudies-literature

Electrogenic sodium bicarbonate cotransporter NBCe1 regulates pancreatic β cell function in type 2 diabetes.

Project description:Pancreatic β cell failure in type 2 diabetes mellitus (T2DM) is attributed to perturbations of the β cell's transcriptional landscape resulting in impaired glucose-stimulated insulin secretion. Recent studies identified SLC4A4 (a gene encoding an electrogenic Na+-coupled HCO3- cotransporter and intracellular pH regulator, NBCe1) as one of the misexpressed genes in β cells of patients with T2DM. Thus, in the current study, we set out to test the hypothesis that misexpression of SLC4A4/NBCe1 in T2DM β cells contributes to β cell dysfunction and impaired glucose homeostasis. To address this hypothesis, we first confirmed induction of SLC4A4/NBCe1 expression in β cells of patients with T2DM and demonstrated that its expression was associated with loss of β cell transcriptional identity, intracellular alkalinization, and β cell dysfunction. In addition, we generated a β cell-selective Slc4a4/NBCe1-KO mouse model and found that these mice were protected from diet-induced metabolic stress and β cell dysfunction. Importantly, improved glucose tolerance and enhanced β cell function in Slc4a4/NBCe1-deficient mice were due to augmented mitochondrial function and increased expression of genes regulating β cell identity and function. These results suggest that increased β cell expression of SLC4A4/NBCe1 in T2DM plays a contributory role in promotion of β cell failure and should be considered as a potential therapeutic target.

| S-EPMC8409580 | biostudies-literature

Functional Roles of Electrogenic Sodium Bicarbonate Cotransporter NBCe1 in Ocular Tissues.

Project description:Electrogenic Na(+)-HCO(3) (-) cotransporter NBCe1 is expressed in several tissues such as kidney, eye, and brain, where it may mediate distinct biological processes. In particular, NBCe1 in renal proximal tubules is essential for the regulation of systemic acid/base balance. On the other hand, NBCe1 in eye may be indispensable for the maintenance of tissue homeostasis. Consistent with this view, homozygous mutations in NBCe1 cause severe proximal renal tubular acidosis associated with ocular abnormalities such as band keratopathy, glaucoma, and cataract. The widespread expression of NBCe1 in eye suggests that the inactivation of NBCe1 per se may be responsible for the occurrence of these ocular abnormalities. In this review, we discuss about physiological and pathological roles of NBCe1 in eye.

| S-EPMC3394102 | biostudies-literature

mTOR-regulated phosphorylation of electrogenic sodium bicarbonate cotransporter 1, NBCe1, at S255-257 regulates NBCe1 functional expression in mouse cortical astrocyte

Project description:The electrogenic sodium bicarbonate cotransporter 1, NBCe1 (SLC4A4), is the major bicarbonate transporter expressed in astrocytes. It is highly sensitive for bicarbonate, and the main regulator of intracellular-, extracellular-, and synaptic pH, thereby modulating neuronal excitability. However, despite these essential functions, the molecular mechanisms underlying NBCe1-mediated astrocytic response to extracellular pH changes are mostly unknown. Using primary cortical astrocyte cultures we have investigated the effect of long-term extracellular metabolic alkalosis on regulation of NBCe1 using functional assays and phosphoproteomic analysis.

2019-07-15 | PXD014034 | Pride

Whole-Genome Sequence of the Metastatic PC3 and LNCaP Human Prostate Cancer Cell Lines.

Project description:The bone metastasis-derived PC3 and the lymph node metastasis-derived LNCaP prostate cancer cell lines are widely studied, having been described in thousands of publications over the last four decades. Here, we report short-read whole-genome sequencing (WGS) and de novo assembly of PC3 (ATCC CRL-1435) and LNCaP (clone FGC; ATCC CRL-1740) at ?70 × coverage. A known homozygous mutation in TP53 and homozygous loss of PTEN were robustly identified in the PC3 cell line, whereas the LNCaP cell line exhibited a larger number of putative inactivating somatic point and indel mutations (and in particular a loss of stop codon events). This study also provides preliminary evidence that loss of one or both copies of the tumor suppressor Capicua (CIC) contributes to primary tumor relapse and metastatic progression, potentially offering a treatment target for castration-resistant prostate cancer (CRPC). Our work provides a resource for genetic, genomic, and biological studies employing two commonly-used prostate cancer cell lines.

| S-EPMC5473753 | biostudies-literature

TGF-? signaling directly regulates transcription and functional expression of the electrogenic sodium bicarbonate cotransporter 1, NBCe1 (SLC4A4), via Smad4 in mouse astrocytes.

Project description:The electrogenic sodium bicarbonate cotransporter NBCe1 (SLC4A4) expressed in astrocytes regulates intracellular and extracellular pH. Here, we introduce transforming growth factor beta (TGF-?) as a novel regulator of NBCe1 transcription and functional expression. Using hippocampal slices and primary hippocampal and cortical astrocyte cultures, we investigated regulation of NBCe1 and elucidated the underlying signaling pathways by RT-PCR, immunoblotting, immunofluorescence, intracellular H(+ ) recording using the H(+ ) -sensitive dye 2',7'-bis-(carboxyethyl)-5-(and-6)-carboxyfluorescein, mink lung epithelial cell (MLEC) assay, and chromatin immunoprecipitation. Activation of TGF-? signaling significantly upregulated transcript, protein, and surface expression of NBCe1. These effects were TGF-? receptor-mediated and suppressed following inhibition of JNK and Smad signaling. Moreover, 4-aminopyridine (4AP)-dependent NBCe1 regulation requires TGF-?. TGF-? increased the rate and amplitude of intracellular H+ changes upon challenging NBCe1 in wild-type astrocytes but not in cortical astrocytes from Slc4a4-deficient mice. A Smad4 binding sequence was identified in the NBCe1 promoter and Smad4 binding increased after activation of TGF-? signaling. The data show for the first time that NBCe1 is a direct target of TGF-?/Smad4 signaling. Through activation of the canonical pathway TGF-? acts directly on NBCe1 by binding of Smad4 to the NBCe1 promoter and regulating its transcription, followed by increased protein expression and transport activity.

| S-EPMC5518200 | biostudies-literature

Genomic analyses of the metastasis-derived prostate cancer cell lines LNCaP, VCaP, and PC3-AR.

Project description:BackgroundThe lymph node metastasis-derived LNCaP, the bone metastasis-derived PC3 (skull), and VCaP (vertebral) cell lines are widely used as preclinical models of human prostate cancer (CaP) and have been described in more than 19,000 publications. Here, we report on short-read whole-genome sequencing and genomic analyses of LNCaP, VCaP, and PC3 cells stably transduced with WT AR (PC3-AR).MethodsLNCaP, VCaP, and PC3-AR cell lines were sequenced to an average depth of more than 30-fold using Illumina short-read sequencing. Using various computational methods, we identified and compared the single-nucleotide variants, copy-number profiles, and the structural variants observed in the three cell lines.ResultsLNCaP cells are composed of multiple subpopulations, which results in nonintegral copy number states and a high mutational load when the data is analyzed in bulk. All three cell lines contain pathogenic mutations and homozygous deletions in genes involved in DNA mismatch repair, along with deleterious mutations in cell-cycle, Wnt signaling, and other critical cellular processes. PC3-AR cells have a truncating mutation in TP53 and do not express the p53 protein. The VCaP cells contain a homozygous gain-of-function mutation in TP53 (p.R248W) that promotes cancer invasion, metastasis, and progression and has also been observed in prostate adenocarcinomas. In addition, we detect the signatures of chromothripsis of the q arms of chromosome 5 in both PC3-AR and VCaP cells, strengthening the association of TP53 inactivation with chromothripsis reported in other systems.ConclusionsOur work provides a resource for genetic, genomic, and biological studies employing these commonly-used prostate cancer cell lines.

| S-EPMC8792310 | biostudies-literature

Transport activity of the sodium bicarbonate cotransporter NBCe1 is enhanced by different isoforms of carbonic anhydrase.

Project description:Transport metabolons have been discussed between carbonic anhydrase II (CAII) and several membrane transporters. We have now studied different CA isoforms, expressed in Xenopus oocytes alone and together with the electrogenic sodium bicarbonate cotransporter 1 (NBCe1), to determine their catalytic activity and their ability to enhance NBCe1 transport activity. pH measurements in intact oocytes indicated similar activity of CAI, CAII and CAIII, while in vitro CAIII had no measurable activity and CAI only 30% of the activity of CAII. All three CA isoforms increased transport activity of NBCe1, as measured by the transport current and the rate of intracellular sodium rise in oocytes. Two CAII mutants, altered in their intramolecular proton pathway, CAII-H64A and CAII-Y7F, showed significant catalytic activity and also enhanced NBCe1 transport activity. The effect of CAI, CAII, and CAII mutants on NBCe1 activity could be reversed by blocking CA activity with ethoxyzolamide (EZA, 10 µM), while the effect of the less EZA-sensitive CAIII was not reversed. Our results indicate that different CA isoforms and mutants, even if they show little enzymatic activity in vitro, may display significant catalytic activity in intact cells, and that the ability of CA to enhance NBCe1 transport appears to depend primarily on its catalytic activity.

| S-EPMC3208603 | biostudies-literature

CryoEM structure of the human SLC4A4 sodium-coupled acid-base transporter NBCe1.

Project description:Na+-coupled acid-base transporters play essential roles in human biology. Their dysfunction has been linked to cancer, heart, and brain disease. High-resolution structures of mammalian Na+-coupled acid-base transporters are not available. The sodium-bicarbonate cotransporter NBCe1 functions in multiple organs and its mutations cause blindness, abnormal growth and blood chemistry, migraines, and impaired cognitive function. Here, we have determined the structure of the membrane domain dimer of human NBCe1 at 3.9 Å resolution by cryo electron microscopy. Our atomic model and functional mutagenesis revealed the ion accessibility pathway and the ion coordination site, the latter containing residues involved in human disease-causing mutations. We identified a small number of residues within the ion coordination site whose modification transformed NBCe1 into an anion exchanger. Our data suggest that symporters and exchangers utilize comparable transport machinery and that subtle differences in their substrate-binding regions have very significant effects on their transport mode.

| S-EPMC5834491 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data