The Experts below are selected from a list of 16155 Experts worldwide ranked by ideXlab platform
Omer Faruk Karatas - One of the best experts on this subject based on the ideXlab platform.
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Differential expression of ABCB1, ABCG2, and KLF4 as putative indicators for paclitaxel resistance in human epithelial type 2 cells
Molecular Biology Reports, 2021Co-Authors: Mehmet Bugrahan Duz, Omer Faruk KaratasAbstract:Laryngeal squamous cell carcinoma (LSCC) is the second most common malignancy of the head and neck region in the USA with a declining 5-year survival rate. Paclitaxel resistance of tumors including LSCC still stands as a vital cause for poor clinical outcome in patients. In the current study, our aim was to explore the expressions of ATP-binding cassette transporters and stemness associated genes in human epithelial type 2 (Hep-2) cells with paclitaxel resistance. Resistant cells were developed via treatment with increasing doses of paclitaxel to acquire four sub-lines resistant to one-, two-, four-, and eightfold concentrations of paclitaxel (1×, 2×, 4×, 8×). Then, we profiled the expressions of ten selected ABC transporters (ABCA5, ABCB1, ABCB6, ABCC1, ABCC2, ABCC3, ABCC5, ABCC10, ABCF2, and ABCG2) and four stem cell markers (SOX2, OCT4, KLF, and CXCR4) using quantitative real time polymerase chain reaction in paclitaxel resistant cells to look for a link between these markers and chemoresistance. We demonstrated that ABCB1 and ABCG2 expressions gradually elevated and reached a maximum level in Taxol 8× cells. Considering stem cell markers, KLF4 expression elevated significantly, as soon as parental cells acquired resistance to the lowest dose of paclitaxel and its expression elevated stepwise. Expression levels of other tested ATP-binding cassette transporters and stem cell markers also elevated, although at different steps of paclitaxel resistance acquisition. Our findings suggest that higher expressions of ABCB1, ABCG2, and KLF4 might be considered as putative indicators for paclitaxel resistance in LSCC patients.
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Expression profile of stem cell markers and ABC transporters in 5-fluorouracil resistant Hep-2 cells
Molecular Biology Reports, 2020Co-Authors: Omer Faruk KaratasAbstract:Resistance of laryngeal squamous cell carcinoma cells to traditional therapeutic regimens still remains to be a major reason for therapeutic failure in patients. In this study, we aimed at investigating the expression profiles of ATP-binding cassette (ABC) transporters and stem cell markers in 5-fluorouracil (5-FU) resistant laryngeal Hep-2 cells. We treated parental Hep-2 cells, with stepwise increased doses of 5-FU for almost 1 year to develop 5-FU resistant sub-lines with resistance against varying levels of 5-FU concentrations (4 sub-lines resistant to 1, 2, 4, and eightfold of 5-FU). Then, we measured the expression levels of 10 genes from ABC transporters family and 4 stem cell associated markers using quantitative reverse transcription polymerase chain reaction (qRT-PCR) to find out a potential relationship between these markers and chemoresistance. We found that stemness-associated markers had elevated expressions from the beginning of 5-FU resistance acquisition. Their expressions elevated stepwise while parental Hep-2 cells got resistance to higher doses of 5-FU. Expressions of tested ABC transporters (ABCA5, ABCB1, ABCB6, ABCC1, ABCC2, ABCC3, ABCC5, ABCC10 and ABCF2, and ABCG2) were also deregulated in 5-FU resistant Hep-2 cells. Although their expressions remained unaltered at the beginning of acquisition of resistance, expressions of ABC transporters except from ABCB6 increased significantly when cells became resistant to higher doses of 5-FU. Our results suggest that enrichment of cells with stemness characteristics and upregulation of ABC transporters might be amongst the crucial contributors of chemoresistance in laryngeal cancer cells.
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Expression profile of stem cell markers and ABC transporters in 5-fluorouracil resistant Hep-2 cells
Molecular Biology Reports, 2020Co-Authors: Omer Faruk KaratasAbstract:Resistance of laryngeal squamous cell carcinoma cells to traditional therapeutic regimens still remains to be a major reason for therapeutic failure in patients. In this study, we aimed at investigating the expression profiles of ATP-binding cassette (ABC) transporters and stem cell markers in 5-fluorouracil (5-FU) resistant laryngeal Hep-2 cells. We treated parental Hep-2 cells, with stepwise increased doses of 5-FU for almost 1 year to develop 5-FU resistant sub-lines with resistance against varying levels of 5-FU concentrations (4 sub-lines resistant to 1, 2, 4, and eightfold of 5-FU). Then, we measured the expression levels of 10 genes from ABC transporters family and 4 stem cell associated markers using quantitative reverse transcription polymerase chain reaction (qRT-PCR) to find out a potential relationship between these markers and chemoresistance. We found that stemness-associated markers had elevated expressions from the beginning of 5-FU resistance acquisition. Their expressions elevated stepwise while parental Hep-2 cells got resistance to higher doses of 5-FU. Expressions of tested ABC transporters (ABCA5, ABCB1, ABCB6, ABCC1, ABCC2, ABCC3, ABCC5, ABCC10 and ABCF2, and ABCG2) were also deregulated in 5-FU resistant Hep-2 cells. Although their expressions remained unaltered at the beginning of acquisition of resistance, expressions of ABC transporters except from ABCB6 increased significantly when cells became resistant to higher doses of 5-FU. Our results suggest that enrichment of cells with stemness characteristics and upregulation of ABC transporters might be amongst the crucial contributors of chemoresistance in laryngeal cancer cells.
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The role of ATP‐binding cassette transporter genes in the progression of prostate cancer
The Prostate, 2015Co-Authors: Omer Faruk Karatas, Mehmet Bugrahan Duz, Esra Guzel, Michael Ittmann, Mustafa OzenAbstract:BACKGROUND Prostate cancer (PCa) is the most commonly diagnosed neoplasm and the second leading cause of cancer-related death among men in developed countries. There is no clear evidence showing the success of current screening tests in reducing mortality of PCa. In this study, we aimed to profile expressions of nine ABC transporters, ABCA5, ABCB1, ABCB6, ABCC1, ABCC2, ABCC3, ABCC5, ABCC10, and ABCF2, in recurrent, non-recurrent PCa and normal prostate tissues. METHODS A total of 77 (39 recurrent, 38 non-recurrent) radical prostatectomy and 20 normal prostate samples, obtained from Baylor College of Medicine Prostate Cancer program, were included into the study and divided into two independent groups as test and validation sample sets. Differential expression of selected ABC transporters was assessed using quantitative real-time PCR (qRT-PCR). Pearson's correlation test, receiver operating characteristics (ROC) analysis and Kaplan–Meier test were used for statistical analysis. RESULTS QRT-PCR results demonstrated the elevated expression of ABCA5, ABCB1, ABCB6, ABCC1, and ABCC2 as well as reduced expression of ABCC3 in PCa samples compared to normal prostate tissues. In addition, we found deregulation of ABCB1, ABCB6, ABCC3, and ABCC10 in recurrent PCa samples and validated differential expression of ABCB6, ABCC3, and ABCC10 in recurrent PCa compared to non-recurrent PCa. Pearson's correlation, ROC and Kaplan–Meier analysis revealed the power of these three ABC transporters for estimating prognosis of PCa. CONCLUSIONS We demonstrated differential expression of ABC transporters both in tumor versus normal and recurrent versus non-recurrent comparisons. Our data suggest ABCB6, ABCC3, and ABCC10 as valuable predictors of PCa progression. Prostate © 2015 Wiley Periodicals, Inc.
Olena K. Vatamaniuk - One of the best experts on this subject based on the ideXlab platform.
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n terminal extension and c terminal domains are required for ABCB6 hmt 1 protein interactions function in cadmium detoxification and localization to the endosomal recycling system in caenorhabditis elegans
Frontiers in Physiology, 2018Co-Authors: Sungjin Kim, Anuj K. Sharma, Olena K. VatamaniukAbstract:The chronic exposure of humans to toxic metals such as cadmium from food and air causes dysfunction of vital organs, neurodegenerative conditions, and cancer. In this regard, members of the ABCB sub-family of the ATP-binding cassette (ABC) transporter superfamily, ABCB6/HMT-1, are acutely required for the detoxification of heavy metals and are present in genomes of many organisms including the nematode worm, Caenorhabditis elegans and humans. We showed previously that C. elegans ABCB6/HMT-1 detoxifies cadmium, copper, and arsenic, and is expressed in liver-like cells, the coelomocytes, head neurons and intestinal cells, which are the cell types that are affected by heavy metal poisoning in humans. The subcellular localization of ABCB6/HMT-1 proteins is unclear. ABCB6/HMT-1 proteins have a distinguishing topology: in addition to one transmembrane domain and one nucleotide-binding domain, they possess a hydrophobic N-terminal extension (NTE) domain encompassing five to six transmembrane spans. The role of the NTE domain in the function of ABCB6/HMT-1 in the native organism remains to be investigated. We used a versatile, multicellular model system, C. elegans, to establish the subcellular localization of ABCB6/HMT-1 and refine its structure-function studies in the native organism. We show that ABCB6/HMT-1 localizes mainly to the apical recycling endosomes and, in part, to early and late endosomes of intestinal cells. We also show that ABCB6/HMT-1 lacking the NTE domain is mistargeted to the plasma membrane and is unable to confer cadmium resistance. Although the NTE domain is essential for ABCB6/HMT-1 interaction with itself, the absence of NTE does not fully prevent this interaction. As a result, ABCB6/HMT-1 lacking the NTE domain, and expressed in wild-type worms or co-expressed with the full-length polypeptide, inactivates and mistargets the full-length ABCB6/HMT-1. We also show that the 43 amino acid residue stretch at the COOH-terminus is required for the ABCB6/HMT-1 interaction with itself and cadmium detoxification function. These results suggest that both NTE and COOH-terminus must be present to allow the protein to interact with itself and confer cadmium resistance. Considering that ABCB6/HMT-1 proteins are highly conserved, this study advances our understanding of how these proteins function in cadmium resistance in different species. Furthermore, these studies uncover the role of the endosomal-recycling system in cadmium detoxification.
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Data_Sheet_1_N-Terminal Extension and C-Terminal Domains Are Required for ABCB6/HMT-1 Protein Interactions, Function in Cadmium Detoxification, and Localization to the Endosomal-Recycling System in Caenorhabditis elegans.PDF
2018Co-Authors: Sungjin Kim, Anuj K. Sharma, Olena K. VatamaniukAbstract:The chronic exposure of humans to toxic metals such as cadmium from food and air causes dysfunction of vital organs, neurodegenerative conditions, and cancer. In this regard, members of the ABCB sub-family of the ATP-binding cassette (ABC) transporter superfamily, ABCB6/HMT-1, are acutely required for the detoxification of heavy metals and are present in genomes of many organisms including the nematode worm, Caenorhabditis elegans and humans. We showed previously that C. elegans ABCB6/HMT-1 detoxifies cadmium, copper, and arsenic, and is expressed in liver-like cells, the coelomocytes, head neurons and intestinal cells, which are the cell types that are affected by heavy metal poisoning in humans. The subcellular localization of ABCB6/HMT-1 proteins is unclear. ABCB6/HMT-1 proteins have a distinguishing topology: in addition to one transmembrane domain and one nucleotide-binding domain, they possess a hydrophobic N-terminal extension (NTE) domain encompassing five to six transmembrane spans. The role of the NTE domain in the function of ABCB6/HMT-1 in the native organism remains to be investigated. We used a versatile, multicellular model system, C. elegans, to establish the subcellular localization of ABCB6/HMT-1 and refine its structure-function studies in the native organism. We show that ABCB6/HMT-1 localizes mainly to the apical recycling endosomes and, in part, to early and late endosomes of intestinal cells. We also show that ABCB6/HMT-1 lacking the NTE domain is mistargeted to the plasma membrane and is unable to confer cadmium resistance. Although the NTE domain is essential for ABCB6/HMT-1 interaction with itself, the absence of NTE does not fully prevent this interaction. As a result, ABCB6/HMT-1 lacking the NTE domain, and expressed in wild-type worms or co-expressed with the full-length polypeptide, inactivates and mistargets the full-length ABCB6/HMT-1. We also show that the 43 amino acid residue stretch at the COOH-terminus is required for the ABCB6/HMT-1 interaction with itself and cadmium detoxification function. These results suggest that both NTE and COOH-terminus must be present to allow the protein to interact with itself and confer cadmium resistance. Considering that ABCB6/HMT-1 proteins are highly conserved, this study advances our understanding of how these proteins function in cadmium resistance in different species. Furthermore, these studies uncover the role of the endosomal-recycling system in cadmium detoxification.
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N-Terminal Extension and C-Terminal Domains Are Required for ABCB6/HMT-1 Protein Interactions, Function in Cadmium Detoxification, and Localization to the Endosomal-Recycling System in Caenorhabditis elegans
Frontiers Media S.A., 2018Co-Authors: Sungjin Kim, Anuj K. Sharma, Olena K. VatamaniukAbstract:The chronic exposure of humans to toxic metals such as cadmium from food and air causes dysfunction of vital organs, neurodegenerative conditions, and cancer. In this regard, members of the ABCB sub-family of the ATP-binding cassette (ABC) transporter superfamily, ABCB6/HMT-1, are acutely required for the detoxification of heavy metals and are present in genomes of many organisms including the nematode worm, Caenorhabditis elegans and humans. We showed previously that C. elegans ABCB6/HMT-1 detoxifies cadmium, copper, and arsenic, and is expressed in liver-like cells, the coelomocytes, head neurons and intestinal cells, which are the cell types that are affected by heavy metal poisoning in humans. The subcellular localization of ABCB6/HMT-1 proteins is unclear. ABCB6/HMT-1 proteins have a distinguishing topology: in addition to one transmembrane domain and one nucleotide-binding domain, they possess a hydrophobic N-terminal extension (NTE) domain encompassing five to six transmembrane spans. The role of the NTE domain in the function of ABCB6/HMT-1 in the native organism remains to be investigated. We used a versatile, multicellular model system, C. elegans, to establish the subcellular localization of ABCB6/HMT-1 and refine its structure-function studies in the native organism. We show that ABCB6/HMT-1 localizes mainly to the apical recycling endosomes and, in part, to early and late endosomes of intestinal cells. We also show that ABCB6/HMT-1 lacking the NTE domain is mistargeted to the plasma membrane and is unable to confer cadmium resistance. Although the NTE domain is essential for ABCB6/HMT-1 interaction with itself, the absence of NTE does not fully prevent this interaction. As a result, ABCB6/HMT-1 lacking the NTE domain, and expressed in wild-type worms or co-expressed with the full-length polypeptide, inactivates and mistargets the full-length ABCB6/HMT-1. We also show that the 43 amino acid residue stretch at the COOH-terminus is required for the ABCB6/HMT-1 interaction with itself and cadmium detoxification function. These results suggest that both NTE and COOH-terminus must be present to allow the protein to interact with itself and confer cadmium resistance. Considering that ABCB6/HMT-1 proteins are highly conserved, this study advances our understanding of how these proteins function in cadmium resistance in different species. Furthermore, these studies uncover the role of the endosomal-recycling system in cadmium detoxification
Partha Krishnamurthy - One of the best experts on this subject based on the ideXlab platform.
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Efficient purification and reconstitution of ATP binding cassette transporter B6 (ABCB6) for functional and structural studies.
The Journal of biological chemistry, 2013Co-Authors: Hemantkumar Chavan, Mohiuddin Md. Taimur Khan, George P. Tegos, Partha KrishnamurthyAbstract:The mitochondrial ATP binding cassette transporter ABCB6 has been associated with a broad range of physiological functions, including growth and development, therapy-related drug resistance, and the new blood group system Langereis. ABCB6 has been proposed to regulate heme synthesis by shuttling coproporphyrinogen III from the cytoplasm into the mitochondria. However, direct functional information of the transport complex is not known. To understand the role of ABCB6 in mitochondrial transport, we developed an in vitro system with pure and active protein. ABCB6 overexpressed in HEK293 cells was solubilized from mitochondrial membranes and purified to homogeneity. Purified ABCB6 showed a high binding affinity for MgATP (Kd = 0.18 μM) and an ATPase activity with a Km of 0.99 mM. Reconstitution of ABCB6 into liposomes allowed biochemical characterization of the ATPase including (i) substrate-stimulated ATPase activity, (ii) transport kinetics of its proposed endogenous substrate coproporphyrinogen III, and (iii) transport kinetics of substrates identified using a high throughput screening assay. Mutagenesis of the conserved lysine to alanine (K629A) in the Walker A motif abolished ATP hydrolysis and substrate transport. These results suggest a direct interaction between mitochondrial ABCB6 and its transport substrates that is critical for the activity of the transporter. Furthermore, the simple immunoaffinity purification of ABCB6 to near homogeneity and efficient reconstitution of ABCB6 into liposomes might provide the basis for future studies on the structure/function of ABCB6.
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Efficient Purification and Reconstitution of ATP Binding Cassette Transporter B6 (ABCB6) for Functional and
2013Co-Authors: Hemantkumar Chavan, Partha Krishnamurthy, George P. Tegos, Taimur Khan, Fromthe ‡ DepartmentofpharmacologyAbstract:The mitochondrial ATP binding cassette transporter ABCB6 has been associated with a broad range of physiological functions, including growth and development, therapy-related drug resistance, and the new blood group system Langereis. ABCB6 has been proposed to regulate heme synthesis by shuttling coproporphyrinogen III from the cytoplasm into the mitochondria. However, direct functional information of the transport complex is not known. To understand the role of ABCB6 in mitochondrial transport, we developed an in vitro system with pure and active protein. ABCB6 overexpressed in HEK293 cells was solubilized from mitochondrial membranes and purified to homogeneity. Purified ABCB6 showed a high binding affinity for MgATP (Kd 0.18 M) and an ATPase activity with a Km of 0.99 mM. Reconstitution of ABCB6 into liposomes allowed biochemical characterization of the ATPase including (i) substratestimulated ATPase activity, (ii) transport kinetics of its proposed endogenous substrate coproporphyrinogen III, and (iii) transport kinetics of substrates identified using a high throughput screening assay. Mutagenesis of the conserved lysine to alanine (K629A) in the Walker A motif abolished ATP hydrolysis and substrate transport. These results suggest a direct interaction between mitochondrial ABCB6 and its transport substrates
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polycyclic aromatic hydrocarbons pahs mediate transcriptional activation of the atp binding cassette transporter ABCB6 gene via the aryl hydrocarbon receptor ahr
Journal of Biological Chemistry, 2012Co-Authors: Hemantkumar Chavan, Partha KrishnamurthyAbstract:Abstract Liver is endowed with a mechanism to induce hepatic cytochromes P450 (CYP450s) in response to therapeutic drugs and environmental contaminants, leading to increased detoxification and elimination of the xenobiotics. Each CYP450 is composed of an apoprotein moiety and a heme prosthetic group, which is required for CYP450 activity. Thus, under conditions of CYP450 induction, there is a coordinate increase in heme biosynthesis to compensate for the increased expression of CYP450s. ABCB6, a mitochondrial ATP binding cassette transporter, which regulates coproporphyrinogen transport from the cytoplasm into the mitochondria to complete heme biosynthesis, represents a previously unrecognized rate-limiting step in heme biosynthesis. However, it is not known if exposure to drugs and environmental contaminants induces ABCB6 expression, to assure an adequate and apparently coordinated supply of heme for the generation of functional cytochrome holoprotein. In the present study, we demonstrate that polycyclic aromatic hydrocarbons (PAHs), the widely distributed environmental toxicants shown to induce porphyrin accumulation causing hepatic porphyria, up-regulate ABCB6 expression in both mice and humans. Using siRNA technology and ABCB6 knock-out mice, we demonstrate that PAH-mediated increase in hepatic porphyrins is compromised in the absence of ABCB6. Moreover, in vivo studies in aryl hydrocarbon receptor (AhR) knock-out mice demonstrate that PAH induction of ABCB6 is mediated by AhR. Promoter activation studies combined with electrophoretic mobility shift assay and chromatin immunoprecipitation assay demonstrate direct interactions between the AhR binding sites in the ABCB6 promoter and the AhR receptor, implicating drug activation mechanisms for ABCB6 similar to those found in inducible cytochrome P450s. These studies are the first to describe direct transcriptional activation of both mouse and human ABCB6 by xenobiotics.
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Abstract 3044: A new paradigm for ABCB6: Role in growth and proliferation
Molecular and Cellular Biology, 2012Co-Authors: Kishore Polireddy, Hemant Chavan, Partha KrishnamurthyAbstract:ABCB6 is a mitochondrial ATP binding cassette (ABC) transporter whose expression is upregulated in a broad spectrum of primary human cancers, including hepatocellular carcinoma (HCC). We recently demonstrated that in hepatoma cells ABCB6 expression promotes growth and proliferation. In contrast, loss of ABCB6 expression in these hepatomas precipitates a delay in progression through G2/M phase of the cell cycle. As histone H3 is specifically phopshorylated during mitosis, we took advantage of the H3 phosphorylated antibody to separate G2 phase of the cell cycle from the M-phase. Using this approach, we demonstrate that loss of ABCB6 expression leads to a delay in M phase and not the G2 phase of the cell cycle. However, it is not clear how a mitochondrial transporter, originally characterized as a transporter of porphyrins, regulates cell growth. We have proposed two broad hypotheses to evaluate the mechanism by which ABCB6 regulates growth and proliferation; 1) ABCB6 functionally interacts with one or more of the cell cycle regulatory components, thus precipitating M phase delay; 2) ABCB6s’ location and function as a mitochondrial membrane transporter alters mitochondrial dynamics leading to some level of control over cellular proliferation. To test our proposed first hypothesis we have begun a systematic evaluation of the expression and function of cell cycle regulatory components in response to ABCB6 expression. Results from these studies demonstrate that ABCB6 expression alters the expression of cyclin-dependent kinase inhibitors p16, p21 and p27. In addition, yeast two-hybrid screening and protein interaction studies reveal that ABCB6 interacts with p21 and a novel tumor susceptibility protein-101 (TSG-101). TSG 101 co-localizes with microtubule organizing centers and mitotic spindles during mitosis and its deficiency is associated with abnormalities in microtubules, genetic instability and carcinogenesis. In this report, we provide evidence of the physical and functional interactions between ABCB6 and cell cycle regulatory components and explore the significance of these interactions to growth and proliferation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3044. doi:1538-7445.AM2012-3044
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Functional significance of the ATP-binding cassette transporter B6 in hepatocellular carcinoma
Molecular oncology, 2011Co-Authors: Kishore Polireddy, Hemantkumar Chavan, Bashar Abdulkarim, Partha KrishnamurthyAbstract:ABCB6 is a mitochondrial transporter that regulates porphyrin biosynthesis. ABCB6 expression is upregulated in hepatocellular carcinoma (HCC) but the significance of this upregulation to HCC is not known. In the present study, we investigated: 1) ABCB6 expression in 18 resected human hepatocellular carcinoma (HCC) tissues and 3 human hepatoma cell lines; 2) pattern of ABCB6 expression during liver disease progression; and 3) functional significance of ABCB6 expression to HCC using the hepatoma cell line Huh7. ABCB6 expression was determined by real-time quantitative reverse transcription-polymerase chain reaction and western blotting. ABCB6 expression was upregulated in all the HCC specimens and the three-hepatoma cell lines. Increased ABCB6 expression correlated with liver disease progression with the pattern of expression being HCC > cirrhosis > steatosis. Small hairpin RNA (shRNA)-mediated knockdown of ABCB6 in Huh7 cells lead to decreased cellular proliferation and colony formation. Attenuation of ABCB6 expression did not affect Huh7 apoptosis but lead to a delay in G2/M phase of the cell cycle. In contrast, ABCB6 overexpression resulted in increased growth and proliferation of Huh7 cells. Since ABCB6 expression is induced in multiple tumor types we explored the role of ABCB6 in other cancer cells. ShRNA mediated knockdown of ABCB6 in HEK293 and K562 cells reduced cellular proliferation leading to a delay in G2/M phase, while ABCB6 overexpression promoted cell growth and proliferation. Collectively, these findings, obtained by loss of function and gain of function analysis, suggest that ABCB6 plays a role in cell growth and proliferation by targeting the cell cycle.
L. Ramos - One of the best experts on this subject based on the ideXlab platform.
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corrigendum to isolation and sex steroid effects on the expression of the atp binding cassette transporter ABCB6 in harderian glands of hamster mesocricetus auratus comparative biochemistry and physiology part a 232 2019 40 46
Comparative Biochemistry and Physiology A-molecular & Integrative Physiology, 2019Co-Authors: L. Mares, F. Vilchis, B. Chávez, L. RamosAbstract:Abstract A TP- B inding C assette, subfamily B , member 6 (ABCB6) is a transporter that is upregulated by elevated intracellular porphyrin concentrations. In the Harderian gland (HG), the synthesis of porphyrins appears to be under the influence of gonadal steroids and to exhibit a dimorphic pattern. To explore whether ABCB6 is also influenced by sex steroids, we isolated its specific cDNA sequence and investigated its mRNA levels in the HGs of hamsters. ABCB6's cDNA sequence presents an open reading frame (ORF) of 2529 bp that encodes a predicted 842-amino acid (aa) protein with a molecular weight of 93 kDa. Multiple sequence alignments showed that ABCB6's aa sequence is highly conserved and shares the highest homology (93%) with mouse ABCB6. RT-qPCR analysis indicated that ABCB6 is expressed in all the tissues examined, exhibiting high expression levels in the liver, adrenal glands, and testis. The mRNA concentrations of ABCB6 in HGs were very similar between males and in females; similarly, gonadectomy and treatment with sex steroids appear to scarcely affect ABCB6 mRNA levels. The intraglandular content of ABCB6 mRNA showed discrete, though non-significant, variations through the estrous cycle. The results provide evidence that gonadal steroids have a minimal physiological role on the regulation of ABCB6 expression and might indicate that this transporter has a small effect on porphyrin trafficking in the HGs of hamsters. The authors would like to apologise for any inconvenience caused.
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Corrigendum to “isolation and sex steroid effects on the expression of the ATP-binding cassette transporter ABCB6 in Harderian glands of hamster (Mesocricetus auratus)” [Comparative Biochemistry and Physiology, Part A 232 (2019) 40–46]
Comparative biochemistry and physiology. Part A Molecular & integrative physiology, 2019Co-Authors: L. Mares, F. Vilchis, B. Chávez, L. RamosAbstract:Abstract A TP- B inding C assette, subfamily B , member 6 (ABCB6) is a transporter that is upregulated by elevated intracellular porphyrin concentrations. In the Harderian gland (HG), the synthesis of porphyrins appears to be under the influence of gonadal steroids and to exhibit a dimorphic pattern. To explore whether ABCB6 is also influenced by sex steroids, we isolated its specific cDNA sequence and investigated its mRNA levels in the HGs of hamsters. ABCB6's cDNA sequence presents an open reading frame (ORF) of 2529 bp that encodes a predicted 842-amino acid (aa) protein with a molecular weight of 93 kDa. Multiple sequence alignments showed that ABCB6's aa sequence is highly conserved and shares the highest homology (93%) with mouse ABCB6. RT-qPCR analysis indicated that ABCB6 is expressed in all the tissues examined, exhibiting high expression levels in the liver, adrenal glands, and testis. The mRNA concentrations of ABCB6 in HGs were very similar between males and in females; similarly, gonadectomy and treatment with sex steroids appear to scarcely affect ABCB6 mRNA levels. The intraglandular content of ABCB6 mRNA showed discrete, though non-significant, variations through the estrous cycle. The results provide evidence that gonadal steroids have a minimal physiological role on the regulation of ABCB6 expression and might indicate that this transporter has a small effect on porphyrin trafficking in the HGs of hamsters. The authors would like to apologise for any inconvenience caused.
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Isolation and sex steroid effects on the expression of the ATP-binding cassette transporter ABCB6 in Harderian glands of hamster (Mesocricetus auratus).
Comparative biochemistry and physiology. Part A Molecular & integrative physiology, 2019Co-Authors: L. Mares, F. Vilchis, B. Chávez, L. RamosAbstract:ATP-Binding Cassette, subfamily B, member 6 (ABCB6) is a transporter that is upregulated by elevated intracellular porphyrin concentrations. In the Harderian gland (HG), the synthesis of porphyrins appears to be under the influence of gonadal steroids and to exhibit a dimorphic pattern. To explore whether ABCB6 is also influenced by sex steroids, we isolated its specific cDNA sequence and investigated its mRNA levels in the HGs of hamsters. ABCB6's cDNA sequence presents an open reading frame (ORF) of 2529 bp that encodes a predicted 842-amino acid (aa) protein with a molecular weight of 93 kDa. Multiple sequence alignments showed that ABCB6's aa sequence is highly conserved and shares the highest homology (93%) with mouse ABCB6. RT-qPCR analysis indicated that ABCB6 is expressed in all the tissues examined, exhibiting high expression levels in the liver, adrenal glands, and testis. The mRNA concentrations of ABCB6 in HGs were very similar between males and in females; similarly, gonadectomy and treatment with sex steroids appear to scarcely affect ABCB6 mRNA levels. The intraglandular content of ABCB6 mRNA showed discrete, though non-significant, variations through the estrous cycle. The results provide evidence that gonadal steroids have a minimal physiological role on the regulation of ABCB6 expression and might indicate that this transporter has a small effect on porphyrin trafficking in the HGs of hamsters.
Nora Kucsma - One of the best experts on this subject based on the ideXlab platform.
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The human ABCB6 protein is the functional homologue of HMT-1 proteins mediating cadmium detoxification
Cellular and Molecular Life Sciences, 2019Co-Authors: Zsófia Rakvács, Nora Kucsma, Melinda Gera, Barbara Igriczi, Katalin Kiss, János Barna, Dániel Kovács, Tibor Vellai, László Bencs, Johannes M. ReiseckerAbstract:ABCB6 belongs to the family of ATP-binding cassette (ABC) transporters, which transport various molecules across extra- and intra-cellular membranes, bearing significant impact on human disease and pharmacology. Although mutations in the ABCB6 gene have been linked to a variety of pathophysiological conditions ranging from transfusion incompatibility to pigmentation defects, its precise cellular localization and function is not understood. In particular, the intracellular localization of ABCB6 has been a matter of debate, with conflicting reports suggesting mitochondrial or endolysosomal expression. ABCB6 shows significant sequence identity to HMT-1 (heavy metal tolerance factor 1) proteins, whose evolutionarily conserved role is to confer tolerance to heavy metals through the intracellular sequestration of metal complexes. Here, we show that the cadmium-sensitive phenotype of Schizosaccharomyces pombe and Caenorhabditis elegans strains defective for HMT-1 is rescued by the human ABCB6 protein. Overexpression of ABCB6 conferred tolerance to cadmium and As(III) (As_2O_3), but not to As(V) (Na_2HAsO_4), Sb(V), Hg(II), or Zn(II). Inactivating mutations of ABCB6 abolished vacuolar sequestration of cadmium, effectively suppressing the cadmium tolerance phenotype. Modulation of ABCB6 expression levels in human glioblastoma cells resulted in a concomitant change in cadmium sensitivity. Our findings reveal ABCB6 as a functional homologue of the HMT-1 proteins, linking endolysosomal ABCB6 to the highly conserved mechanism of intracellular cadmium detoxification.
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Role of the N-terminal transmembrane domain in the endo-lysosomal targeting and function of the human ABCB6 protein
The Biochemical journal, 2015Co-Authors: Katalin E Kiss, Anna Brozik, Nora Kucsma, Gábor E. Tusnády, Ptissam Bergam, Guillaume Van Niel, Gergely SzakacsAbstract:ATP-binding cassette, subfamily B (ABCB) 6 is a homodimeric ATP-binding cassette (ABC) transporter present in the plasma membrane and in the intracellular organelles. The intracellular localization of ABCB6 has been a matter of debate, as it has been suggested to reside in the mitochondria and the endo-lysosomal system. Using a variety of imaging modalities, including confocal microscopy and EM, we confirm the endo-lysosomal localization of ABCB6 and show that the protein is internalized from the plasma membrane through endocytosis, to be distributed to multivesicular bodies and lysosomes. In addition to the canonical nucleotide-binding domain (NBD) and transmembrane domain (TMD), ABCB6 contains a unique N-terminal TMD (TMD0), which does not show sequence homology to known proteins. We investigated the functional role of these domains through the molecular dissection of ABCB6. We find that the folding, dimerization, membrane insertion and ATP binding/hydrolysis of the core–ABCB6 complex devoid of TMD0 are preserved. However, in contrast with the full-length transporter, the core–ABCB6 construct is retained at the plasma membrane and does not appear in Rab5-positive endosomes. TMD0 is directly targeted to the lysosomes, without passage to the plasma membrane. Collectively, our results reveal that TMD0 represents an independently folding unit, which is dispensable for catalysis, but has a crucial role in the lysosomal targeting of ABCB6.
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Screening the Expression of ABCB6 in Erythrocytes Reveals an Unexpectedly High Frequency of Lan Mutations in Healthy Individuals
PloS one, 2014Co-Authors: Magdalena Koszarska, Katalin E Kiss, Nora Kucsma, Melinda Gera, György Várady, Géza Antalffy, Hajnalka Andrikovics, Attila Tordai, Maciej Studzian, Dominik StrapagielAbstract:Lan is a high-incidence blood group antigen expressed in more than 99.9% of the population. Identification of the human ABC transporter ABCB6 as the molecular basis of Lan has opened the way for studies assessing the relation of ABCB6 function and expression to health and disease. To date, 34 ABCB6 sequence variants have been described in association with reduced ABCB6 expression based on the genotyping of stored blood showing weak or no reactivity with anti-Lan antibodies. In the present study we examined the red blood cell (RBC) surface expression of ABCB6 by quantitative flow cytometry in a cohort of 47 healthy individuals. Sequencing of the entire coding region of the ABCB6 gene in low RBC ABCB6 expressors identified a new allele (IVS9+1G>A, affecting a putative splice site at the boundary of exon 9) and two nonsynonymous SNPs listed in the SNP database (R192Q (rs150221689) and G588 S (rs145526996)). The R192Q mutation showed co-segregation with reduced RBC ABCB6 expression in a family, and we found the G588 S mutation in a compound heterozygous individual with undetectable ABCB6 expression, suggesting that both mutations result in weak or no expression of ABCB6 on RBCs. Analysis of the intracellular expression pattern in HeLa cells by confocal microscopy indicated that these mutations do not compromise overall expression or the endolysosomal localization of ABCB6. Genotyping of two large cohorts, containing 235 and 1039 unrelated volunteers, confirmed the high allele frequency of Lan-mutations. Our results suggest that genetic variants linked to lower or absent cell surface expression of ABCB6/Langereis may be more common than previously thought.
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shifting the paradigm the putative mitochondrial protein ABCB6 resides in the lysosomes of cells and in the plasma membrane of erythrocytes
PLOS ONE, 2012Co-Authors: Katalin E Kiss, Anna Brozik, Nora Kucsma, Alexandra Toth, Melinda Gera, Laurence Berry, Alice Vallentin, Henri Vial, Michel Vidal, Gergely SzakacsAbstract:ABCB6, a member of the adenosine triphosphate–binding cassette (ABC) transporter family, has been proposed to be responsible for the mitochondrial uptake of porphyrins. Here we show that ABCB6 is a glycoprotein present in the membrane of mature erythrocytes and in exosomes released from reticulocytes during the final steps of erythroid maturation. Consistent with its presence in exosomes, endogenous ABCB6 is localized to the endo/lysosomal compartment, and is absent from the mitochondria of cells. Knock-down studies demonstrate that ABCB6 function is not required for de novo heme biosynthesis in differentiating K562 cells, excluding this ABC transporter as a key regulator of porphyrin synthesis. We confirm the mitochondrial localization of ABCB7, ABCB8 and ABCB10, suggesting that only three ABC transporters should be classified as mitochondrial proteins. Taken together, our results challenge the current paradigm linking the expression and function of ABCB6 to mitochondria.
