scholarly journals Structure of Ycf1p reveals the transmembrane domain TMD0 and the regulatory region of ABCC transporters

2021 ◽  
Vol 118 (21) ◽  
pp. e2025853118
Author(s):  
Sarah C. Bickers ◽  
Samir Benlekbir ◽  
John L. Rubinstein ◽  
Voula Kanelis
Keyword(s):  
Abc Transporters ◽  
Posttranslational Modifications ◽  
Transmembrane Domain ◽  
Regulatory Region ◽  
Multidrug Resistance Proteins ◽  
Protein Activity ◽  
Resistance Proteins ◽  
Binding Domains ◽  
Abc Protein ◽  
R Region

ATP binding cassette (ABC) proteins typically function in active transport of solutes across membranes. The ABC core structure is composed of two transmembrane domains (TMD1 and TMD2) and two cytosolic nucleotide binding domains (NBD1 and NBD2). Some members of the C-subfamily of ABC (ABCC) proteins, including human multidrug resistance proteins (MRPs), also possess an N-terminal transmembrane domain (TMD0) that contains five transmembrane α-helices and is connected to the ABC core by the L0 linker. While TMD0 was resolved in SUR1, the atypical ABCC protein that is part of the hetero-octameric ATP-sensitive K+ channel, little is known about the structure of TMD0 in monomeric ABC transporters. Here, we present the structure of yeast cadmium factor 1 protein (Ycf1p), a homolog of human MRP1, determined by electron cryo-microscopy (cryo-EM). A comparison of Ycf1p, SUR1, and a structure of MRP1 that showed TMD0 at low resolution demonstrates that TMD0 can adopt different orientations relative to the ABC core, including a ∼145° rotation between Ycf1p and SUR1. The cryo-EM map also reveals that segments of the regulatory (R) region, which links NBD1 to TMD2 and was poorly resolved in earlier ABCC structures, interacts with the L0 linker, NBD1, and TMD2. These interactions, combined with fluorescence quenching experiments of isolated NBD1 with and without the R region, suggest how posttranslational modifications of the R region modulate ABC protein activity. Mapping known mutations from MRP2 and MRP6 onto the Ycf1p structure explains how mutations involving TMD0 and the R region of these proteins lead to disease.

scholarly journals Structure of Ycf1p reveals the transmembrane domain TMD0 and the regulatory R region of ABCC transporters

2021 ◽  
Author(s):  
Sarah C. Bickers ◽  
Samir Benlekbir ◽  
John L. Rubinstein ◽  
Voula Kanelis
Keyword(s):  
Transmembrane Domain ◽  
Katp Channels ◽  
K Channel ◽  
Multidrug Resistance Proteins ◽  
Protein Activity ◽  
Post Translational Modifications ◽  
Resistance Proteins ◽  
Binding Domains ◽  
Abc Protein ◽  
R Region

AbstractATP binding cassette (ABC) proteins typically function in active transport of solutes across membranes. The ABC core structure is comprised of two transmembrane domains (TMD1 and TMD2) and two cytosolic nucleotide binding domains (NBD1 and NBD2). Some members of the C-subfamily of ABC (ABCC) proteins, including human multidrug resistance proteins (MRPs), also possess an N-terminal transmembrane domain (TMD0) that contains five transmembrane α-helices and is connected to the ABC core by the L0 linker. While TMD0 was resolved in SUR1, the atypical ABCC protein that is part of the hetero-octameric ATP-sensitive K+ channel, little is known about the structure of TMD0 in monomeric ABC transporters. Here, we present the structure of yeast cadmium factor 1 protein (Ycf1p), a homologue of human MRP1, determined by electron cryomicroscopy (cryo-EM). Comparison of Ycf1p, SUR1, and a structure of MRP1 that showed TMD0 at low resolution demonstrates that TMD0 can adopt different orientations relative to the ABC core, including a 145° rotation between Ycf1p and SUR1. The cryo-EM map also reveals that segments of the regulatory (R) region, which links NBD1 to TMD2 and was poorly resolved in earlier ABCC structures, interacts with the L0 linker, NBD1, and TMD2. These interactions, combined with fluorescence quenching experiments of isolated NBD1 with and without the R region, suggests how post-translational modifications of the R region modulate ABC protein activity. Mapping known mutations from MRP2 and MRP6 onto the Ycf1p structure explains how mutations involving TMD0 and the R region of these proteins lead to disease.Statement of SignificanceThe Ycf1p structure provides an atomic model for the TMD0 domain of ABCC transporters and for two segments of the regulatory (R) region that links NBD1 to TMD2. The orientation of TMD0 in Ycf1p differs from that seen in SUR1, the regulatory ABCC protein in KATP channels, demonstrating flexibility in TMD0/ABC core contacts. The structure suggests how post-translational modifications of the R region modulate ABC protein activity and provides a mechanistic understanding of several diseases that occur due to mutation of human homologues of Ycf1p.

scholarly journals The ABCB Multidrug Resistance Proteins Do Not Contribute to Ivermectin Detoxification in the Colorado Potato Beetle, Leptinotarsa decemlineata (Say)

Insects ◽  
2020 ◽  
Vol 11 (2) ◽  
pp. 135
Author(s):  
Grant Favell ◽  
Jeremy N. McNeil ◽  
Cam Donly
Keyword(s):  
Multidrug Resistance ◽  
Colorado Potato Beetle ◽  
Leptinotarsa Decemlineata ◽  
Expression Profiles ◽  
Malpighian Tubule ◽  
Multidrug Resistance Proteins ◽  
Protein Activity ◽  
Resistance Proteins ◽  
Potato Beetle ◽  
Leptinotarsa Decemlineata Say

The Colorado potato beetle, Leptinotarsa decemlineata (Say), is a significant agricultural pest that has developed resistance to many insecticides that are used to control it. Investigating the mechanisms of insecticide detoxification in this pest is important for ensuring its continued control, since they may be contributors to such resistance. Multidrug resistance (MDR) genes that code for the ABCB transmembrane efflux transporters are one potential source of insecticide detoxification activity that have not been thoroughly examined in L. decemlineata. In this study, we annotated the ABCB genes found in the L. decemlineata genome and then characterized the expression profiles across midgut, nerve, and Malpighian tubule tissues of the three full transporters identified. To investigate if these genes are involved in defense against the macrocyclic lactone insecticide ivermectin in this insect, each gene was silenced using RNA interference or MDR protein activity was inhibited using a chemical inhibitor, verapamil, before challenging the insects with a dose of ivermectin. Survival of the insects did not significantly change due to gene silencing or protein inhibition, suggesting that MDR transporters do not significantly contribute to defense against ivermectin in L. decemlineata.

scholarly journals Role of the ABC transporter TruMDR2 in terbinafine, 4-nitroquinoline N-oxide and ethidium bromide susceptibility in Trichophyton rubrum

2006 ◽  
Vol 55 (8) ◽  
pp. 1093-1099 ◽  
Author(s):  
Ana Lúcia Fachin ◽  
Monica S. Ferreira-Nozawa ◽  
Walter Maccheroni ◽  
Nilce M. Martinez-Rossi
Keyword(s):  
Abc Transporter ◽  
Ethidium Bromide ◽  
Abc Transporters ◽  
Trichophyton Rubrum ◽  
Single Copy ◽  
Drug Susceptibility ◽  
Transmembrane Segments ◽  
Binding Domains ◽  
Abc Protein ◽  
Identical Nucleotide

A single-copy gene, designated TruMDR2, encoding an ATP-binding cassette (ABC) transporter was cloned and sequenced from the dermatophyte Trichophyton rubrum. The ORF of TruMDR2 was 4048 nt and the deduced amino acid sequence showed high homology with ABC transporters involved in drug efflux in other fungi. The encoded ABC protein predicted 12 transmembrane segments (TMSs) and two almost identical nucleotide-binding domains (NBDs) arranged in two halves in a (TMS6–NBD)2 configuration and could be classified as a member of the multidrug-resistance (MDR) class of ABC transporters. Northern blot analyses revealed an increased level of transcription of the TruMDR2 gene when mycelium was exposed to acriflavine, benomyl, ethidium bromide, ketoconazole, chloramphenicol, griseofulvin, fluconazole, imazalil, itraconazole, methotrexate, 4-nitroquinoline N-oxide (4NQO) or tioconazole. Disruption of the TruMDR2 gene rendered the mutant more sensitive to terbinafine, 4NQO and ethidium bromide than the control strain, suggesting that this transporter plays a role in modulating drug susceptibility in T. rubrum.

scholarly journals Characterization of Oligomeric Human Half-ABC Transporter ATP-binding Cassette G2

2004 ◽  
Vol 279 (19) ◽  
pp. 19781-19789 ◽  
Author(s):  
Junkang Xu ◽  
Yang Liu ◽  
Youyun Yang ◽  
Susan Bates ◽  
Jian-Ting Zhang
Keyword(s):  
Abc Transporters ◽  
Transmembrane Domain ◽  
Gel Filtration ◽  
Nucleotide Binding ◽  
Sucrose Density Gradient ◽  
Atp Binding ◽  
Cancer Resistance ◽  
Binding Domains ◽  
Resistance Protein ◽  
Atp Binding Cassette

Human ATP-binding cassette G2 (ABCG2, also known as mitoxantrone resistance protein, breast cancer-resistance protein, ABC placenta) is a member of the superfamily of ATP-binding cassette (ABC) transporters that have a wide variety of substrates. Overexpression of human ABCG2 in model cancer cell lines causes multidrug resistance by actively effluxing anticancer drugs. Unlike most of the other ABC transporters which usually have two nucleotide-binding domains and two transmembrane domains, ABCG2 consists of only one nucleotide-binding domain followed by one transmembrane domain. Thus, ABCG2 has been thought to be a half-transporter that may function as a homodimer. In this study, we characterized the oligomeric feature of human ABCG2 using non-denaturing detergent perfluoro-octanoic acid and Triton X-100 in combination with gel filtration, sucrose density gradient sedimentation, and gel electrophoresis. Unexpectedly, we found that human ABCG2 exists mainly as a tetramer, with a possibility of a higher form of oligomerization. Monomeric and dimeric ABCG2 did not appear to be the major form of the protein. Further immunoprecipitation analysis showed that the oligomeric ABCG2 did not contain any other proteins. Taken together, we conclude that human ABCG2 likely exists and functions as a homotetramer.

scholarly journals Comparison of carbohydrate ABC importers from Mycobacterium tuberculosis

BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Lilia I. De la Torre ◽  
José G. Vergara Meza ◽  
Sindy Cabarca ◽  
André G. Costa-Martins ◽  
Andrea Balan
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Abc Transporters ◽  
Binding Proteins ◽  
Substrate Binding ◽  
Regulatory Region ◽  
Nucleotide Binding ◽  
Transmembrane Domains ◽  
Pathogenic Species ◽  
Binding Domains

Abstract Background Mycobacterium tuberculosis, the etiological agent of tuberculosis, has at least four ATP-Binding Cassette (ABC) transporters dedicated to carbohydrate uptake: LpqY/SugABC, UspABC, Rv2038c-41c, and UgpAEBC. LpqY/SugABC transporter is essential for M. tuberculosis survival in vivo and potentially involved in the recycling of cell wall components. The three-dimensional structures of substrate-binding proteins (SBPs) LpqY, UspC, and UgpB were described, however, questions about how these proteins interact with the cognate transporter are still being explored. Components of these transporters, such as SBPs, show high immunogenicity and could be used for the development of diagnostic and therapeutic tools. In this work, we used a phylogenetic and structural bioinformatics approach to compare the four systems, in an attempt to predict functionally important regions. Results Through the analysis of the putative orthologs of the carbohydrate ABC importers in species of Mycobacterium genus it was shown that Rv2038c-41c and UgpAEBC systems are restricted to pathogenic species. We showed that the components of the four ABC importers are phylogenetically separated into four groups defined by structural differences in regions that modulate the functional activity or the interaction with domain partners. The regulatory region in nucleotide-binding domains, the periplasmic interface in transmembrane domains and the ligand-binding pocket of the substrate-binding proteins define their substrates and segregation in different branches. The interface between transmembrane domains and nucleotide-binding domains show conservation of residues and charge. Conclusions The presence of four ABC transporters in M. tuberculosis dedicated to uptake and transport of different carbohydrate sources, and the exclusivity of at least two of them being present only in pathogenic species of Mycobacterium genus, highlights their relevance in virulence and pathogenesis. The significant differences in the SBPs, not present in eukaryotes, and in the regulatory region of NBDs can be explored for the development of inhibitory drugs targeting the bacillus. The possible promiscuity of NBDs also contributes to a less specific and more comprehensive control approach.

Expression and localization of the multidrug resistance proteins MRP2 and MRP3 in human gallbladder epithelia

2001 ◽  
Vol 120 (5) ◽  
pp. A93-A93
Author(s):  
D ROST ◽  
J KONIG ◽  
G WEISS ◽  
E KLAR ◽  
W STREMMEL ◽  
...  
Keyword(s):  
Multidrug Resistance ◽  
Multidrug Resistance Proteins ◽  
Human Gallbladder ◽  
Resistance Proteins

scholarly journals Molecular Action of Polyphenols in Leukaemia and Their Therapeutic Potential

2021 ◽  
Vol 22 (6) ◽  
pp. 3085
Author(s):  
Hamza A. Alaswad ◽  
Amani A. Mahbub ◽  
Christine L. Le Maitre ◽  
Nicola Jordan-Mahy
Keyword(s):  
Oxidative Dna Damage ◽  
Therapeutic Potential ◽  
Multidrug Resistance Proteins ◽  
Resistance Proteins ◽  
Cancer Cell Death ◽  
Natural Agents ◽  
Activation Of Transcription ◽  
Molecular Action ◽  
Decrease Cell Proliferation ◽  
Chemotherapy Agents

Leukaemia is a malignant disease of the blood. Current treatments for leukaemia are associated with serious side-effects. Plant-derived polyphenols have been identified as potent anti-cancer agents and have been shown to work synergistically with standard chemotherapy agents in leukaemia cell lines. Polyphenols have multiple mechanisms of action and have been reported to decrease cell proliferation, arrest cell cycle and induce apoptosis via the activation of caspase (3, 8 and 9); the loss of mitochondrial membrane potential and the release of cytochrome c. Polyphenols have been shown to suppress activation of transcription factors, including NF-kB and STAT3. Furthermore, polyphenols have pro-oxidant properties, with increasing evidence that polyphenols inhibit the antioxidant activity of glutathione, causing oxidative DNA damage. Polyphenols also induce autophagy-driven cancer cell death and regulate multidrug resistance proteins, and thus may be able to reverse resistance to chemotherapy agents. This review examines the molecular mechanism of action of polyphenols and discusses their potential therapeutic targets. Here, we discuss the pharmacological properties of polyphenols, including their anti-inflammatory, antioxidant, anti-proliferative, and anti-tumour activities, and suggest that polyphenols are potent natural agents that can be useful therapeutically; and discuss why data on bioavailability, toxicity and metabolism are essential to evaluate their clinical use.

scholarly journals Spanning the gap: unraveling RSC dynamics in vivo

2021 ◽  
Author(s):  
Heinz Neumann ◽  
Bryan J. Wilkins
Keyword(s):  
Cell Cycle ◽  
Posttranslational Modifications ◽  
Transcriptional Activation ◽  
Binding Mode ◽  
Binding Modes ◽  
Binding Domains ◽  
Binding Interface ◽  
The Many ◽  
And Function

AbstractMultiple reports over the past 2 years have provided the first complete structural analyses for the essential yeast chromatin remodeler, RSC, providing elaborate molecular details for its engagement with the nucleosome. However, there still remain gaps in resolution, particularly within the many RSC subunits that harbor histone binding domains.Solving contacts at these interfaces is crucial because they are regulated by posttranslational modifications that control remodeler binding modes and function. Modifications are dynamic in nature often corresponding to transcriptional activation states and cell cycle stage, highlighting not only a need for enriched spatial resolution but also temporal understanding of remodeler engagement with the nucleosome. Our recent work sheds light on some of those gaps by exploring the binding interface between the RSC catalytic motor protein, Sth1, and the nucleosome, in the living nucleus. Using genetically encoded photo-activatable amino acids incorporated into histones of living yeast we are able to monitor the nucleosomal binding of RSC, emphasizing the regulatory roles of histone modifications in a spatiotemporal manner. We observe that RSC prefers to bind H2B SUMOylated nucleosomes in vivo and interacts with neighboring nucleosomes via H3K14ac. Additionally, we establish that RSC is constitutively bound to the nucleosome and is not ejected during mitotic chromatin compaction but alters its binding mode as it progresses through the cell cycle. Our data offer a renewed perspective on RSC mechanics under true physiological conditions.

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