ABC transporters and the hallmarks of cancer:roles in cancer aggressiveness beyond multidrug resistance

The ATP-binding cassette transporters(ABC transporters)have been intensely studied over the past 50 years for their involvement in the multidrug resistance(MDR)phenotype,especially in cancer.They are frequently overexpressed in both naive and post-treatment tumors,and hinder effective chemotherapy by reducing drug accumulation in cancer cells.In the last decade however,several studies have established that ABC transporters have additional,fundamental roles in tumor biology;there is strong evidence that these proteins are involved in transporting tumor-enhancing molecules and/or in protein-protein interactions that impact cancer aggressiveness,progression,and patient prognosis.This review highlights these studies in relation to some well-described cancer hallmarks,in an effort to re-emphasize the need for further investigation into the physiological functions of ABC transporters that are critical for tum or development.Unraveling these new roles offers an opportunity to define new strategies and targets for therapy,which would include endogenous substrates or signaling pathways that regulate these proteins.

Metabolomic profiles in a green alga (Raphidocelis subcapitata) following erythromycin treatment: ABC transporters and energy metabolism

A recent study showed that erythromycin(ERY)exposure caused hormesis in a model alga(Raphidocelis subcapitata)where the growth was promoted at an environmentally realistic concentration(4μg/L)but inhibited at two higher concentrations(80 and 120μg/L),associated with opposite actions of certain signaling pathways(e.g.,xenobiotic metabolism,DNA replication).However,these transcriptional alterations remain to be investigated and verified at the metabolomic level.This study uncovered metabolomic profiles and detailed toxic mechanisms of ERY in R.subcapitata using untargetedmetabolomics.Themetabolomic analysis showed that metabolomic pathways including ABC transporters,fatty acid biosynthesis and purine metabolism were associated with growth promotion in algae treated with 4μg/L ERY.An overcompensation was possibly activated by the low level of ERY in algae where more resources were reallocated to efficiently restore the temporary impairments,ultimately leading to the outperformance of growth.By contrast,algal growth inhibition in the 80 and 120μg/L ERY treatments was likely attributed to the dysfunction of metabolomic pathways related to ABC transporters,energy metabolism and metabolism of nucleosides.Apart from binding of ERY to the 50S subunit of ribosomes to inhibit protein translation as in bacteria,the data presented here indicate that inhibition of protein translation and growth performance of algae by ERY may also result from the suppression of amino acid biosynthesis and aminoacyl-tRNA biosynthesis.This study provides novel insights into the dose-dependent toxicity of ERY on R.subcapitata.

An Arabidopsis ABC Transporter Mediates Phosphate Deficiency-Induced Remodeling of Root Architecture by Modulating Iron Homeostasis in Roots

The remodeling of root architecture is a major developmental response of plants to phosphate （Pi） deficiency and is thought to enhance a plant＇s ability to forage for the available Pi in topsoil. The underlying mechanism controlling this response, however, is poorly understood. In this study, we identified an Arabidopsis mutant, hps 10 （hypersensitive to Pi starvation 10）, which is morphologically normal under Pi sufficient condition but shows increased inhibition of primary root growth and enhanced production of lateral roots under Pi defi- ciency, hpslO is a previously identified allele （als3-3） of the ALUMINUM SENSITIVE3 （ALS3） gene, which is involved in plant tolerance to aluminum toxicity. Our results show that ALS3 and its interacting protein AtSTAR1 form an ABC transporter complex in the tonoplast. This protein complex mediates a highly electro- genic transport in Xenopus oocytes. Under Pi deficiency, als3 accumulates higher levels of Fe3＋ in its roots than the wild type does. In Arabidopsis, LPR1 （LOW PHOSPHATE ROOT1） and LPR2 encode ferroxidases, which when mutated, reduce Fe3＋ accumulation in roots and cause root growth to be insensitive to Pi defi- ciency. Here, we provide compelling evidence showing that ALS3 cooperates with LPR1/2 to regulate Pi deficiency-induced remodeling of root architecture by modulating Fe homeostasis in roots.

ABC efflux transporters at blood-central nervous system barriers and their implications for treating spinal cord disorders

The barriers present in the interfaces between the blood and the central nervous system form a major hurdle for the pharmacological treatment of central nervous system injuries and diseases.The family of ATP-binding cassette(ABC)transporters has been widely studied regarding efflux of medications at blood-central nervous system barriers.These efflux transporters include P-glycoprotein(abcb1),‘breast cancer resistance protein'(abcg2)and the various‘multidrug resistance-associated proteins'(abccs).Understanding which efflux transporters are present at the blood-spinal cord,blood-cerebrospinal fluid and cerebrospinal fluid-spinal cord barriers is necessary to determine their involvement in limiting drug transfer from blood to the spinal cord tissue.Recent developments in the blood-brain barrier field have shown that barrier systems are dynamic and the profile of barrier defenses can alter due to conditions such as age,disease and environmental challenge.This means that a true understanding of ABC efflux transporter expression and localization should not be one static value but instead a range that represents the complex patient subpopulations that exist.In the present review,the blood-central nervous system barrier literature is discussed with a focus on the impact of ABC efflux transporters on:(i)protecting the spinal cord from adverse effects of systemically directed drugs,and(ii)limiting centrally directed drugs from accessing their active sites within the spinal cord.

Thermodynamics of ABC transporters

ABC transporters form the largest of all transporter families, and their structural study has made tremen- dous progress over recent years. However, despite such advances, the precise mechanisms that determine the energy-coupling between ATP hydrolysis and the con- formational changes following substrate binding remain to be elucidated. Here, we present our thermodynamic analysis for both ABC importers and exporters, and introduce the two new concepts of differential-binding energy and elastic conformational energy into the dis- cussion. We hope that the structural analysis of ABC transporters will henceforth take thermodynamic aspects of transport mechanisms into account as well.

Beauvericin counteracted multi-drug resistant Candida albicans by blocking ABC transporters

Multi-drug resistance of pathogenic microorganisms is becoming a serious threat,particularly to immunocompromised populations.The high mortality of systematic fungal infections necessitates novel antifungal drugs and therapies.Unfortunately,with traditional drug discovery approaches,only echinocandins was approved by FDA as a new class of antifungals in the past two decades.Drug efflux is one of the major contributors to multi-drug resistance,the modulator of drug efflux pumps is considered as one of the keys to conquer multi-drug resistance.In this study,we combined structure-based virtual screening and whole-cell based mechanism study,identified a natural product,beauvericin(BEA)as a drug efflux pump modulator,which can reverse the multi-drug resistant phenotype of Candida albicans by specifically blocking the ATP-binding cassette(ABC)transporters;meantime,BEA alone has fungicidal activity in vitro by elevating intracellular calcium and reactive oxygen species(ROS).It was further demonstrated by histopathological study that BEA synergizes with a sub-therapeutic dose of ketoconazole(KTC)and could cure the murine model of disseminated candidiasis.Toxicity evaluation of BEA,including acute toxicity test,Ames test,and hERG(human ether-a-go-go-related gene)test promised that BEA can be harnessed for treatment of candidiasis,especially the candidiasis caused by ABC overexpressed multi-drug resistant C.albicans.

Two ABCI family transporters,OsABCI15 and OsABCI16,are involved in grain-filling in rice

Seed development is critical for plant reproduction and crop yield,with panicle seed-setting rate,grain-filling,and grain weight being key seed characteristics for yield improvement.However,few genes are known to regulate grain filling.Here,we identify two adenosine triphosphate(ATP)-binding cassette(ABC)I-type transporter genes,OsABCI15 and OsABCI16,involved in rice grain-filling.Both genes are highly expressed in developing seeds,and their proteins are localized to the plasma membrane and cytosol.Interestingly,knockout of OsABCI15 and OsABCI16 results in a significant reduction in seed-setting rate,caused predominantly by the severe empty pericarp phenotype,which differs from the previously reported low seed-setting phenotype resulting from failed pollination.Further analysis indicates that OsABCI15 and OsABCI16 participate in ion homeostasis and likely export ions between filial tissues and maternal tissues during grain filling.Importantly,overexpression of OsABCI15 and OsABCI16 enhances the seed-setting rate and grain yield in transgenic plants and decreases ion accumulation in brown rice.Moreover,the OsABCI15/16 orthologues in maize exhibit a similar role in kernel development,as demonstrated by their disruption in transgenic maize.Therefore,ourfindings reveal the important roles of two ABC transporters in cereal grain filling,highlighting their value in crop yield improvement.

High glucose decreases the expression of ATP-binding cassette transporter G1 in human vascular smooth muscle cells

Objective：ATP-binding cassette transporters（ABC） A1 and G1 play an important role in mediating cholesterol efflux and preventing macrophage foam cell formation. In this study, we examined the regulation of ABC transporters by high glucose in human vascular smooth muscle cells（VSMCs）, the other precursor of foam cells. Methods：Incubation of human VSMCs with D-glucose（5 to 30 mM） for 1 to 7 days in the presence or absence of antioxidant and nuclear factor（NF）- κ B inhibitors, the expressions of ABCA1 and ABCG1 were analyzed by real time PCR and Western blotting. Results：High glucose decreased ABCG1 mRNA and protein expression in cultured VSMCs, whereas the expression of ABCA1 was not significantly decreased. Down-regulation of ABCG1 mRNA expression by high glucose was abolished by antioxidant N-acetyl-L-cysteine（NAC） and NF- κ B inhibitors, BAY 11-7085 and tosyl-phenylalanine chloromethyl-ketone（TPCK）. Conclusion：High glucose suppresses the expression of ABCG1 in VSMCs, which is the possible mechanism of VSMC derived foam cell transformation.

Molecular Modelling of Human Multidrug Resistance Protein 5 (ABCC5)

Multidrug resistance protein 5 (MRP5/ABCC5) is a 161 kDa member of the super family of ATP-binding cassette (ABC) superfamily of transmembrane transporters that is clinically relevant for its ability to confer multidrug resistance by actively e?uxing anticancer drugs. ABCC5 has also been identified as an efflux transporter of cGMP (cyclic guanosine monophosphate). Elevated intracellular levels of cGMP in cancer cells have been implicated in several clinical studies, that may induce apoptosis, and as a result many different cancer cells seem to overcome this deleterious effect by increased efflux of cGMP through ABCC5. Thus inhibition of ABCC5 may have cytotoxic effects mediated through cGMP and it will also increase the intracellular concentration of other drugs that are aimed for the treatment of cancer which are otherwise exported out of the cells. Considering the functional importance and lack of X-ray crystal structure of ABCC5, present work was undertaken to construct 3D structure of protein using homology modeling protocol of YASARA structure (V. 16.3.28). In this study, five different ABC templates (PDB ID’s: 4F4C, 4Q9H, 4M1M, 4M2T and 4KSD) were used for homology modeling. Five models were constructed on each template and a hybrid model was built using all five templates. All models were refined and ranked as per their overall Z-score. The top ranked ABBC5 model was based on template 4Q9H that had 91.2% of residues in allowed regions as revealed by PROCHECK-NMR and the QMEAN score was 0.54 which indicated a reliable model. The results of the study and the proposed model can be further used for elucidating the structural and functional aspects of ABCC5 and to gain more insights to the molecular basis of ABCC5 inhibition through docking studies.

Arabidopsis ABCG14 forms a homodimeric transporter for multiple cytokinins and mediates long-distance transport of isopentenyladeninetype cytokinins

Cytokinins(CKs),primarily trans-zeatin(tZ)and isopentenyladenine(iP)types,play critical roles in plant growth,development,and various stress responses.Long-distance transport of tZ-type CKs meidated by Arabidopsis ATP-binding cassette transporter subfamily G14(AtABCG14)has been well studied;however,less is known about the biochemical properties of AtABCG14 and its transporter activity toward iP-type CKs.Here we reveal the biochemical properties of AtABCG14 and provide evidence that it is also required for long-distance transport of iP-type CKs.AtABCG14 formed homodimers in human(Homo sapiens)HEK293T,tobacco(Nicotiana tabacum),and Arabidopsis cells.Transporter activity assays of AtABCG14 in Arabidopsis,tobacco,and yeast(Saccharomyces cerevisiae)showed that AtABCG14 may directly transport multiple CKs,including iP-and tZ-type species.AtABCG14 expression was induced by iP in a tZ-type CKdeficient double mutant(cypDM)of CYP735A1 and CYP735A2.The atabcg14 cypDM triple mutant exhibited stronger CK-deficiency phenotypes than cypDM.Hormone profiling,reciprocal grafting,and 2H6-iP isotope tracer experiments showed that root-to-shoot and shoot-to-root long-distance transport of iP-type CKs were suppressed in atabcg14 cypDM and atabcg14.These results suggest that AtABCG14 participates in three steps of the circular long-distance transport of iP-type CKs:xylem loading in the root for shootward transport,phloem unloading in the shoot for shoot distribution,and phloemunloading in the root for root distribution.We found that AtABCG14 displays transporter activity toward multiple CK species and revealed its versatile roles in circular long-distance transport of iP-type CKs.These findings provide newinsights into the transport mechanisms of CKs and other plant hormones.

Rice Stomatal Closure Requires Guard Cell Plasma Membrane ATP-Binding Cassette Transporter RCN1/OsABCG5

Water stress is one of the major environmental stresses that affect agricultural production worldwide. Water loss from plants occurs primarily through stomatal pores. Here, we report that an Oryza sativa half-size ATP-binding cassette （ABC） subfamily G protein, RCN 1/OsABCG5, is involved in stomatal closure mediated by phytohormone abscisic acid （ABA） accumulation in guard cells. We found that the GFP-RCN1/ OsABCG5-fusion protein was localized at the plasma membrane in guard cells. The percentage of guard cell pairs containing both ABA and GFP-RCN1/OsABCG5 increased after exogenous ABA treatment, whereas they were co-localized in guard cell pairs regardless of whether exogenous ABA was applied. ABA application resulted in a smaller increase in the percentage of guard cell pairs containing ABA in rcnl mutant （A684P） and RCN1-RNAi than in wild-type plants. Furthermore, polyethylene glycol （drought stress）-inducible ABA accumulation in guard cells did not occur in rcnl mutants. Stomata closure mediated by exogenous ABA application was strongly reduced in rcnl mutants. Finally, rcnl mutant plants had more rapid water loss from detached leaves than the wild-type plants. These results indicate that in response to drought stress, RCN1/OsABCG5 is involved in accumulation of ABA in guard cells, which is indispensable for stomatal closure.

NaPDR1 and NaPDR1-like are essential for the resistance of Nicotiana attenuata against fungal pathogen Alternaria alternata

Pleiotropic drug resistance(PDR) transporters are widely distributed membrane proteins catalyzing the export or import of a diverse array of molecules, and are involved in many plant responses to biotic and abiotic stresses. However, it is unclear whether PDRs are involved in Nicotiana attenuata resistance to the necrotic fungal pathogen Alternaria alternata. In this study, transcriptional levels of both NaPDR1 and NaPDR1-like were highly induced in N. attenuata leaves after A. alternata inoculation. Interestingly,silencing NaPDR1 or NaPDR1-like individually had little effect on N. attenuata resistance to A. alternata;however, when both genes were co-silenced plants became highly susceptible to the fungus, which was associated with elevated JA and ethylene responses. Neither NaPDR1 nor NaPDR1-like was significantly elicited by exogenous treatment with methyl jasmonate(MeJA), whereas both were highly induced by ethylene. The elicitation levels of both genes by A. alternata were significantly reduced in plants with impaired JA or ethylene signaling pathways. Thus, we conclude that both NaPDR1 and NaPDR1-like function redundantly to confer resistance against A. alternata in N. attenuata, and the elicitation of the transcripts of both genes by the fungus is partially dependent on ethylene and jasmonate signaling.

The roles of the human ATP-binding cassette transporters P-glycoprotein and ABCG2 in multidrug resistance in cancer and at endogenous sites: future opportunities for structure-based drug design of inhibitors

The ATP-binding cassette(ABC)transporters P-glycoprotein(P-gp)and ABCG2 are multidrug transporters that confer drug resistance to numerous anti-cancer therapeutics in cell culture.These findings initially created great excitement in the medical oncology community,as inhibitors of these transporters held the promise of overcoming clinical multidrug resistance in cancer patients.However,clinical trials of P-gp and ABCG2 inhibitors in combination with cancer chemotherapeutics have not been successful due,in part,to flawed clinical trial designs resulting from an incomplete molecular understanding of the multifactorial basis of multidrug resistance(MDR)in the cancers examined.The field was also stymied by the lack of high-resolution structural information for P-gp and ABCG2 for use in the rational structure-based drug design of inhibitors.Recent advances in structural biology have led to numerous structures of both ABCG2 and P-gp that elucidated more clearly the mechanism of transport and the polyspecific nature of their substrate and inhibitor binding sites.These data should prove useful helpful for developing even more potent and specific inhibitors of both transporters.As such,although possible pharmacokinetic interactions would need to be evaluated,these inhibitors may show greater effectiveness in overcoming ABC-dependent multidrug resistance in combination with chemotherapeutics in carefully selected subsets of cancers.Another perhaps even more compelling use of these inhibitors may be in reversibly inhibiting endogenously expressed P-gp and ABCG2,which serve a protective role at various blood-tissue barriers.Inhibition of these transporters at sanctuary sites such as the brain and gut could lead to increased penetration by chemotherapeutics used to treat brain cancers or other brain disorders and increased oral bioavailability of these agents,respectively.

Identification of two ABCC transporters involved in malathion detoxification in the red flour beetle,Tribolium castaneum

ABC transporters have been suggested to be involved in insecticide detoxification in different insect species mainly based on the indirect observation of transcriptional upregulation of ABC gene expression in response to insecticide exposure.Previous studies performed by us and others in the red flour beetle,Tribolium castaneum,have analyzed the function of TcABCA-C and TcABCG-H genes using RNA interference(RNAi)and demonstrated that specific TcABCA and TcABCC genes are involved in the elimination of the pyrethroid tefluthrin and the benzoylurea diflubenzuron,because gene silencing increased the beetle's susceptibility to the insecticides.In this study,we focused on the potential functions of TcABCA-C genes in detoxification of the pyrethroid cyfluthrin(CF),the organophosphate malathion(MAL)and the diacylhdyazine tebufenozide(TBF).Analysis of transcript levels of selected TcABCA-C genes in response to treatment with these three chemically unrelated insecticides revealed that some genes were particularly upregulated after insecticide treatment.In addition,the ABC inhibitor verapamil synergized significantly the toxicity of MAL but only negligibly CF and TBF toxicities.Finally,silencing of two TcABCC genes by RNAi revealed a significant increase in susceptibility to MAL.In contrast,we did not observe a significant increase in insecticide-induced mortalities when knocking down TcABC genes in larvae treated with CF or TBF,although they were upregulated in response to insecticide treatment.Our results suggest that two pleiotropic ABCC transporters expressed in metabolic and excretory tissues contribute to the elimination of MAL.

Cryo-EM structures for the Mycobacterium tuberculosis iron-loaded siderophore transporter IrtAB

The adenosine 5'-triphosphate(ATP)-binding cassette(ABC)transporter,IrtAB,plays a vital role in the replication and viability of Mycobacterium tuberculosis(Mtb),where its function is to import iron-loaded siderophores.Unusually,it adopts the canonical type IV exporter fold.Herein,we report the structure of unliganded Mtb IrtAB and its structure in complex with ATP,ADP,or ATP analogue(AMP-PNP)at resolutions ranging from 2.8 to 3.5Å.The structure of IrtAB bound ATP-Mg2+shows a“head-to-tail”dimer of nucleotide-binding domains(NBDs),a closed amphipathic cavity within the transmembrane domains(TMDs),and a metal ion liganded to three histidine residues of IrtA in the cavity.Cryo-electron microscopy(Cryo-EM)structures and ATP hydrolysis assays show that the NBD of IrtA has a higher affinity for nucleotides and increased ATPase activity compared with IrtB.Moreover,the metal ion located in the TM region of IrtA is critical for the stabilization of the conformation of IrtAB during the transport cycle.This study provides a structural basis to explain the ATP-driven conformational changes that occur in IrtAB.

Plant pleiotropic drug resistance transporters:Transport mechanism, gene expression,and function

Pleiotropic drug resistance （PDR） transporters belonging to the ABCG subfamily of ATP-binding cassette （ABC） transporters are identified only in fungi and plants. Members of this family are expressed in plants in response to various biotic and abiotic stresses and transport a diverse array of moleculesacross membranes, Although their detailed transport mechanism is largely unknown, they play important roles in detoxification processes, preventing water loss, transport of phytohormones, and secondary metabolites. This review provides insights into transport mechanisms of plant PDR transporters, their expression profiles, and multitude functions in plants.

The pharmacological impact of ATP-binding cassette drug transporters on vemurafenib-based therapy

Melanoma is the most serious type of skin cancer and one of the most common cancers in the world.Advanced melanoma is often resistant to conventional therapies and has high potential for metastasis and low survival rates.Vemurafenib is a small molecule inhibitor of the BRAF serine-threonine kinase recently approved by the United States Food and Drug Administration to treat patients with metastatic and unresectable melanomas that carry an activating BRAF(V600E)mutation.Many clinical trials evaluating other therapeutic uses of vemurafenib are still ongoing.The ATP-binding cassette(ABC)transporters are membrane proteins with important physiological and pharmacological roles.Collectively,they transport and regulate levels of physiological substrates such as lipids,porphyrins and sterols.Some of them also remove xenobiotics and limit the oral bioavailability and distribution of many chemotherapeutics.The overexpression of three major ABC drug transporters is the most common mechanism for acquired resistance to anticancer drugs.In this review,we highlight some of the recent findings related to the effect of ABC drug transporters such as ABCB1 and ABCG2 on the oral bioavailability of vemurafenib,problems associated with treating melanoma brain metastases and the development of acquired resistance to vemurafenib in cancers harboring the BRAF(V600E)mutation.

Emerging consensus on the mechanism of polyspecific substrate recognition by the multidrug transporter P-glycoprotein

P-glycoprotein(P-gp or ABCB1)is a member of the broad family of ABC transporters.P-gp participates in the establishment of physiological barriers limiting cellular access of a large number of toxic compounds.It thus plays important roles in the pharmacokinetics of these compounds.Cancer cells and cells infected by viruses exploit the presence of P-gp to fend off drug treatment,rendering them multidrug-resistant.Overcoming multidrug resistance caused by expression of ABC transporters has gained increasing attention in the field of drug development.Recently,studies of P-gp,especially from structural investigations by both cryo-electron microscopy and X-ray crystallography,have provided high-resolution mechanistic details for the function of this transporter.Structures with increasing resolution and accuracy in various substrate-and inhibitor-bound forms are available for analysis and a consensus on the mechanism of substrate polyspecificity is emerging.The use of new structural information may aid development of P-gp inhibitors as well as compounds that may bypass P-gp action.

ABCA1 is associated with the development of acquired chemotherapy resistance and predicts poor ovarian cancer outcome

Aim:This study investigated the ATP binding cassette(ABC)transporter(ABCA1,ABCB1,ABCB3,ABCC2 and ABCG2)expression in high grade serous ovarian cancer(HGSOC)tissues,cell lines and primary cells to determine their potential relationship with acquired chemotherapy resistance and patient outcome.Methods:ABC transporter mRNA and protein expression(ABCA1,ABCB1,ABCB3,ABCC2 and ABCG2)was assessed in publicly available datasets and in a tissue microarray(TMA)cohort of HGSOC at diagnosis,respectively.ABC transporter mRNA expression was also assessed in chemosensitive ovarian cancer cell lines(OVCAR-5 and CaOV3)versus matching cell lines with acquired carboplatin resistance and in primary HGSOC cells from patients with chemosensitive disease at diagnosis(n=10)as well as patients with acquired chemotherapy resistance at relapse(n=6).The effects of the ABCA1 inhibitor apabetalone in carboplatin-sensitive and-resistant cell lines were also investigated.Results:High ABCA1 mRNA and protein expression was found to be significantly associated with poor patient outcome.ABCA1 mRNA and protein levels were significantly increased in ovarian cancer cell lines(OVCAR-5 CBPR and CaOV3 CBPR)with acquired carboplatin resistance.ABCA1 mRNA was significantly increased in primary HGSOC cells obtained from patients with acquired chemotherapy resistance.Apabetalone treatment reduced ABCA1 protein expression and increased the sensitivity of both parental and carboplatin-resistant ovarian cancer cells to carboplatin.Conclusion: These results suggest that inhibiting ABCA1 transporter may be useful in overcoming acquired chemotherapy resistance and improving outcome for patients with HGSOC.

New xylose transporters support the simultaneous consumption of glucose and xylose in Escherichia coli

Glucose and xylose are two major components of lignocellulose.Simultaneous consumption of glucose and xylose is critical for engineered microorganisms to produce fuels and chemicals from lignocellulosic biomass.Although many production limitations have been resolved,glucose‐induced inhibition of xylose transport remains a challenge.In this study,a cell growthbased screening strategy was designed to identify xylose transporters uninhibited by glucose.The glucose pathway was genetically blocked in Escherichia coli so that glucose functions only as an inhibitor and cells need xylose as the carbon source for survival.Through adaptive evolution,omics analysis and reverse metabolic engineering,a new phosphoenolpyruvate:carbohydrate phosphotransferase system(PTS)galactitol transporter(GalABC,encoded by EcolC_1640,EcolC_1641,and EcolC_1642 genes)that is not inhibited by glucose was identified.Inactivation of adenylate cyclase led to increased expression of the EcolC_1642 gene,and a point mutation in gene EcolC_1642(N13S)further enhanced xylose transport.During the second round of gene mining,AraE and a new ABC transporter(AraFGH)of xylose were identified.A point mutation in the transcription regulator araC(L156I)caused increased expression of araE and araFGH genes without arabinose induction,and a point mutation in araE(D223Y)further enhanced xylose transport.These newly identified xylose transporters can support the simultaneous consumption of glucose and xylose and have potential use in producing chemicals from lignocellulose.