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.

Based on the concept presented in this review, TKI anticancer drugs are considered as compounds recognized by the cellular mechanisms handling xenobiotics. Accordingly, novel anticancer therapies should equally focus on the effectiveness of target inhibition and exploration of potential interactions of the designed molecules by membrane transporters. Thus, targeted hydrophobic small molecule compounds should also be screened to evade xenobiotic-sensing cellular mechanisms.

The tight control of proliferation, survival, and apoptosis of bone marrow progenitor cells has a major role in allowing normal hematopoiesis. A variety of cytokines, including GM-CSF 1 and IL-3, regulate viability, proliferation, differentiation, and function of hemopoietic cells (1). Hemopoietic cells undergo apoptotic cell death in the absence of growth factors, whereas an escape from this regulation may result in leukemogenesis.The human GM-CSF/IL-3-dependent myelo-erythroid cell line, TF-1, has been reported to undergo apoptosis upon hormone deprivation (2). Apoptosis of these cells can be suppressed by re-administration of GM-CSF, IL-3, or IL-4 (3) or overexpression of Bcl-2 (4). The alteration of intracellular Ca 2ϩ concentration control cellular processes as diverse as proliferation, development, contraction, or secretion. However, when exceeding its normal spatial and temporal boundaries, Ca 2ϩ can result in cell death through both necrosis and apoptosis (7).The purpose of this study was to delineate how Ca 2ϩ can have multiple roles in cell survival and growth. Ca 2ϩ functions directly through regulation of gene expression, e.g. c-fos (8) or via activating Ca 2ϩ -dependent enzymatic action, e.g. calcium/ calmodulin-dependent kinases II and IV in neurons, calcineurin in lymphocytes, and in conjunction with other signaling pathways such as those regulated through MAPK and PI3K * This work was supported by the Hungarian Research Foundation (Grant OTKA T029291) by the Scientific Research Council, Ministry of Health (Grant ETT 206/2001), and by NRDP 1/024, Ministry of Education, Hungary. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.ʈ To whom correspondence should be addressed. Tel./Fax: 36-1-372-4353; E-mail: magocsi@biomembrane.hu.1 The abbreviations used are: GM-CSF, granulocyte-macrophage colony-stimulating factor; AP-1, activator protein-1; BAPTA-AM, 1,2-bis(o-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid tetra(acetoxymethyl)ester; [Ca 2ϩ ] i , cytoplasmic free calcium concentration; CPA, cyclopiazonic acid; CREB, cAMP response element binding protein; MAPK, mitogen-activated protein kinase; ERK1/2, extracellular signalregulated kinase; MEK, MAPK/ERK kinase; Egr-1, early growth response protein-1; Elk-1, Ets domain protein-1; p90RSK, 90-kDa ribosomal S6 kinase; PD98059, 2Ј-amino-3-methoxyflavone (MEK-1 inhibitor); UO126, 1,4-diamino-2,3-dicyano-1,4-bis[2-aminophenylthio]-butadiene (MEK1/2 inhibitor); PI3K, phosphoinositide 3-kinase; SRE, serum response element; STAT-5, signal transducer and activator of transcription-5; IL-3, interleukin-3; FACS, fluorescence-activated cell

The role of extracellular vesicles (EVs) in mediating the immunosuppressory properties of mesenchymal stem cells (MSCs) has recently attracted remarkable scientific interest. The aim of this work was to analyze the transport mechanisms of membrane and cytoplasmic components between T lymphocytes and adipose tissue-derived MSCs (AD-MSCs), by focusing on the role of distinct populations of EVs, direct cell-cell contacts, and the soluble mediators per se in modulating T lymphocyte function. We found that neither murine thymocytes and human primary T cells nor Jurkat lymphoblastoid cells incorporated appreciable amounts of MSC-derived microvesicles (MVs) or exosomes (EXOs). Moreover, these particles had no effect on the proliferation and IFN-γ production of in vitro-stimulated primary T cells. In contrast, AD-MSCs incorporated large amounts of membrane components from T cells as an intensive uptake of EXOs and MVs could be observed. Interestingly, we found a bidirectional exchange of cytoplasmic components between human AD-MSCs and primary T lymphocytes, mediated by tunneling nanotubes (TNTs) derived exclusively from the T cells. In contrast, TNTs couldn't be observed between AD-MSCs and the Jurkat cells. Our results reveal a novel and efficient way of intercellular communication between MSCs and T cells, and may help a better understanding of the immunomodulatory function of MSCs.

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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.

No abstract

ABCG2 is a plasma membrane multidrug transporter with an established role in the cancer drug-resistance phenotype. This protein is expressed in a variety of tissues, including several types of stem cell. Although ABCG2 is not essential for life, knock-out mice were found to be hypersensitive to xenobiotics and had reduced levels of the side population of hematopoietic stem cells. Previously we have shown that ABCG2 is present in human embryonic stem cell (hESC) lines, with a heterogeneous expression pattern. In this study we examined this heterogeneity, and investigated whether it is related to stress responses in hESCs. We did not find any difference between expression of pluripotency markers in ABCG2-positive and negative hESCs; however, ABCG2-expressing cells had a higher growth rate after cell separation. We found that some harmful conditions (physical stress, drugs, and UV light exposure) are tolerated much better in the presence of ABCG2 protein. This property can be explained by the transporter function which eliminates potential toxic metabolites accumulated during stress conditions. In contrast, mild oxidative stress in hESCs caused rapid internalization of ABCG2, indicating that some environmental factors may induce removal of this transporter from the plasma membrane. On the basis of these results we suggest that a dynamic balance of ABCG2 expression at the population level has the advantage of enabling prompt response to changes in the cellular environment. Such actively maintained heterogeneity might be of evolutionary benefit in protecting special cell types, including pluripotent stem cells.