Multidrug resistance phenotypes and MRS2 mitochondrial magnesium channel: two players from one stemness?

Commentary to:Human mitochondrial Mrs2 protein promotes multidrug resistance in gastric cancer cells by regulating p27, cyclin D1 expression and cytochrome C releaseYu Chen, Xufeng Wei, Pengfei Yan, Ying Han , Kai-Chun Wu, Shiren Sun, Daiming Fa

Missense mutations in the perforin (PRF1) gene as a cause of hereditary cancer predisposition

Perforin, a pore-forming toxin released from secretory granules of NK cells and CTLs, is essential for their cytotoxic activity against infected or cancerous target cells. Bi-allelic loss-of-function mutations in the perforin gene are invariably associated with a fatal immunoregulatory disorder, familial haemophagocytic lymphohistiocytosis type 2 (FHL2), in infants. More recently, it has also been recognized that partial loss of perforin function can cause disease in later life, including delayed onset FHL2 and haematological malignancies. Herein we report a family in which a wide range of systemic inflammatory and neoplastic manifestations have occurred across three generations. We found that disease was linked to two missense perforin gene mutations (encoding A91V, R410W) that cause protein misfolding and partial loss of activity. These cases link the partial loss of perforin function with some solid tumours that are known to be controlled by the immune system, as well as haematological cancers. Our findings also demonstrate that perforin gene mutations can contribute to hereditary cancer predisposition

Mechanism For Copper(II)-Mediated Disaggregation Of A Porphyrin J-Aggregate

J-aggregates of anionic meso-tetrakis(4-sulfonatophenyl)porphyrin form at intermediate pH (2.3–3.1) in the presence of NiSO₄ or ZnSO₄ (ionic strength, I.S. = 3.2 M). These aggregates convert to monomeric porphyrin units via metallation with copper(II) ions. The kinetics for the disassembly process, as monitored by UV/vis spectroscopy, exhibits zeroth-order behavior. The observed zeroth-order rate constants show a two-term dependence on copper(II) ion concentrations: linear and second order. Also observed is an inverse dependence on hydrogen ion concentration. Activation parameters have been determined for the disassembly process leading to ΔH^≠ = (+163 ± 15) kJ·mol⁻¹ and ΔS^≠ = (+136 ± 11) J·K⁻¹. A mechanism is proposed in which copper(II) cation is in pre-equilibrium with a reactive site at the rim of the J-aggregate. An intermediate copper species is thus formed that eventually leads to the final metallated porphyrin either through an assisted attack of a second metal ion or through a direct insertion of the metal cation into the macrocycle core

Lipid specificity of the immune effector perforin

Perforin is a pore forming protein used by cytotoxic T lymphocytes to remove cancerous or virus-infected cells during immune response. During the response, the lymphocyte membrane becomes refractory to perforin function by accumulating densely ordered lipid rafts and externalizing negatively charged lipid species. The dense membrane packing lowers the capacity of perforin to bind, and negatively charged lipids scavenge any residual protein before pore formation. Using atomic force microscopy on model membrane systems, we here provide insight into the molecular basis of perforin lipid specificity

Production of anti-breast cancer monoclonal antibodies using a glutathione-S-transferase-MUC1 bacterial fusion protein.

Two murine Mabs VA1(IgG1) and VA2(IgG1) were produced against a bacterial fusion protein comprising glutathione S-transferase and five tandem repeats of the MUC1 protein. Using the immunoperoxidase staining technique, VA1 detected 46/53 and VA2 detected 48/53 breast cancers and both also reacted with a range of other human epithelial carcinomas. In addition VA1 gave weak reactions with normal breast tissues whereas VA2 was non-reactive and could be a relatively tumour specific antibody for breast cancer. The antibodies were also tested by ELISA-VA1 reacted weakly with glycosylated HMFG but strongly with deglycosylated HMFG, whereas VA2 reacted strongly with both forms of HMFG. The reactivities of the two Mabs with synthetic peptides of the MUC1 tandem repeat were used to map the epitopes recognised by VA1 (amino acids RPAPGS) and VA2 (amino acids DTRPA). The use of fusion proteins provides another means of immunisation to produce anti-tumour antibodies

TRPM6 is essential for magnesium uptake and epithelial cell function in the colon

Intestinal magnesium (Mg) uptake is essential for systemic Mg homeostasis. Colon cells express the two highly homologous transient receptor potential melastatin type (TRPM) 6 and 7 Mg2+ channels, but their precise function and the consequences of their mutual interaction are not clear. To explore the functional role of TRPM6 and TRPM7 in the colon, we used human colon cell lines that innately express both channels and analyzed the functional consequences of genetic knocking-down, by RNA interference, or pharmacological inhibition, by NS8593, of either channel. TRPM7 silencing caused an increase in Mg2+ influx, and correspondingly enhanced cell proliferation and migration, while downregulation of TRPM6 did not affect significantly either Mg2+ influx or cell proliferation. Exposure to the specific TRPM6/7 inhibitor NS8593 reduced Mg2+ influx, and consequently cell proliferation and migration, but Mg supplementation rescued the inhibition. We propose a model whereby in colon cells the functional Mg2+ channel at the plasma membrane may consist of both TRPM7 homomers and TRPM6/7 heteromers. A different expression ratio between the two proteins may result in different functional properties. Altogether, our findings confirm that TRPM6 cannot be replaced by TRPM7, and that TRPM6/7 complexes and TRPM6/7-mediated Mg2+ influx are indispensable in human epithelial colon cells

Cytotoxic T lymphocyte–induced killing in the absence of granzymes A and B is unique and distinct from both apoptosis and perforin-dependent lysis

Cytotoxic T lymphocyte (CTL)–induced death triggered by the granule exocytosis pathway involves the perforin-dependent delivery of granzymes to the target cell. Gene targeting has shown that perforin is essential for this process; however, CTL deficient in the key granzymes A and B maintain the ability to kill their targets by granule exocytosis. It is not clear how granzyme AB−/− CTLs kill their targets, although it has been proposed that this occurs through perforin-induced lysis. We found that purified granzyme B or CTLs from wild-type mice induced classic apoptotic cell death. Perforin-induced lysis was far more rapid and involved the formation of large plasma membrane protrusions. Cell death induced by granzyme AB−/− CTLs shared similar kinetics and morphological characteristics to apoptosis but followed a distinct series of molecular events. Therefore, CTLs from granzyme AB−/− mice induce target cell death by a unique mechanism that is distinct from both perforin lysis and apoptosis