1. Human and murine neuroblastoma cell lines were used to investigate, by the whole-cell patch-clamp technique, the properties of a novel inward-rectifying K+ current (IIR) in the adjustment of cell resting potential (Vrest), which was in the range -40 to -20 mV. 2. When elicited from a holding potential of 0 mV, IIR was completely inactivated with time constants ranging from 13 ms at -140 mV to 4 5 s at -50 mV. The steady-state inactivation curve (h(V)) was found to be independent of [Nae]0 and [K+]O (2-80 mM) and could be fitted to a Boltzmann curve with a steep slope factor of 5-6, and a V½ around Vrest. synchronization in the GO-GQ phase of the cell cycle, or at the G1-S boundaries, the cells reduced their variability of h(V). The same occurred after cell synchronization in G1 by treatment with retinoic acid. 5. The experimental data could be fitted to a classical model of an inward rectifier, after removing the dependence of conductance activation on (V -EK), and incorporating an inactivation with an intrinsic voltage dependence. Moreover, the model predicts, for this novel inward rectifier and in contrast with the classical inward rectifier, the incapacity of maintaining, in physiological media, a Vr,st more negative than -35 to -40 mV, which is an important feature of cancer cells.

The role of K ؉ channel activity during cell cycle progression has become a research topic of considerable interest. Blocking of K ؉ channels inhibits the proliferation of many cell types, although the mechanism of this inhibition is unclear. There is speculation that K ؉ channels differentially regulate the electrical potential of the plasma membrane (V m ) during proliferation. We have demonstrated that in tumor cells the value of V m is clamped to rather depolarized values by K ؉ channels belonging to the HERG family. We report here that tumor cell lines preferentially express the herg1 gene and a truncated, N-deleted form that corresponds to herg1b. This alternative transcript is also expressed in human primary acute myeloid leukemias. Both HERG1 and HERG1B proteins are expressed on the plasma membrane of tumor cells and can form heterotetramers. The expression of HERG protein isoforms is strongly cell cycle-dependent, accounting for variations in HERG currents along the mitotic cycle. Moreover, the blocking of HERG channels dramatically impairs cell growth of HERG-bearing tumor cells. These results suggest that modulated expression of different K ؉ channels is the molecular basis of a novel mechanism regulating neoplastic cell proliferation.

The acquisition of the capacity to invade surrounding tissues confers a more malignant phenotype to tumor cells and is necessary for the establishment of metastases. The understanding of the molecular mechanisms underlying cell invasion in human solid tumors such as colorectal cancers could provide not only more sensitive prognostic analyses but also novel molecular targets for cancer therapy.We report in this article that K ؉ ion channels belonging to the HERG family are important determinants for the acquisition of an invasive phenotype in colorectal cancers. The herg1 gene and HERG1 protein are expressed in many colon cancer cell lines, and the activity of HERG channels modulates colon cancer cell invasiveness. Moreover, the amount of HERG1 protein expressed on the plasma membrane is directly related to the invasive phenotype of colon cancer cells.Finally, both the herg1 gene and HERG1 protein were expressed in a high percentage of primary human colorectal cancers, with the highest incidence occurring in metastatic cancers, whereas no expression could be detected either in normal colonic mucosa or in adenomas.

An important target in the understanding of the pathogenesis of acute myeloid leukemias (AML) relies on deciphering the molecular features of normal and leukemic hemopoietic progenitors. In particular, the analysis of the mechanisms involved in the regulation of cell proliferation is decisive for the establishment of new targeted therapies. To gain further insight into this topic we report herein a novel approach by analyzing the role of HERG K + channels in the regulation of hemopoietic cell proliferation. These channels, encoded by the human ether-agò -gò -related gene (herg), belong to a family of K + channels, whose role in oncogenesis has been recently demonstrated. We report here that herg is switched off in normal peripheral blood mononuclear cells (PBMNC) as well as in circulating CD34 + cells, however, it is rapidly turned on in the latter upon induction of the mitotic cycle. Moreover, herg appears to be constitutively activated in leukemic cell lines as well as in the majority of circulating blasts from primary AML. Evidence is also provided that HERG channel activity regulates cell proliferation in stimulated CD34 + as well as in blast cells from AML patients. These results open new perspectives on the pathogenetic role of HERG K + channels in leukemias.

Bone marrow cell liquid cultures were incubated at various oxygen concentrations ranging from 0% to 18% (air). The total number of cells in culture (CT) at the end of a 6-day incubation was found to be directly proportional to the oxygen concentration. As compared with air- incubated controls, cells recovered from severely hypoxic (1% oxygen) day-5 liquid cultures showed (1) the same day-7 colony-formation efficiency in semisolid culture (neutrophilic/monocytic colonies) or in spleen; (2) a higher day-14 spleen colony-formation efficiency; (3) an enhanced radio-protection ability; and (4) an increased marrow repopulation ability, as measured by determining either total cell number in recipient marrow MRAcell, or the capacity of the latter of generating day-7 neutrophilic/monocytic colonies in secondary in vitro assays (MRACFU-NM). Taking into account CT, the absolute numbers of progenitors in culture were also computed. The results showed that, with respect to time 0, incubation in air produced an increase in the number of day-7 CFUs and a decrease in the number of the other progenitors, whereas in hypoxic cultures all types of progenitors decreased. However, as compared with air-incubated controls, all progenitors, except cells sustaining MRACFU-NM, were reduced in hypoxic cultures. The degree of reduction paralleled the position of the progenitor in the hematopoietic hierarchy, being maximum for day-7 CFUs and null for cells sustaining MRACFU-NM, which, in fact, were better preserved in hypoxic cultures.

1. The regular firing of a Hodgkin-Huxley neurone endowed with fast Na¤ and delayed K¤ channels can be converted into adapting firing by appending HERG (human eag-related gene) channels. 2. The computer model predictions were verified by studying the firing properties of F-11 DRG neurone ² neuroblastoma hybrid cells induced to differentiate by long-term exposure to retinoic acid. These cells, which express HERG currents (IHERG), show clear spike-frequency adaptation of their firing when current clamped with long depolarizations. 3. In agreement with the prediction, the selective blocking of IHERG by class III antiarrhythmic drugs always led to the disappearance of the spike-frequency adaptation, and the conversion of adapting firing to regular firing. 4. It is proposed that, in addition to their role in the repolarization of the heart action potential, HERG channels may sustain a process of spike-frequency adaptation, and hence contribute to the control of burst duration in a way that is similar to that of the K¤ currents, IAHP, IC and IM. In addition to the known cardiac arrhythmia syndrome (LQT2), genetic mutations or an altered HERG expression could lead to continuous hyperexcitable states sustained by the inability of nerve or endocrine cells to accommodate to repetitive stimuli. This might help in clarifying the pathogenesis of still undefined idiopathic familial epilepsies.

Adhesive receptors of the integrin family are primarily involved in cell-extracellular matrix adhesion. Additionally, integrins trigger multiple signaling pathways that are involved in cell migration, proliferation, survival, and differentiation. We previously demonstrated that the activation of integrins containing the ␤ 1 subunit leads to a selective increase in potassium currents carried by the human ether-a-go-go-related gene (hERG) channels in neuroblastoma and leukemia cells; this current activation modulates adhesion-dependent differentiation in these cells. We hypothesized that the cross-talk between integrins and hERG channels could be traced back to the assembly of a macromolecular signaling complex comprising the two proteins. We tested this hypothesis in both SH-SY5Y neuroblastoma cells and in human embryonic kidney 293 cells stably transfected with hERG1 and, therefore, expressing only the full-length hERG1 protein on the plasma membrane. The ␤ 1 integrin and hERG1 coprecipitate in these cells and colocalize in both intracellular and surface membrane compartments. The two proteins also coprecipitate with caveolin-1, suggesting the localization of the complex in lipid rafts/caveolae. hERG1-transfected cells undergo an activation of hERG currents after ␤ 1 integrinmediated adhesion to fibronectin; concomitant with this activation, the focal adhesion kinase associates with the hERG1 protein and becomes tyrosine phosphorylated. Using hERG1-specific inhibitors, we show that the tyrosine phosphorylation of focal adhesion kinase is strictly dependent on hERG channel activity. Similarly, the activity of the small GTPase Rac1 turned out to be dependent on hERG currents. On the whole, these data indicate that the hERG1 protein associates with ␤ 1 integrins and modulates adhesion receptor signaling.

1. The relationships between the K+ inward rectifier current present in neuroblastoma cells (IIR)