title: Leukemia inhibitory factor regulates trafficking of T-type Ca2+ channels.
abstract: Neuropoietic cytokines such as ciliary neurotrophic factor (CNTF) and leukemia inhibitory factor (LIF) stimulate the functional expression of T-type Ca(2+) channels in developing sensory neurons. However, the molecular and cellular mechanisms involved in the cytokine-evoked membrane expression of T-type Ca(2+) channels are not fully understood. In this study we investigated the role of LIF in promoting the trafficking of T-type Ca(2+) channels in a heterologous expression system. Our results demonstrate that transfection of HEK-293 cells with the rat green fluorescent protein (GFP)-tagged T-type Ca(2+) channel α(1H)-subunit resulted in the generation of transient Ca(2+) currents. Overnight treatment of α(1H)-GFP-transfected cells with LIF caused a significant increase in the functional expression of T-type Ca(2+) channels as indicated by changes in current density. LIF also evoked a significant increase in membrane fluorescence compared with untreated cells. Disruption of the Golgi apparatus with brefeldin A inhibited the stimulatory effect of LIF, indicating that protein trafficking regulates the functional expression of T-type Ca(2+) channels. Trafficking of α(1H)-GFP was also disrupted by cotransfection of HEK-293 cells with the dominant-negative form of ADP-ribosylation factor (ARF)1 but not ARF6, suggesting that ARF1 regulates the LIF-evoked membrane trafficking of α(1H)-GFP subunits. Trafficking of T-type Ca(2+) channels required transient activation of the JAK and ERK signaling pathways since stimulation of HEK-293 cells with LIF evoked a considerable increase in the phosphorylation of the downstream JAK targets STAT3 and ERK. Pretreatment of HEK-293 cells with the JAK inhibitor P6 or the ERK inhibitor U0126 blocked ERK phosphorylation. Both P6 and U0126 also inhibited the stimulatory effect of LIF on T-type Ca(2+) channel expression. These findings demonstrate that cytokines like LIF promote the trafficking of T-type Ca(2+) channels.
authors.affiliation: Department of Biology, University of Vermont, Burlington, Vermont, USA.
authors.forename: Deblina
authors.initials: D
authors.lastname: Dey
authors.forename: Andrew
authors.initials: A
authors.lastname: Shepherd
authors.forename: Judith
authors.initials: J
authors.lastname: Pachuau
authors.forename: Miguel
authors.initials: M
authors.lastname: Martin-Caraballo
mesh_terms: D000818:Animals; D020747:Calcium Channels, T-Type / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D002642:Chick Embryo; D004789:Enzyme Activation / Q000235:genetics; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D053605:Leukemia Inhibitory Factor / Q000502:physiology*; D020935:MAP Kinase Signaling System / Q000502:physiology; D021381:Protein Transport / Q000235:genetics / Q000502:physiology; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology
chemical_list: C479952:CACNA1H protein, human; D020747:Calcium Channels, T-Type; D053605:Leukemia Inhibitory Factor; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D048049:Extracellular Signal-Regulated MAP Kinases