Open AccessLeukemia inhibitory factor regulates trafficking of T-type Ca2+channels
Author(s) -
Deblina Dey,
Andrew K. Shepherd,
Judith Pachuau,
Miguel MartinCaraballo
Publication year - 2011
Publication title -
american journal of physiology. cell physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.432
H-Index - 181
eISSN - 1522-1563
pISSN - 0363-6143
DOI - 10.1152/ajpcell.00115.2010
Subject(s) - leukemia inhibitory factor , microbiology and biotechnology , hek 293 cells , brefeldin a , transfection , chemistry , green fluorescent protein , mapk/erk pathway , ciliary neurotrophic factor , signal transduction , biology , cell culture , endoplasmic reticulum , neurotrophic factors , golgi apparatus , biochemistry , embryonic stem cell , receptor , genetics , gene
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.