On cell signalling mechanism of Mycobacterium leprae soluble antigen (MLSA) in Jurkat T cells.

We investigated the role of Mycobaterium leprae soluble antigen (MLSA) in the modulation of calcium signalling, phosphorylation of mitogen-activated protein (MAP) kinases and IL-2 mRNA expression in human Jurkat T cells. We observed that MLSA induced an increase in free intracellular calcium concentrations, [Ca2+]i, via opening CRAC (Ca2+-release activated- Ca2+) channels. Furthermore, MLSA failed to potentiate both thapsigargin- and anti-CD3 antibodies-induced capacitative calcium influx in Jurkat T cells. We observed that MLSA failed to affect the degree of phosphorylation of two MAP kinases, i.e., ERK1/ERK2, stimulated by anti-CD3 antibodies alone or phorbol 12-myristate 13-acetate (PMA) alone. In order to mimic co-stimulation of T cells, we stimulated them by both PMA and anti-CD3 antibodies. MLSA significantly curtailed the phosphorylation of ERK1/ERK2, stimulated by both PMA and anti-CD3 antibodies in Jurkat T cells. Also MLSA was found to reduce the transcription of IL-2 gene in PMA plus anti-CD3 antibodies-activated Jurkat T cells. Our finding demonstrates that Ca2+ influx via CRAC channels, inhibition of ERK1/ERK2 phosphorylation and IL-2 gene transcription may be implicated in immunosuppressive effects of MLSA antigen.

Fatty acyl-CoA-acyl-CoA-binding protein complexes activate the Ca2+ release channel of skeletal muscle sarcoplasmic reticulum.

We previously reported that fatty acyl-CoA esters activate ryanodine receptor/Ca2+ release channels in a terminal cisternae fraction from rabbit skeletal muscle [Fulceri, Nori, Gamberucci, Volpe, Giunti and Benedetti (1994) Cell Calcium 15, 109-116]. Skeletal muscle cytosol contains a high-affinity fatty acyl-CoA-binding protein (ACBP) [Knudsen, Hojrup, Hansen, H.O., Hansen, H.F. and Roepstorff (1989) Biochem. J. 262, 513-519]. We show here that palmitoyl-CoA (PCoA) in a complex with a molar excess of bovine ACBP causes a discrete Ca2+ efflux or allows Ca2+ release from the Ca2+-preloaded terminal cisternae fraction by sub-optimal caffeine concentrations. Both effects were abolished by elevating the free [Mg2+] in the system, which inhibits the Ca2+ release channel activity. Sensitization towards caffeine was a function of both the concentration of the complex and the [PCoA]-to-[ACBP] ratio. In all experimental conditions the calculated free [PCoA] was no more than 50 nM, and such concentrations by themselves were inactive on Ca2+ release channels. The KD for PCoA binding was approx. 2 nM for bovine and yeast ACBP, and slightly higher (8 nM) for rat ACBP. The PCoA-rat ACBP complex behaved in the same manner as the PCoA-bovine ACBP complex, whereas the ester complexed with yeast ACBP was more active in activating/sensitizing Ca2+ efflux. A non-hydrolysable analogue of PCoA bound to (bovine) ACBP also sensitized the Ca2+ release channel towards caffeine. These findings indicate that fatty acyl-CoA-ACBP complexes either interact directly with one or more components in the terminal cisternae membranes or, through interaction with the component(s), donate the fatty acyl-CoA esters to high-affinity binding sites of the membrane, thus affecting (and possibly regulating) Ca2+ release channel activity.