A Ca2+ transport system associated with the plasma membrane of Dictyostelium discoideum is activated by different chemoattractant receptors.

Abstract

Abstract. Amebae of Dictyostelium exhibit a transient uptake of extracellular Ca 2+ rv5 s after activation of surface folate or cAMP receptors (Bumann, J., B. Wurster, and D. Malchow. 1984. J. Cell Biol. 98:173-178). To further characterize these Ca 2+ entry systems, we analyzed 45Ca2+ uptake by resting and activated amebae. Like the surface chemoreceptors, folate-and cAMP-induced Ca 2+ uptake responses were developmentally regulated; the former response was evident in vegetative but not aggregation-competent cells, whereas the latter response displayed the opposite pattern of expression. In contrast, other characteristics of these Ca2+-uptake pathways were remarkably similar. Both systems (a) exhibited comparable kinetic properties, (b) displayed a high specificity for Ca 2+, and (c) were inhibited effectively by Ruthenium Red, sodium azide, and carbonylcyanide m-chlorophenylhydrazone. These results, together with the finding that vegetative cells transformed with a plasmid expressing the surface cAMP receptor exhibit a cAMP-induced Ca 2+ uptake, suggest that different chemoreceptors activate a single Ca 2+ entry pathway. Additional pharmacological and ion competition studies indicated that receptor-mediated Ca 2+ entry probably does not involve a Na+/Ca 2+ exchanger or voltageactivated channels. Chemoattractant binding appears to generate intracellular signals that induce activation and adaption of the Ca2+-uptake response. Analysis of putative signaling mutants suggests that Ca 2÷ entry is not regulated by the guanine nucleotide-binding (G) protein subunits God or Got2, or by G protein-mediated changes in intracellular cAMP or guanosine 3,'5'-cyclic monophosphate (cGMP).I N many higher eukaryotic cells, hormones and neurotransmitters induce changes in cytosolic free Ca 2+ by promoting mobilization of sequestered intracellular Ca 2+ and/or by stimulating entry of extracellular Ca 2+ across the plasmalemma. Ca 2+ mobilization responses can be mediated by increases in cytosolic Ca 2+ (18) or by agonistinduced production of inositol 1,4,5-trisphosphate, which releases Ca 2+ from the ER (4) and other cellular compartments (27, 48). Enhanced uptake of extracellular Ca 2+ occurs following activation of voltage-regulated (40) or ligandgated (3) Ca 2+ channels, and likely serves a crucial role both in Ca 2+ signaling events and in replenishing agonist-depleted stores (for review, see reference 22). Importantly, many of these Ca2+-uptake systems, both voltage-dependent and voltage-independent, appear to be regulated by extracellular factors (for review, see reference 43). In addition, certain of these Ca 2+ channels appear to be modulated directly by guanine nucleotide-binding protein (G protein) t subunits while others are regulated indirectly by second messengers such as 1. Abbreviations used in this paper: CCCP, carbonylcyanide m-chlorophenylhydrazone; G protein, guanine nucleotide-binding protein.cAMP, guanosine 3',5'-cyclic monophosphate (cGMP), and inositol polyphosphates (43).In the lower eukaryote, Dictyostel...

A G-protein beta-subunit is essential for Dictyostelium development.

“…For instance, although these cells do not sense cAMP, they are able to carry out chemotaxis to other attractants such as folic acid, yeast, and bacteria extracts (Kesbeke et al 1988). cAMP receptor-mediated Ca 2+ influxes and ligand-induced receptor phosphorylation also occur normally in the g~2-cell lines (Milne and Coukell 1991;Pupillo et al 1992;Milne and Devreotes 1993}. None of these responses are affected by disruption of seven of the eight ~-subunits (G~6 has not been disrupted) (Kumagai et al 1991;Hadwiger and Firtel 1992;M.…”

A model for cAMP-mediated cGMP response in Dictyostelium discoideum.

“…Activation of the receptor alters both the Vm,a and the Km of the transport. The values of these constants have been measured (Millne and Coukell, 1991) and are presented in Table 1.…”

The IplA Ca++ Channel OfDictyostelium discoideumIs Necessary For Ca++, But Not cAMP Chemotaxis, And Plays A Fundamental Role In Natural Aggregation

“…Together, these results indicate that IplA is localized primarily in vesicular membranes in the cytoplasm and suggest low levels in the plasma membrane. (Wick et al, 1978;Milne and Coukell, 1991;Milne and Devreotes, 1993;Groner and Malchow, 1996 (Fig. 1B).…”