Two distinct intracellular Ca2+-release components act in opposite ways in the regulation of the auxin-dependent MIA biosynthesis in Catharanthus roseus cells

Abstract

Calcium-mediated signalling is ubiquitous in both animals and plants. Changes in cytoplasmic free Ca(2+) concentration couple diverse arrays of stimuli to their specific responses, the specificity of the stimulus being determined by integrated actions between multiple Ca(2+) mobilization pathways. In this work, a pharmacological approach is reported, aimed at deciphering the role of calcium as a second messenger in the transduction pathway leading to the inhibitory effect of 2,4-dichlorophenoxyacetic acid (2,4-D), in regulating monoterpene indole alkaloid (MIA) biosynthesis in Catharanthus roseus cells. It is demonstrated here that auxin-dependent MIA biosynthesis is differentially regulated by two distinct calcium release components from internal stores in C. roseus showing pharmacological profiles similar to those displayed by animal RyR and IP3 channels. MIA biosynthesis is stimulated by caffeine (Ca(2+)-release activator through RyR channels) and by heparin and TMB8 (Ca(2+)-release inhibitors of IP3 channels) whereas MIA biosynthesis is inhibited by mastoparan (Ca(2+)-release activator of IP3 channels) and by ruthenium red and DHBP (Ca(2+)-release inhibitors of RyR channels). Furthermore, calcium, as 2,4-D, acts on MIA biosynthesis by regulating the monoterpene moiety of the MIA biosynthesis pathway since calcium channel modulators preferentially modulate g10h expression, the gene encoding the enzyme of the secoiridoid monoterpene pathway, that is the major target of 2,4-D action. In addition, the simultaneous use of caffeine (an activator of RyR channel in animals) and TMB8 (an inhibitor of the IP3 channel) in 2,4-D treated cells triggers a synergistic effect on MIA accumulation. This finding suggests an opposite and co-ordinated action of multiple Ca(2+)-release pathways in 2,4-D signal transduction, adding a new level of complexity to calcium signalling in plants and questioning the existence of RyR and IP3 channels in plants.

Molecular Mechanism for Human Sperm Chemotaxis Mediated by Progesterone

“…However, these channels can be effectively blocked by tetracaine [47], which did not interfere with P-mediated chemotactic response (see below), thus it seems that CNG may not be responsible for Ca 2+ influx during chemotaxis mediated by P. In contrast, the SOC channel blocker SKF96365 inhibited human sperm chemotaxis at a doses consistent with its effects on SOCs, similarly to the reported block of chemotaxis in ascidian sperm [48]. Furthermore, the antagonist TMB-8 (known to inhibit both IP 3 R [20], [22] and RyR [21] calcium channels) significantly inhibited chemotaxis at 10 µM, a dose consistent with effects on store mobilization and modest compared to that reported to arrest intracellular calcium oscillations in human sperm stimulated by micromolar P [18]. Thus, it appears likely that store mobilization and activation of SOCs occurs during P-induced chemotaxis of human sperm and underlies at least part of the requirement for Ca 2+ influx.…”

“…Cells were pre-treated with TMB-8, which inhibits stored Ca 2+ mobilization in somatic cells [20]–[22]. This compound caused a clear, dose-dependent inhibition of chemotaxis that was statistically significant at 10 µM (Fig.…”

Deciphering the Evolution, Cell Biology and Regulation of Monoterpene Indole Alkaloids

“…Because auxin inhibits MIA biosynthesis in cell culture, predominantly at secologanin biosynthesis level, downstream signalling has been investigated (Poutrain, Mazars, Thiersault, Rideau, & Pichon, 2009). Changes in free cytoplasmic calcium concentration ([Ca 2þ ] cyt ) occur during many physiological processes and particularly in auxin signalling (Singla, Chugh, Khurana, & Khurana, 2006).…”

Ca2+signals: The versatile decoders of environmental cues

“…There is differential regulation of auxin-dependent MIA biosynthesis by functioning of two distinct calcium release components showing pharmacological profiles similar to those displayed by animal RyR and IP3 channels. The existence of RyR and IP3 channels in plants make calcium signaling to be a more complex mechanism (Poutrain et al, 2009).…”