Cell division in the unicellular microalga Dunaliella viridis depends on phosphorylation of extracellular signal-regulated kinases (ERKs)

Abstract

In mammalian cells, MAPKs are involved in both stress response (JNK and p38 pathways) and cell proliferation and differentiation [extracellular signal-regulated kinase (ERK)] through protein kinase cascades. Exposure of Dunaliella viridis cell cultures to PD98059, a very specific inhibitor of the ERK signalling pathway, resulted in a total arrest of cell proliferation and a complete dephosphorylation of ERK. As shown by flow cytometry analysis of propidium iodide-stained cells, PD98059 stopped mitosis at the G(2) phase after the S phase has been completed. Multiple physiological parameters such as cell motility and reducing power generation (NADPH) clearly indicate that the treated cells are wholly viable. Exposure of D. viridis to environmental stresses that impair cell division, such as hyperosmotic shock, nitrogen starvation, or sublethal UV irradiation, caused a marked decrease in the phospho-ERK levels as detected by western blot. Two 400 bp polynucleotides from D. viridis with high homologies to published sequences of ERK1 and ERK2 were cloned, sequenced, and submitted to GenBank. Northern blot analysis revealed two mRNA bands of approximately 1.9 kb, consistent with the expected size of ERK proteins ( approximately 40 kDa). Sequence analysis showed that they contained several mitogen-activated protein kinase (MAPK) conserved domains, including II, III, VIb, VII, and the double phosphorylation motif. Interestingly, in D. viridis, this motif was T*DY* instead of the canonic T*EY*. Based on this finding, ERK plant sequences can be divided into two groups, one termed the T*DY* branch and the other termed the T*EY* branch. The molecular and functional data presented here suggest that ERK is a very ancient signalling pathway and that it was already present in the last common ancestor of all eukaryotic cells.

Phosphorylation of MAPK-like proteins in three intertidal macroalgae under stress conditions

“…This behaviour in HC thalli under temperature stress was similar to that al ready described in Dunaliella tertiolecta after exposure to UVR (García-Gómez et al 2012), where a transient increase (24 h) of p38 phosphorylation was detected. In E. elongata, ERK phosphorylation de creased in all C and N combinations after exposure to elevated temperature (+ 4°C), which could indicate that cell division and algal growth decrease compared to initial conditions at ambient temperature (Jiménez et al 2007, García-Gómez et al 2012). In the case of U. rigida, ERK phosphorylation was higher at temperature+ 4°C than at ambient temperature (except in LCLN), which may indicate that this species benefits from increased temperature if C and N are not limiting.…”

“…In addition, presence of ERK-like MAPK was detected in all species. Active cell division occurs only when ERK is phosphorylated, and its dephos phorylation leads to cell division arrest, followed by cell death (Jiménez et al 2007).…”

“…Despite the fact that the study of signal transduction in algae is very recent, different studies with both macro-and microalgae (Jiménez et al 2004, 2007, García-Gómez et al 2012 have clearly shown the presence of p38-like, JNK-like and ERK-like components in algae, as well as their crucial role in acclimation to stress and in cell division (Jiménez et al 2004, 2007, Parages 2012. A procedure has been developed to detect the presence of these MAPK homologues in several species of Chlorophyta, Rhodophyta and Phaeophyta , revealing that phosphory lation of p38-and JNK-like MAPKs occurs in Arctic kelps in response to increasing UVR exposure and temperature (Parages et al 2013) as well as in intertidal macroalgae at the onset of emersion and during periods of the highest irradiation (Parages et al 2014).…”

Metabolic switching: synergistic induction of carotenogenesis in the aerial microalga, Vischeria helvetica, under environmental stress conditions by inhibitors of fatty acid biosynthesis

“…In the green microalga D. tertiolecta, two kinds of MAPK-like proteins, p38 and c-jun Nterminal kinase (JNK), are phosphorylated during environmental stresses including osmotic and thermal shock, ultraviolet irradiation, and nitrogen starvation (Jiménez et al 2004). The phosphorylation levels of extracellular signal-regulated kinase (ERK)-like proteins in D. viridis decrease markedly after exposure to stresses such as hyperosmotic shock, sublethal ultraviolet irradiation, and nitrogen deficiency that impaired cell division (Jiménez et al 2007).…”

“…6. JNK-like proteins are associated with cell survival, cell death, DNA repair, and metabolism in D. tertiolecta (Jiménez et al 2007;García-Gómez et al 2012). Although the influence of the JNK pathway on carotenoid biosynthesis in microalgae is not well understood, it may play an important role in regulating carotenogenesis.…”