Abstract
Calcium acts as a universal second messenger in both developmental processes and responses to environmental stresses. Previous research has shown that a number of stimuli can induce [Ca2+] increases in both the cytoplasm and nucleus in plants. However, the relationship between cytosolic and nucleosolic calcium signaling remains obscure. Here, we generated transgenic plants containing a fusion protein, comprising rat parvalbumin (PV) with either a nuclear export sequence (PV-NES) or a nuclear localization sequence (NLS-PV), to selectively buffer the cytosolic or nucleosolic calcium. Firstly, we found that the osmotic stress-induced cytosolic [Ca2+] increase (OICIcyt) and the salt stress-induced cytosolic [Ca2+] increase (SICIcyt) were impaired in the PV-NES lines compared with the Arabidopsis wildtype (WT). Similarly, the osmotic stress-induced nucleosolic [Ca2+] increase (OICInuc) and salt stress-induced nucleosolic [Ca2+] increase (SICInuc) were also disrupted in the NLS-PV lines. These results indicate that PV can effectively buffer the increase of [Ca2+] in response to various stimuli in Arabidopsis. However, the OICIcyt and SICIcyt in the NLS-PV plants were similar to those in the WT, and the OICInuc and SICInuc in the PV-NES plants were also same as those in the WT, suggesting that the cytosolic and nucleosolic calcium dynamics are mutually independent. Furthermore, we found that osmotic stress- and salt stress-inhibited root growth was reduced dramatically in the PV-NES and NLS-PV lines, while the osmotic stress-induced increase of the lateral root primordia was higher in the PV-NES plants than either the WT or NLS-PV plants. In addition, several stress-responsive genes, namely CML37, DREB2A, MYB2, RD29A, and RD29B, displayed diverse expression patterns in response to osmotic and salt stress in the PV-NES and NLS-PV lines when compared with the WT. Together, these results imply that the cytosolic and nucleosolic calcium signaling coexist to play the pivotal roles in the growth and development of plants and their responses to environment stresses.
Highlights
Calcium is commonly accepted as a ubiquitous second messenger in eukaryotic organisms in which it regulates diverse biological processes, such as fertilization, pollen tube elongation, proliferation, neural signaling, and learning (Berridge et al, 2000; Bootman et al, 2001; Cullen and Lockyer, 2002)
We found that enhanced yellow fluorescent protein (eYFP)-PV-nuclear export signal (NES) was restricted to the cytoplasm, whereas eYFP-nuclear localized signal (NLS)-PV was colocalized with the nuclear marker (Figure 1A)
These results proved that the subcellular localization of PV-NES or NLS-PV is specific to the cytoplasm or nucleus in the transgenic plants, which lay the foundation for its selective buffering of [Ca2+]cyt or [Ca2+]nuc
Summary
Calcium is commonly accepted as a ubiquitous second messenger in eukaryotic organisms in which it regulates diverse biological processes, such as fertilization, pollen tube elongation, proliferation, neural signaling, and learning (Berridge et al, 2000; Bootman et al, 2001; Cullen and Lockyer, 2002). The Ca2+ signature in plants has two basic patterns: the first is the circadian Ca2+ oscillation that occurs at the whole-tissue level (Johnson et al, 1995), which is mainly regulated by the CASIP3 pathway in Arabidopsis (Tang et al, 2007) and is measured by an aequorin-based calcium indicator (Knight et al, 1991); the second is those short-term Ca2+ increases or spikes which respond to various abiotic and biotic stimuli, namely light, high and low temperatures, touch, salt and drought, osmotic stress, plant hormones, fungal elicitors, and nodulation factors (Sanders et al, 1999), which can be measured by aequorin and fluorescence resonance energy transfer (FRET)-based yellow
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