Phosphorylation of a Renatured Protein from Etiolated Wheat Leaf Protoplasts Is Modulated by Blue and Red Light

Abstract

The central role played by light in the coordination of plant growth and development has led to intense interest in the detection and transduction of light signals (Chory, 1993). The primary receptor of R is phytochrome (Hart, 1988) and multiple forms are known to exist, which may control different cellular processes. Increasing evidence suggests that one receptor for B may be a protein kinase (Gallagher et al., 1988). Since both R and B initiate separate but often interrelated physiological responses, it has been assumed that the transduction pathways may be distinct, but the identity of either remains unclear (Trewavas and Gilroy, 1991).It has long been thought that [Ca2+i] may act in transduction of photomorphogenetic signals resulting from R due to its important role in animal cells (Haupt and Weisenseel, 1976). Early studies involving 45Ca2+ flux measurements and autoradiography (Weisenseel and Ruppert, 1977; Dreyer and Weisenseel, 1979; Hale and Roux, 1980) provided indirect evidence to support this hypothesis. Following the detection of a plasma membrane-located Ca-dependent protein kinase in plant cells (Hetherington and Trewavas, 1982), Roux (1984) Grimm et al. (1989), who showed that phytochrome often co-purifies with a 60-kD Ca-dependent protein kinase. Thummler et al. (1992) demonstrated that Ceratodon phytochrome, unlike other plant phytochromes, has a conserved protein kinase sequence.When etiolated wheat (Tviticum aestivum L.) leaf protoplasts are irradiated with R, their volume increases by up to 20% (Bossen et al., 1988). It is believed that R-induced protoplast swelling employs the same molecular mechanism as R-induced leaf unrolling (Zhou et al., 1990). Using Casensitive fluorescent dyes and laser confocal scanning imaging, we have been able to show that R induces [Ca2+], transients lasting several minutes in single etiolated wheat leaf protoplasts (Shacklock et al., 1992). We mimicked these transients by photolysis of incorporated caged Ca (nitr-5) and incorporated caged inositol trisphosphate and showed that these artificially produced transients increased the volume of the protoplasts, thus mimicking the physiological effects of R. By incubating etiolated wheat leaf protoplasts in 32Pi for time periods between 15 s and 15 min, we detectedR-induced changes in the phosphorylation of two peptides with approximate electrophoretic mobilities of 60 and 70 kD (Fallon et al., 1993). Phosphorylation of these two peptides increased during 45 s of R irradiation and showed kinetics similar to those of the detected [Ca2+], transients. Again, we were able to mimic the R-increased phosphorylation of these two peptides by photolyzing incorporated caged Ca.The 60-and 70-kD peptides were the primary species labeled in these short incubation periods; only after several hours of incubation in 32Pi was the labeling of numerous other peptides detected. This suggested the possibility that the very rapid labeling of the 60-and 70-kD peptides was the result of autophosphorylation, which is often faster t...

Light activates a 46‐kDa MAP kinase‐like protein kinase in soybean cell culture

“…Protein phosphorylation and dephosphorylation events are correlated with several light responses in a number of plants [19], and photoreceptor‐mediated phosphorylation of a number of mostly unidentified proteins has been demonstrated [20,21]. Recent reports have suggested light‐dependent phosphorylation of several transcription factors that interact with promoter elements of light‐regulated genes.…”

An increase in cytosolic calcium ion concentration precedes hypoosmotic shock‐induced activation of protein kinases in tobacco suspension culture cells

“…In plants, it has been reported that phytohormones and external stimuli, such as light, cold shock, fungal elicitor and wounding, induce the phosphorylation of specific proteins and activate protein kinases [1][2][3][4][5][6][7]. Transient elevation of cytosolic free calcium concentration ([Ca 2+ ] cyt ) has been reported to be caused by external stimuli such as cold shock, fungal elicitor and wounding as well [8,9].…”

Light perception in higher plants

“…Signal transduction in pulvinar movement and guard cell swelling, processes which are regulated by blue light, is beyond the scope of this review, and I refer the reader to recent reviews on this topic [23,147]. Phytochromemediated phosphorylation of a number of mostly unidentified proteins has been demonstrated [148][149][150][151]. Phosphorylation and translocation between the cytosol and the nucleus of the bZIP transcription factor CPRF2 from parsley [152][153][154] and import of the bZIP factor GBF2 from Arabidopsis into the nucleus [155] is also light-regulated.…”