In vivo phosphorylation sites of the tobacco calcium-dependent protein kinases NtCDPK2 and NtCDPK3 were determined in response to biotic or abiotic stress. Stress-inducible phosphorylation was exclusively located in the variable N termini, where both kinases were phosphorylated differentially despite 91% overall sequence identity. In NtCDPK2, serine 40 and threonine 65 were phosphorylated within 2 min after stress. Whereas Thr 65 is subjected to intra-molecular in vivo autophosphorylation, Ser 40 represents a target for a regulatory upstream protein kinase, and correct NtCDPK2 membrane localization is required for Ser 40 phosphorylation. NtCDPK3 is phosphorylated at least at two sites in the N terminus by upstream kinase(s) upon stress stimulus, first at Ser 54 , a site not present in NtCDPK2, and also at a second undetermined site not identical to Ser 40 . Domain swap experiments established that differential phosphorylation of both kinases is exclusively determined by the respective N termini. A cell death-inducing response was only observed upon expression of a truncated variant lacking the junction and calcium-binding domain of NtCDPK2 (VK2). This response required protein kinase activity and was reduced when subcellular membrane localization was disturbed by a mutation in the myristoylation and palmitoylation site. Our data indicate that CDPKs are integrated in stressdependent protein kinase signaling cascades, and regulation of CDPK function in response to in vivo stimulation is dependent on its membrane localization. Calcium-dependent protein kinases (CDPKs)2 form a gene family of 34 and 29 members in Arabidopsis thaliana and rice, respectively. They possess a conserved modular structure of four domains (VKJC): an N-terminal variable domain (V), which for many isoforms contains a myristoylation and palmitoylation motif mediating membrane localization; a serine/ threonine protein kinase domain (K); an autoinhibitory junction peptide (J); and a calcium-sensing calmodulin-like domain (C) at the C terminus (1). These structural features as well as published biochemical and functional data suggest that the regulatory switch for CDPK in vivo activation includes (i) the binding of calcium leading to the release of the autoinhibitory domain from the active site, (ii) CDPK protein (auto)phosphorylation, and (iii) specific subcellular localization (2-6).CDPKs have been associated with the regulation of diverse processes of plant growth and development, nutrient primary metabolism, and signaling in biotic and abiotic stress responses and water/ion transport (7-9). Only recently, the analysis of genetic knock-out mutants in Arabidopsis allowed the assignment of a biological function in abscisic acid signaling, stomatal aperture,andsaltstresstolerancetoArabidopsisCPK4(calciumdependent protein kinase 4) and CPK11 (10), CPK3 and CPK6 (11), and CPK23 (12), respectively. However, none of these studies addressed the biochemical mechanism of how the CDPK enzyme itself is regulated by (auto)phosphorylation in planta.In vitro...

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Soil test crop response (STCR) correlation studies were carried out in Vindhyan alluvial plain during 2001 to 2004 taking IR-36 as test crop to quantify rice production in the context of the variability of soil properties and use of balanced fertilizers based on targeted yield concept. The soils were developed on gently sloping alluvial plain with different physiographic settings and notable variation in drainage condition. Soil properties show moderate variation in texture (loamy to clay), organic carbon content (4.4 to 9.8 g/kg), cation exchange capacity (10.2 to 22.4 cmol (p+)/kg) and pH (5.3 to 6.4). Soil fertility status for N is low to medium (224 to 348 kg/ha), P is medium to high (87 to 320 kg/ha) and K ranges from medium to high (158 to 678 kg/ha). Database regarding nutrient requirement in kg/t of grain produce (NR), the percent contribution from the soil available nutrients [CS (%)] and the percent contribution from the applied fertilizer nutrients [CF (%)] were computed for calibrating and formulating fertilizer recommendations. Validity of the yield target for 7 and 8 t/ha was tested in farmers' fields and yields targets varied at less than 10%. The percent achievement of targets aimed at different level was more than 90%, indicating soil test based fertilizer recommendation approach was economically viable within the agro-ecological zone with relatively uniform cropping practices and socio-economic conditions.

GnCDPK specifically uses the myosin light chain synthetic peptide (MLCpep), which is the phosphate-accepting domain of smooth muscle myosin light chains (KKRPQRATSNVFS), as an exogenous substrate under in vitro experimental conditions. In this report we show that GnCDPK undergoes intramolecular autophosphorylation. This self-phosphorylation occurs in threonine residues in a Ca 2؉ -dependent (K 0.5 ‫؍‬ 0.5 M) and calmodulin-independent manner. The kinase activity toward MLCpep and its sensitivity to Ca 2؉ were unaffected by prior autophosphorylation when measured under saturating ATP concentrations. The role of autophosphorylation in the exogenous substrate MLCpep phosphorylation reaction was reinvestigated at low ATP concentrations. A pronounced lag time of 1 to 2 min, followed by a linear increase of activity for 7.5 min, was seen in the initial rate of MLCpep phosphorylation under such suboptimal conditions. Prior autophosphorylation completely abolished this lag phase, and a sharp rise of exogenous substrate phosphorylation was seen from the 1st min. Our results suggest that autophosphorylation is a prerequisite for the activation of GnCDPK.