Posted on

Spk seeds

Rice SPK, a calmodulin-like domain protein kinase, is required for storage product accumulation during seed development: phosphorylation of sucrose synthase is a possible factor

Suc, an end product of photosynthesis, is metabolized by Suc synthase in sink organs as an initial step in the biosynthesis of storage products. Suc synthase activity is known to be regulated by reversible phosphorylation, but the details of this process are unclear at present. Rice SPK, a calcium-dependent protein kinase, is expressed uniquely in the endosperm of immature seed, and its involvement in the biosynthetic pathways of storage products was suggested. Antisense SPK transformants lacked the ability to accumulate storage products such as starch, but produced watery seed with a large amount of Suc instead, as the result of an inhibition of Suc degradation. Analysis of in vitro phosphorylation indicated that SPK phosphorylated specifically a Ser residue in Suc synthase that has been shown to be important for its activity in the degradation of Suc. This finding suggests that SPK is involved in the activation of Suc synthase. It appears that SPK is a Suc synthase kinase that may be important for supplying substrates for the biosynthesis of storage products.

Figures

Detection of SPK mRNA in…

Detection of SPK mRNA in an Immature Rice Seed by in Situ Hybridization.…

Detection of SPK mRNA in an Immature Rice Seed by in Situ Hybridization. (A) The region expressing Spk is shown in purple. The specific RNA of the antisense strand was used as a probe. (B) and (C) Magnified images corresponding to the boxes in (A). (D) and (E) Results of the control experiment using sense strand RNA as a probe. AL, aleurone layer; EM, embryo; EN, endosperm.

Effect of the Antisense Spk…

Effect of the Antisense Spk Gene on Its Transformants. Phenotypic features of immature…

Effect of the Antisense Spk Gene on Its Transformants. Phenotypic features of immature seed 2 weeks after pollination of the wild type (cv Nipponbare) (A) and of a representative antisense transformant (ASPK19) (B) are shown. HU, hull; EN, endosperm; EN′, watery endosperm.

Protein Accumulation in Immature Seed…

Protein Accumulation in Immature Seed of the Transformants and Wild-Type Plants. (A) Protein…

Protein Accumulation in Immature Seed of the Transformants and Wild-Type Plants. (A) Protein accumulated in immature seed of the transformant ASPK19 and the wild type detected by SDS-PAGE. Lane 1, crude extract; lane 2, soluble fraction; lane 3, insoluble fraction. Each lane contained one-fifth of a total crude extract or of each fraction that was prepared independently from a single grain. (B) Protein composition of immature seed of the transformant ASPK19 or the wild type. Each lane contained 25 μg of total protein from the crude extract. (C) Detection of RBE1 (85 kD) and SPK (68 kD) in immature seed of the transformant ASPK19 or the wild type by protein gel blot analysis. Each lane contained one-fifth of a total crude extract from a single grain. The proteins corresponding to RBE1 and SPK are indicated by arrows. TF, transformant ASPK19; WT, wild type.

See also  X-wing seeds

SPK May Have CDPK Activity.…

SPK May Have CDPK Activity. (A) Calcium dependence of SPK autophosphorylation. The effects…

SPK May Have CDPK Activity. (A) Calcium dependence of SPK autophosphorylation. The effects of the addition of 100 μM Ca 2+ (as CaCl2) or 100 μM EGTA to the reaction (+) were examined. (B) Effects of inhibitors. Results in the presence of 1 μM (lane 1), 5 μM (lane 2), or 10 μM (lane 3) staurosporine (STA), ML-9, W-7, or with no inhibitor (CON) are shown. Each lane contained 3.0 μg of GST–SPK in the reaction mixture.

In Vitro Phosphorylation of GST–Suc…

In Vitro Phosphorylation of GST–Suc Synthase by SPK. In vitro phosphorylation of GST–Suc…

In Vitro Phosphorylation of GST–Suc Synthase by SPK. In vitro phosphorylation of GST–Suc synthase (Susy) and RBE1 (RBE) in the presence or absence of GST–SPK (SPK). The presence of these proteins in the reaction (2.5 μg of GST–SPK, 3.5 μg of GST–Suc synthase, and 3.0 μg of RBE1) is indicated (+). Top, Coomassie blue–stained proteins (CBB); bottom, corresponding autoradiograms (AR).

Detection of in Vitro Phosphorylation…

Detection of in Vitro Phosphorylation of Proteins by SPK in Immature Seed. Proteins…

Detection of in Vitro Phosphorylation of Proteins by SPK in Immature Seed. Proteins on a 10% SDS–polyacrylamide gel are shown as an autoradiogram. Lanes 1 and 2 contained crude extracts from the wild-type plant (cv Nipponbare). Lane 2 shows the results of kinase reactions in the presence of 100 μM Ca 2+ .

Immunoprecipitation of Suc Synthase in…

Immunoprecipitation of Suc Synthase in Immature Seed and Phosphorylation by SPK. Phosphorylation of…

Immunoprecipitation of Suc Synthase in Immature Seed and Phosphorylation by SPK. Phosphorylation of the constituents of immunoprecipitates obtained with anti-Suc synthase antibody is shown. Lanes show the results of reactions in the presence (+) or absence of a crude extract of immature seed (C.E.), anti-Suc synthase antibody (anti-Susy), and GST–SPK. CBB indicates Coomassie blue–stained proteins, and AR shows the corresponding autoradiograms.

In Vitro Phosphorylation of GST–Suc…

In Vitro Phosphorylation of GST–Suc Synthase by the Crude Extract from the Immature…

In Vitro Phosphorylation of GST–Suc Synthase by the Crude Extract from the Immature Seed of the Wild-Type Plant and the Watery Seed of the Antisense Transformants. Each lane contained 3.0 μg of GST–Suc synthase as the substrate. Lanes 1 and 2 contained 8.0 μg of total proteins in the crude extract from the immature seed of cv Nipponbare. Lane 2 also included 1 mM EGTA in the reaction mixture. Lane 3 contained 8.0 μg of total proteins in the crude extract from the watery seed of the antisense transformant ASPK19. Proteins resolved on a 10% SDS–polyacrylamide gel are shown in the autoradiogram.

See also  Fruity juice seeds

Detection of Suc Synthase in…

Detection of Suc Synthase in the Watery Seed of the Antisense Transformant by…

Detection of Suc Synthase in the Watery Seed of the Antisense Transformant by Protein Gel Blot Analysis. Each lane contained one-fifth of a total crude extract from a single grain of the antisense SPK transformant line ASPK19 (TF) and the wild-type plant (WT). The 85-kD protein corresponding to Suc synthase (Susy) is indicated by the arrow.

Determination of the Phosphorylation Site…

Determination of the Phosphorylation Site in Suc Synthase. (A) Alignment of the amino…

Determination of the Phosphorylation Site in Suc Synthase. (A) Alignment of the amino acid sequences of Suc synthases in the region around the potential target site. The consensus amino acid residues, Arg and Ser, are boxed. Zm1, Zm2, Rs1, Rs2, and Rs3 denote Suc synthases encoded by maize Msus1, maize Msus2, rice Rsus1, rice Rsus2, and rice Rsus3, respectively. RtoG, mutant with substitution at the Arg residue; StoV, mutant with substitution at the Ser residue. Numbers indicate the positions of amino acid residues around the potential phosphorylation site. For accession numbers, see Methods. (B) In vitro phosphorylation of the site-specifically mutated and wild-type Suc synthases. Lanes show the reaction with GST–Suc synthase (wild type [WT]), with GST–RtoG (RtoG), and with GST–StoV (StoV). CBB and AR indicate the Coomassie blue–stained proteins and the corresponding autoradiograms, respectively.

RNA maturation of the rice SPK gene may involve trans-splicing

A gene encoding a calcium-dependent seed-specific protein kinase (SPK) is abundantly expressed in developing rice seeds (Kawasaki, T et al. Gene (1993) 129, 183-189). Rice genomic clones encoding SPK were isolated using the entire cDNA fragment as a probe. Physical mapping of these genomic clones indicated that the genomic region corresponding to the entire cDNA was divided into two different regions, SPK-A and SPK-B, located on different rice chromosomes. The results of RACE-PCR analyses showed that the respective transcripts from SPK-A and SPK-B contained additional sequences which were not found in the SPK cDNA, and that these sequences were removed like introns during maturation of the SPK mRNA. These results suggest that two different RNAs were independently transcribed from SPK-A and SPK-B and joined, possibly by trans-splicing.

Similar articles

Kawasaki T, Hayashida N, Baba T, Shinozaki K, Shimada H. Kawasaki T, et al. Gene. 1993 Jul 30;129(2):183-9. doi: 10.1016/0378-1119(93)90267-7. Gene. 1993. PMID: 8325505

Breviario D, Morello L, Giani S. Breviario D, et al. Plant Mol Biol. 1995 Mar;27(5):953-67. doi: 10.1007/BF00037023. Plant Mol Biol. 1995. PMID: 7766885

Asano T, Kunieda N, Omura Y, Ibe H, Kawasaki T, Takano M, Sato M, Furuhashi H, Mujin T, Takaiwa F, Wu Cy CY, Tada Y, Satozawa T, Sakamoto M, Shimada H. Asano T, et al. Plant Cell. 2002 Mar;14(3):619-28. doi: 10.1105/tpc.010454. Plant Cell. 2002. PMID: 11910009 Free PMC article.

See also  Patty cake seeds

Kikuchi H, Hirose S, Toki S, Akama K, Takaiwa F. Kikuchi H, et al. Plant Mol Biol. 1999 Jan;39(1):149-59. doi: 10.1023/a:1006156214716. Plant Mol Biol. 1999. PMID: 10080717

Washio K, Ishikawa K. Washio K, et al. Plant Mol Biol. 1992 Jul;19(4):631-40. doi: 10.1007/BF00026789. Plant Mol Biol. 1992. PMID: 1627776

Cited by 9 articles

Halawa M, Cortleven A, Schmülling T, Heyl A. Halawa M, et al. Sci Rep. 2021 Jan 18;11(1):1722. doi: 10.1038/s41598-020-80223-2. Sci Rep. 2021. PMID: 33462253 Free PMC article.

Singh A, Zahra S, Das D, Kumar S. Singh A, et al. Database (Oxford). 2019 Jan 1;2019:bay135. doi: 10.1093/database/bay135. Database (Oxford). 2019. PMID: 30624648 Free PMC article.

Dubrovina AS, Kiselev KV, Zhuravlev YN. Dubrovina AS, et al. Biomed Res Int. 2013;2013:264314. doi: 10.1155/2013/264314. Epub 2012 Dec 26. Biomed Res Int. 2013. PMID: 23509698 Free PMC article. Review.

Ülker B, Hommelsheim CM, Berson T, Thomas S, Chandrasekar B, Olcay AC, Berendzen KW, Frantzeskakis L. Ülker B, et al. Plant Cell. 2012 Nov;24(11):4314-23. doi: 10.1105/tpc.112.100404. Epub 2012 Nov 9. Plant Cell. 2012. PMID: 23144181 Free PMC article. Review.

Reddy AS, Rogers MF, Richardson DN, Hamilton M, Ben-Hur A. Reddy AS, et al. Front Plant Sci. 2012 Feb 7;3:18. doi: 10.3389/fpls.2012.00018. eCollection 2012. Front Plant Sci. 2012. PMID: 22645572 Free PMC article.

Sour Patch Kiss

This strain has it all – good yield, insane amounts of gooey stickiness and a delicious, loud smell of sour candy. Every grower that we have shown the dried flowers to immediately said they had to have the strain ASAP. Our goal with this strain was to create bag appeal that was second to none that gave an experience to match.

The parents we chose for this are Kimbo Kush and Sour Kush. When I first tried Kimbo Kush, I knew I wanted to work on a project with Kimbo as one of the ingredients. Not only was it incredibly frosty and potent with an amazing smell/taste of Blackberry scones, it created a very distinguishable euphoric, indica-dominant high. I chose Sour Kush as the other parent because it was a strain that would totally stink up a room with a sour funk, was a great yielder and super frosty/potent.

Sour Patch Kiss grows extremely impressive flowers. The buds look like they are dripping in resin. With the proper environment, trichomes will coat even the fan leaves and stems.

The above picture was taken at 6 weeks into flower. It continued to pack on resin all the way up to harvest until the top colas looked like they were wearing a white fur coat.

The looks, smell and taste of Sour Patch Kiss are at a level very few strains can come close to achieving.