Dataset Information

29RNP kinase idetification - Identification and characterization of chloroplast casein kinase II from Oryza sativa (rice)

ABSTRACT: The 29-kDa RNA protein (29RNP), which was originally identified in rice etioplasts, has increased phosphorylation levels during de-etiolation. Here, using heparin-sepharose enriched fractions, we identified a chloroplast kinase that phosphorylated 29RNP. The chloroplast kinase phosphorylated the 29RNP and exhibited CKII biochemical characteristics in a kinase reaction test. Proteomic analysis of the chloroplast heparin-sepharose enriched fractions revealed the presence of 70 proteins, including both abundant proteins, such as plastid encoded polymerase subunits, and low-abundance proteins, such as APO2 and DAG. An inclusion list method using Fourier transform ion cyclotron resonance mass spectrometer ( FT-ICR LTQ MS ) was subsequently used to analyze the chloroplast heparin-sepharose enriched fractions. The 29RNP kinase was positively identified as a CKII alpha family protein by the presence of two unique peptides.

INSTRUMENT(S): LTQ FT

ORGANISM(S): Oryza Sativa (rice)

TISSUE(S): Plant Cell, Leaf

SUBMITTER: Qingtao Lu

LAB HEAD: Lu Qingtao

PROVIDER: PXD000951 | Pride | 2015-04-27

REPOSITORIES: Pride

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Publications

Identification and characterization of chloroplast casein kinase II from Oryza sativa (rice).

Lu Qingtao Q Ding Shunhua S Reiland Sonja S Rödiger Anja A Roschitzki Bernd B Xue Peng P Gruissem Wilhelm W Lu Congming C Baginsky Sacha S

Journal of experimental botany 20141014 1

Plastid casein kinase II is an important regulator of transcription, posttranscriptional processes, and, most likely, different metabolic functions in dicotyledonous species. Here we report the identification and characterization of pCKII from the monocotyledonous species Oryza sativa. OspCKII activity was enriched from isolated rice chloroplasts using heparin-Sepharose chromatography, in which it co-elutes with the transcriptionally active chromosome (TAC) and several ribosomal proteins. Inclus ...[more]

PMID: 25316064

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Project description:Glycerol kinase deficiency (GKD) is an X-linked inborn error of metabolism with metabolic and neurologic crises. Liver shows the highest level of glycerol kinase (GK) activity in humans and mice. Absence of genotype-phenotype correlations in patients with GKD indicate the involvement of modifier genes, including other network partners. To understand the molecular pathogenesis of GKD, we performed microarray analysis on liver mRNA from neonatal glycerol kinase (Gyk) knockout (KO) and wild type (WT) mice. Unsupervised learning revealed the overall gene expression profile of the KO mice was different from that of WT. Real time PCR confirmed differences for selected genes. Functional gene enrichment analysis was used to find 56 increased and 37 decreased gene functional categories. Pathway Assist analysis identified changes in gene expression levels of genes involved in organic acid metabolism indicating that GK was part of the same metabolic network which correlates well with the patients with GKD having metabolic acidemia during their episodic crises. Network component analysis (NCA) showed that transcription factors SREBP-1c, ChREBP, HNF-4alpha, and PPAR-alpha, had increased activity in the Gyk KO mice compared with WT mice; while SREBP-2 was less active in the Gyk KO mice. These studies show that Gyk deletion causes alterations in gene expression of genes in several regulatory networks and is the first time NCA has been used to expand on microarray data from a mouse knockout model of a human disease. Male WT and KO mouse pups were sacrificed on day of life (dol) 3 and each liver was harvested. Total RNA from 4 KO and 4 WT livers was isolated individually. cDNA was synthesized from the poly(A)+ mRNA in the total RNA, Biotin-tagged and fragmented to an average strand length of 100 bases (range 35-200 bases). Ten µg of each cRNA was hybridized onto an Affymetrix mus 430 2.0 GeneChip to analyze differences in liver gene expression between KO and WT mice. Day of life three was chosen because the mice are phenotypically symptomatic with statistically different parameters for hypoglycemia, acidosis; low bicarbonate and decreased base excess. On day of life 2 they are not significantly different from wild type in all of these important clinical phenotypes.

Dataset Information

29RNP kinase idetification - Identification and characterization of chloroplast casein kinase II from Oryza sativa (rice)

Publications

Identification and characterization of chloroplast casein kinase II from Oryza sativa (rice).

Similar Datasets

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