Cloning and characterization of the cDNA coding for the catalytic α subunit of CK2 from tobacco

Salinas, Paula; Bantignies, Brigitte; Tapia, Julio C.; Jordana, Xavier; Holuigue, Loreto

doi:10.1007/978-1-4615-1723-8_16

Cited by 7 publications

(10 citation statements)

References 23 publications

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“…This result allowed us to conclude that the kinase activity purified on heparin-Sepharose was able to phosphorylate PVA CP in a heparin-sensitive manner using GTP as a phosphoryl donor. Moreover, the molecular mass of the identified kinase (39 kD) was identical to that of the recently reported ␣ -catalytic subunit of CK2 from tobacco (tCK2 ␣ ; Salinas et al, 2001). Ultimately, liquid chromatography-tandem mass spectrometry (LC-MS/MS) was use to prove the identity of the 39-kD protein kinase.…”

Section: Identification Of Tobacco Ck2 As the Kinase Involved In Pva mentioning

confidence: 55%

“…We used the well-studied ␣ -catalytic subunit of CK2 from maize (rmCK2 ␣ ; Boldyreff et al, 1993), which is Ͼ 90% identical at the protein level to its homolog from tobacco (Salinas et al, 2001). PVA genome-linked protein (VPg) was included in the assays to verify our previous results indicating that PVA VPg is a substrate for a protein kinase other than PVA CP and TMV MP (Ivanov et al, 2001).…”

Section: Pva Cp Is Phosphorylated By Ck2 In Vitromentioning

confidence: 99%

“…To produce recombinant tCK2 ␣ in bacterial cells, the full-length gene of tCK2 ␣ (Salinas et al, 2001) was amplified from a tobacco (B) PVA CP and TMV MP, but not PVA VPg, are phosphorylated efficiently by the ␣ -catalytic subunit of recombinant maize CK2. (C) Effect of DRB, a cell-permeable inhibitor of CK2, on the phosphorylation of PVA CP in vivo.…”

Section: Expression Purification and Catalytic Activity Of The Recomentioning

confidence: 99%

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Phosphorylation of the Potyvirus Capsid Protein by Protein Kinase CK2 and Its Relevance for Virus Infection [W]

et al. 2003

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We reported previously that the capsid protein (CP) of Potato virus A (PVA) is phosphorylated both in virus-infected plants and in vitro. In this study, an enzyme that phosphorylates PVA CP was identified as the protein kinase CK2. The ␣ -catalytic subunit of CK2 (CK2 ␣ ) was purified from tobacco and characterized using in-gel kinase assays and liquid chromatographytandem mass spectrometry. The tobacco CK2 ␣ gene was cloned and expressed in bacterial cells. Specific antibodies were raised against the recombinant enzyme and used to demonstrate the colocalization of PVA CP and CK2 ␣ in infected tobacco protoplasts. A major site of CK2 phosphorylation in PVA CP was identified by a combination of mass spectrometric analysis, radioactive phosphopeptide sequencing, and mutagenesis as Thr-242 within a CK2 consensus sequence. Amino acid substitutions that affect the CK2 consensus sequence in CP were introduced into a full-length infectious cDNA clone of PVA tagged with green fluorescent protein. Analysis of the mutant viruses showed that they were defective in cell-to-cell and longdistance movement. Using in vitro assays, we demonstrated that CK2 phosphorylation inhibited the binding of PVA CP to RNA, suggesting a molecular mechanism of CK2 action. These results suggest that the phosphorylation of PVA CP by CK2 plays an important regulatory role in virus infection.

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Section: Identification Of Tobacco Ck2 As the Kinase Involved In Pva mentioning

confidence: 55%

Section: Pva Cp Is Phosphorylated By Ck2 In Vitromentioning

confidence: 99%

Section: Expression Purification and Catalytic Activity Of The Recomentioning

confidence: 99%

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Phosphorylation of the Potyvirus Capsid Protein by Protein Kinase CK2 and Its Relevance for Virus Infection [W]

et al. 2003

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“…Casein kinase II (CKII) is a ubiquitous protein in eukaryotic organisms, and shows relatively higher levels of expression in proliferating tissues, such as root meristem. It is directly involved in the salicylic acid (SA) transduction pathway (Salinas et al 2001). Moreover, it has also been proposed that CKII functions in sensing ionic signals and transducing them to the cell cycle machinery (Glover 1998).…”

Section: Up-regulated Proteins Under Waterlogging Stressmentioning

confidence: 99%

A proteomic screen and identification of waterlogging-regulated proteins in tomato roots

Ahsan

Lee

et al. 2007

Plant Soil

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To gain a better understanding of the tomato root response to waterlogging stress at protein level, a proteomic approach has been conducted in combination with physiological analyses. Fiveweek old plants were subjected to waterlogging stress, and roots were collected 24 and 72 h after treatment. Waterlogging stress resulted in a gradual increase of lipid peroxidation and in vivo H 2 O 2 concentration in roots. To elucidate temporal changes in protein expression in tomato roots, proteins were extracted from the root samples, separated by 2-DE, and visualized by staining with Coomassie Brilliant Blue. A total of 35 proteins were diVerentially expressed in tomato roots in response to waterlogging stress, and 29 were identiWed using MALDI-TOF analysis. Of the proteins identiWed, 16 proteins were up-regulated and 13 proteins were down-regulated in waterlogged tomato roots. In addition to previously identiWed waterlogging-regulated proteins, we identiWed several novel proteins, including 3-beta-hydroxylase, phenylalanine ammonia-lyase, glutamyl-tRNA reductase, Xavanone 3-hydroxylase, mitochondrial ATPase alpha subunit, cysteine protease, DWARF1, and NIM1-like protein 2. These newly identiWed proteins are involved in several cellular processes, including hormone and secondary metabolite synthesis, programmed cell death, and stress and defense mechanisms. Five diVerentially accumulated proteins were further analyzed at the mRNA level, which conWrmed the diVerential gene expression, and revealed that the transcript levels were not always consistent with the translation level. The identiWcation of these novel proteins in the waterlogging stress response provides new insight that can lead to a better understanding of the molecular basis of waterlogging-sensitivity in plants. Abbreviations 2-DETwo-dimensional gel electrophoresis ANP Anaerobic polypeptide BR Brassinosteroid MALDI-TOF Matrix assisted laser desorption ionization time-of-Xight PAL Phenylalanine ammonia-lyase PEG Polyethylene glycol ROS Reactive oxygen species SAR Systemic acquired resistance SDS-PAGE Sodium dodecylsulfate polyacrylamide gel electrophoresis TBARS Thiobarbituric acid reactive substance

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“…In plants, genes encoding the α and β subunits belong to multigene families. In Arabidopsis thaliana, four genes encode CK2α (Mizoguchi et al 1993; Salinas et al 2006) and four CK2β (Collinge and Walker 1994; Sugano et al 1998; Salinas et al 2006). In Zea mays , until now, three cDNA clones encoding CK2β and three cDNA clones encoding CK2α (ZmCK2α‐1, ZmCK2α‐2, and ZmCK2α‐3) have been isolated and characterized (Dobrowolska et al 1991; Peracchia et al 1999; Riera et al 2001a,b).…”

Section: Introductionmentioning

confidence: 99%

A novel splicing variant encoding putative catalytic α subunit of maize protein kinase CK2

Łebska

Szczegielniak

Dobrowolska

et al. 2009

Physiologia Plantarum

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A cDNA highly homologous to the known catalytic alpha subunit of protein kinase CK2 was cloned from maize (Zea mays). It was designated ZmCK2alpha-4 (accession no. AAF76187). Sequence analysis shows that ZmCK2alpha-4 and the previously identified ZmCK2alpha-1 (accession no. X61387) are transcribed from the same gene, ZmPKCK2AL (accession no. Y11649), but at different levels in various maize organs and at different stages of development. The cDNA encoding ZmCK2alpha-4 has three potential translation initiation sites. The three putative variants of ZmCK2alpha-4 were expressed in Escherichia coli as GST-fusion proteins and purified from bacterial extracts. In contrast to the previously characterized ZmCK2alphas, the obtained GST:ZmCK2alpha-4 proteins were catalytically inactive as monomers or in the presence of equimolar amounts of the human CK2beta. However, GST:ZmCK2alpha-4 did phosphorylate casein in the presence of a large excess of the beta subunit. The activity of ZmCK2alpha-4 toward casein could also be stimulated by increasing ATP concentration. Modeling studies have shown that there is no interaction between the N-terminal segment of ZmCK2alpha-4 and the activation loop responsible for constitutive catalytic activity of CK2alpha. Preliminary results suggest that ZmCK2alpha-4 may function as a negative regulator of other CK2s, and at certain circumstances as a holoenzyme which catalytic activity is stimulated by specific regulatory subunit(s).

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Cloning and characterization of the cDNA coding for the catalytic α subunit of CK2 from tobacco

Cited by 7 publications

References 23 publications

Phosphorylation of the Potyvirus Capsid Protein by Protein Kinase CK2 and Its Relevance for Virus Infection [W]

Phosphorylation of the Potyvirus Capsid Protein by Protein Kinase CK2 and Its Relevance for Virus Infection [W]

A proteomic screen and identification of waterlogging-regulated proteins in tomato roots

A novel splicing variant encoding putative catalytic α subunit of maize protein kinase CK2

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