Нові генетичні конструкції KIN10-His/KIN11-His як інструмент для встановлення функціональної гомології протеїнкіназ SnRK1 та BSRK

  • О. Є. Краснопьорова Інститут харчової біотехнології і геноміки НАН України, Україна, 04123, м. Київ-123, вул. Осиповського, 2а
  • С. В. Ісаєнков Інститут харчової біотехнології і геноміки НАН України, Україна, 04123, м. Київ-123, вул. Осиповського, 2а
  • П. А. Карпов Інститут харчової біотехнології і геноміки НАН України, Україна, 04123, м. Київ-123, вул. Осиповського, 2а
  • А. І. Ємець Інститут харчової біотехнології і геноміки НАН України, Україна, 04123, м. Київ-123, вул. Осиповського, 2а

Анотація

Aim. The protein kinases SnRK1 from Arabidopsis thaliana are one of the key regulators of plant responses to different types of abiotic stresses. Many functions of these enzymes have not been studied yet. The possible functions of these protein kinases are regulation of cytoskeletal elements. To gain insight into molecular mechanisms of interaction of these enzymes with the cytoskeleton elements and discovery of potential substrates, the genetic constructs pGWB8-KIN10:His and pGWB8-KIN11:His for plant transformation were created. Methods. The coding sequences of KIN10 and KIN11 were cloned using gateway cloning system and other molecular-biological methods including PCR, RT-PCR. Results. The KIN10 and KIN11 His-tag fusions in genetic constructs pGWB8-KIN10:His and pGWB8-KIN11:His were created. Conclusions. We have created plasmid constructs pGWB8-KIN10:His аnd pGWB8-KIN11:His. According to bioinformatical analysis the KIN10 and KIN11 shared high level of homology with human BRSK1. Thus the KIN10 and KIN11might play important role in regulation of cytoskeleton. Created His-tag constructs can be used for identification of new substrates among cytoskeletal and other proteins.
Keywords: SnRK1, protein kinases, plasmid construct, сytoskeletal elements, polyhistidine-tag.

Посилання

Emanuelle S., Hossain M.I., Moller I.E., Pedersen H.L., van de Meene A.M., Doblin M.S., Koay A., Oakhill J.S., Scott J.W., Willats W.G., Kemp B.E., Bacic A., Gooley P.R., Stapleton D.I. SnRK1 from Arabidopsis thaliana is an atypical AMPK. Plant J. 2015. V. 82(2). P. 183–1892. doi: 10.1111/tpj.12813

Karpov P.A., Nadezhdina E.S., Yemets A.I., Matusov V.G., Nyporko A.Yu., Shashina N.Yu., Blume Ya.B. Bioinformatic search of plant microtubule- and cell cycle related serine-threonine protein kinases. BMC Genomics. 2010. 11, Suppl. 1. S 14. doi: 10.1186/1471-2164-11-S1-S14.

Karpov P.A., Rayevsky A.V., Blume Ya.B. Bioinformatic search for plant homologs of the protein kinase Bub1 a key component of the mitotic spindle assembly checkpoint. Cytol. Genetics. 2010. V. 44(6). P. 376–388. doi: 10.3103/S0095452710060095

Witczak C.A., Sharoff C.G., Goodyear L.J. AMP-activated protein kinase in skeletal muscle: From structure and localization to its role as a master regulator of cellular metabolism. Cell. Mol. Life Sci. 2008. 65. P. 3737. doi: 10.1007/s00018-008-8244-6.

Sanz P. Snf1 protein kinase: a key player in the response to cellular stress in yeast. Biochem. Soc. Trans. 2003. V. 31 (Pt 1). P. 178-181. doi: 10.1042/bst0310178

Halford N.G., Hey S.J. Snf1-related protein kinases (SnRKs) act within an intricate network that links metabolic and stress signalling in plants. Biochem. J. 2009. V. 419(2). P. 247-259. doi: 10.1042/BJ20082408

Polge C., Thomas M. SNF1/AMPK/SnRK1 kinases, global regulators at the heart of energy control? Trends. Plant Sci. 2007. V.21(1). P. 20-28. doi: 10.1016/j.tplants.2006.11.005

Nietzsche M., Schießl I., Börnke F. The complex becomes more complex: protein-protein interactions of SnRK1 with DUF581 family proteins provide a framework for cell- and stimulus type-specific SnRK1 signaling in plants. Front. Plant Sci. 2014. V. 5(54). doi: 10.3389/fpls.2014.00054.

Tomé F., Nägele T., Adamo M., Garg A., Marco-llorca C., Nukarinen E., Pedrotti L., Peviani A., Simeunovic A., Tatkiewicz A., Tomar M., Gamm M. The low energy signaling network. Front. Plant Sci. 2014. V. 5. doi: 10.3389/fpls.2014.00353.

Tsai A.Y.-L., Gazzarrini S. Trehalose-6-phosphate and SnRK1 kinases in plant development and signaling: the emerging picture. Front. Plant Sci. 2014. V. 5(119). doi: 10.3389/fpls.2014.00119.

Cho H.Y., Wen T.N., Wang Y.T., Shih M.C. Quantitative phosphoproteomics of protein kinase SnRK1 regulated protein phosphorylation in Arabidopsis under submergence. J. Exp. Bot. 2016. V.67(9). P. 2745-2760. doi: 10.1093/jxb/erw107

Crozet P., Margalha L., Butowt R., Fernandes N., Elias C.A., Orosa B., Tomanov K., Teige M., Bachmair A., Sadanandom A., Baena-González E. SUMOylation represses SnRK1 signaling in Arabidopsis. Plant J. 2016. V.85(1). P. 120-133. doi: 10.1111/tpj.13096

Shin J., Sánchez-Villarreal A., Davis A.M., Du S.X., Berendzen K.W., Koncz C., Ding Z., Li C., Davis S.J. The metabolic sensor AKIN10 modulates the Arabidopsis circadian clock in a light-dependent manner. Plant Cell Environ. 2017. V. 5. doi: 10.1111/pce.12903.

Jamsheer K.M., Laxmi A. Expression of Arabidopsis FCS-Like Zinc finger genes is differentially regulated by sugars, cellular energy level, and abiotic stress. Front. Plant Sci. 2015. V. 24(6) :746. doi: 10.3389/fpls.2015.00746

Chiang C.P., Li C.H., Jou Y., Chen Y.C., Lin Y.C., Yang F.Y., Huang N.C., Yen H.E. Suppressor of K+ transport growth defect 1 (SKD1) interacts with RING-type ubiquitin ligase and sucrose non-fermenting 1-related protein kinase (SnRK1) in the halophyte ice plant. J. Exp. Bot. 2013. V. 64(8). P. 2385-2400. doi: 10.1093/jxb/ert097

Krasnop'orova O.Ie., Isaienkov S.V., Karpov P.A., Iemets' A.I., Blium Ya.B. Kladystychnyy analiz seryn-treoninovoi proteinkinazy KIN10 ta osoblyvosti ii ekspresii v riznykh orhanakh Arabidopsis thaliana. Dop. NAN Ukrainy. 2016. No. 1. P. 81-91. [in Ukrainian]

Bright J.N., Carling D., Thornton C. Investigating the regulation of brain-specific kinases 1 and 2 by phosphorylation. J. Biol. Chem. 2008. V.283(22). P. 14946-14954. doi: 10.1074/jbc.M710381200

Karimi M., Inzé D., Depicker A. GATEWAY vectors for Agrobacterium-mediated plant transformation. Trends. Plant Sci. 2002. V.7(5). P. 193-195. doi: 10.1016/S1360-1385(02)02251-3

Rozen S., Skaletsky H.J. Primer3 on the WWW for general users and for biologist programmers. In: Krawetz S, Misener S (Eds) Bioinformatics Methods and Protocols: Methods in Molecular Biology. Humana Press, Totowa, NJ. 2000. P. 365-386. doi: 10.1385/1-59259-192-2:365

Freuler F., Stettler T., Meyerhofer M., Leder L., Mayr L.M. Development of a novel Gateway-based vector system for efficient, multiparallel protein expression in Escherichia coli. Protein Expr. Purif. 2008. V. 59 (2). P. 232-241. doi: 10.1016/j.pep.2008.02.003

Inoue H., Nojima H., Okayama H. High efficiency transformation of Escherichia coli with plasmids. Gene. 1990. V.96(1). P. 23-28. doi: 10.1016/0378-1119(90)90336-P

Cui W., Liu W., Wu G. A simple method for the transformation of Agrobacterium tumefaciens by foreign DNA. Chin. J. Biotechnol. 1995. V.11(4). P. 267-274.

The UniProt Consortium. The Universal Protein Resource (UniProt). Nucl. Acids Res. 2008. V. 36. P. 190-195. doi: 10.1093/nar/gkm895

Korf I., Yandell M., Bedell J. BLAST. Sebastopol. O'Reilly and Associates, 2003. 368 p.

Larkin M.A., Blackshields G., Brown N.P., Chenna R., McGettigan P.A. McWilliam H., Valentin F., Wallace I.M., Wilm A., Lopez R., Thompson J.D., Gibson T.J., Higgins D.G. Clustal W and ClustalX version 2.0. Bioinformatics. 2007. V. 23. P. 2947-2948. doi: 10.1093/bioinformatics/btm404

Kumar S., Stecher G., Tamura K. MEGA 7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Mol. Biol. Evol. 2016. V. 33. P. 1870-1874. doi: 10.1093/molbev/msw054

Yang J., Yan R., Roy A., Xu D., Poisson J., Zhang Y. The I-TASSER Suite: protein structure and function prediction. Nature Meth. 2015. V. 12(1). P. 7-8. doi: 10.1038/nmeth.3213