StCDPK2 expression and activity reveal a highly responsive potato calcium-dependent protein kinase involved in light signalling
- 276 Downloads
Calcium-dependent protein kinases (CDPKs) are essential calcium sensors. In this work, we have studied StCDPK2 isoform from potato both at gene and protein level. StCdpk2 genomic sequence contains eight exons and seven introns, as was observed for StCdpk1. There is one copy of the gene per genome located in chromosome 7. StCDPK2 encodes an active CDPK of 515 aminoacids, with an apparent MW of 57 kDa, which presents myristoylation and palmitoylation consensus in its N-terminus. StCDPK2 is highly expressed in leaves and green sprouts; enhanced expression was detected under light treatment, which corresponds well with light responsive cis-acting elements found in its promoter sequence. Antibodies against the recombinant StCDPK2::6xHis protein detected this isoform in soluble and particulate fractions from leaves. StCDPK2 autophosphorylation and kinase activity are both calcium dependent reaching half maximal activation at 0.6 μM calcium. The active kinase is autophosphorylated on serine and tyrosine residues and its activity is negatively modulated by phosphatidic acid (PA). Our results reveal StCDPK2 as a signalling element involved in plant growth and development and show that its activity is tightly regulated.
KeywordsAutophosphorylation CDPK Light Phospholipids Solanum tuberosum
Calcium-dependent protein kinase
We thank technical assistance of Karen McClean. We thank Dr. Sara Maldonado for helpful discussion and the technical assistance of Verónica Lainez. We thank Dr Salomé Prat Monguío (CSIC, Madrid, Spain) for the cDNA library from tuberizing stolons and Dr. Ana L. Villasuso and Dr. Estela E. Machado for providing the phospholipids. This work was supported by grants from CONICET, UBA, FONCYT-ANPCYT and the Royal Society of Edinburgh. RMU and PRG are members of CONICET. VG, CG and SB are fellows from CONICET.
- Choi HI, Park HJ, Park JH, Kim S, Im MY, Seo HH, Kim YW, Hwang I, Kim SY (2005) Arabidopsis calcium-dependent protein kinase AtCPK32 interacts with ABF4, a transcriptional regulator of abscisic acid-responsive gene expression, and modulates its activity. Plant Physiol 139:1750–1761CrossRefPubMedGoogle Scholar
- Harris N, Spence J, Oparka KJ (1994) Immunocytochemistry for light and electron microscopy. In: Harris N, Oparka KJ (eds) Plant cell biology: a practical approach. IRL Press at Oxford University Press, Oxford, pp 157–176Google Scholar
- Hein I, McLean K, Chalhoub B, Bryan G (2007) Generation and screening of a BAC library from a diploid potato clone to unravel durable late blight resistance on linkage group IV. Int J Plant Genomics (article ID 51421)Google Scholar