Genomic and functional characterization of StCDPK1

Abstract

StCDPK1 is a calcium dependent protein kinase expressed in tuberizing potato stolons and in sprouting tubers. StCDPK1 genomic sequence contains eight exons and seven introns, the gene structure is similar to Arabidopsis, rice and wheat CDPKs belonging to subgroup IIa. There is one copy of the gene per genome and it is located in the distal portion of chromosome 12. Western blot and immunolocalization assays (using confocal and transmission electron microscopy) performed with a specific antibody against StCDPK1 indicate that this kinase is mainly located in the plasma membrane of swelling stolons and sprouting tubers. Sucrose (4-8%) increased StCDPK1 protein content in non-induced stolons, however the amount detected in swelling stolons was higher. Transgenic lines with reduced expression of StCDPK1 (beta 7) did not differ from controls when cultured under multiplication conditions, but when grown under tuber inducing conditions some significant differences were observed: the beta 7 line tuberized earlier than controls without the addition of CCC (GA inhibitor), developed more tubers than wild type plants in the presence of hormones that promote tuberization in potato (ABA and BAP) and was more insensitive to GA action (stolons were significantly shorter than those of control plants). StCDPK1 expression was induced by GA, ABA and BAP. Our results suggest that StCDPK1 plays a role in GA-signalling and that this kinase could be a converging point for the inhibitory and promoting signals that influence the onset of potato tuberization.

Genome-wide analysis and expression profiling of calcium-dependent protein kinases in potato (Solanum tuberosum)

“…StCDPK1 and StCDPK3, with different substrate specificities and cellular distribution, were found associated with early and induced stolons, respectively. StCDPK1 was expressed in tuberizing stolons and sprouting tubers (Raíces et al 2001;Gargantini et al 2009), whereas expression of StCDPK3 was abundant in stolons, roots and leaves (Grandellis et al 2012). Transcript of another CDPK, StCDPK2, was present in all plant tissues studied and during different developmental processes -sprouting and tuberization (Giammaria et al 2011); its level was the highest in actively growing young leaves.…”

“…StCDPK1 transcript was observed in mesh bulbs and that of StCDPK2 mainly in the leaves. StCDPK1 also plays a role in gibberellic acid (GA) signaling and influences tuberization (Gargantini et al 2009) while StCDPK2 is associated with plant growth and development (Giammaria et al 2011). An application of abscisic acid (ABA) or GA had no significant effect on the activity and expression of StCDPK2 (Giammaria et al 2011), while treatment with jasmonic acid decreased StCDPK2 expression at both these levels (Ulloa et al 2002).…”

StCDPK2 expression and activity reveal a highly responsive potato calcium-dependent protein kinase involved in light signalling

“…Primers were designed based on the gene structure of StCdpk1 (Gargantini et al 2009). A PCR fragment containing StCDPK2 regulatory domain interrupted by three introns was labelled with dCTP using the High Prime Kit (Roche, Indianapolis, IN, USA).…”

“…A PCR fragment containing StCDPK2 regulatory domain interrupted by three introns was labelled with dCTP using the High Prime Kit (Roche, Indianapolis, IN, USA). This fragment together with PCR fragments amplified from StCdpk1 (intron 3, Gargantini et al 2009) or StCdpk3 (intron 6-3 0 utr) genes were used to probe a genomic library from the diploid potato clone HB171. The screening was performed according to Hein et al (2007).…”

“…membranes (Amersham Pharmacia Biotech, Uppsala, Sweden). Membranes were hybridized overnight with a [ 32 P]-labelled full-length StCDPK2 probe or with a 400 nt PCR fragment corresponding to intron 4 (probe 2) at 65°C in Church's buffer according to Gargantini et al (2009).…”