Effects of sucrose on the expression of protein kinase and protein phosphatases in connection with fructan synthesis initation

MARTÍNEZ NOËL G; TOGNETTI J; PONTIS H

Denver, USA

Congreso; ASPB; 2002

The enzyme sucrose-sucrose fructosyl transferase (6-SFT) is responsible for fructan synthesis initiation in cereal leaves. This enzyme is induced by increased sugar concentration, as verified in experiments using detached wheat leaves whose cut end were immersed in sugar solutions. However, the signaling pathway leading to 6-SFT gene induction is still unclear. Previous experiments from our laboratory (Martínez Noël et al., Planta 213:640-646) have shown that sugar mediated induction requires both protein phosphorylation and dephosphorylation, since this process is blocked by including protein kinase and protein phosphatase inhibitors such as staurosporine (Ser/Thr protein kinase inhibitor), w7 (calmodulin antagonist) and okadaic acid (protein phosphatase 1 and 2A inhibitor) into sugar solutions. Addition of mannose, a Pi sequestering agent, to sugar solutions inhibited 6-SFT induction while the addition of Pi did not. The process is also dependent on calcium ions, since the addition of different chelating agents like EDTA or EGTA and the specific Ca+2 chelator BAPTA inhibited sugar dependent induction of 6-SFT. We have cloned a fragment of 190 bp and a fragment of 104 bp corresponding to a calcium dependent protein kinase (CDPK) and a protein phosphatase 1 (PP1), respectively using degenerated primers from leaves treated with sucrose. The CDPK is inducible by sucrose and the PP1 is downregulated by the sugar as shown by RT-PCR experiments. Besides, we have measured the activity of the protein phosphatase 2A (PP2A), the most sensitive phosphatase to okadaic acid, and it decreased with the treatment with sucrose. We are currently studying protein phosphatase and protein kinase activities in connection with sugar mediated 6-SFT induction. Supported by ANPCyT and FIBA