SPICK2, a homologue of the weakly-inward-rectifying Shaker-like Arabidopsis K channel, AKT2, is a candidate K+-influx channel participating in light- and clock-regulated leaf movements of the legume, Samanea saman. Light and the biological clock regulate the in situ K+-influx channel activity differentially in extensor and flexor halves of the pulvinus (the S. saman leaf motor organ), and also - though differently - the transcript level of SPICK2 in the pulvinus. This disparity between the in situ channel activity versus its candidate transcript, along with the sequence analysis of SPICK2, suggest an in situ regulation of the activity of SPICK2, possibly by phosphorylation and/or by interaction with cAMP. Consistent with this (i) the activity of the voltage-dependent K+-selective fraction of the inward current in extensor and flexor cells was affected differentially in whole-cell patch-clamp assays promoting phosphorylation (using the protein phosphatase inhibitor okadaic acid); (ii) several proteins in isolated plasma membrane-enriched vesicles of the motor cells underwent phosphorylation without an added kinase in conditions similar to patch-clamp; and (iii) the SPICK2 protein was phosphorylated in vitro by the catalytic subunit of the broad-range cAMP-dependent protein kinase. All of these results are consistent with the notion that SPICK2 is the K+-influx channel, and is regulated in vivo directly by phosphorylation.
Bibliographical noteFunding Information:
We are grateful to Professor A Harmon for comments related to CDPKs, to Drs S Podell and M Gribskov for comments on the plant PKA, to Mr Juraj Sklenar for instruction on the preparation of membrane vesicles, to Professors Z Adam, M Cohen-Armon, H Fromm, D Hananshvili, Y Zik, and Y Lee, Drs Z Arazi, R Gurevitz, O Oesterzaetzer, and I. Sakler, and to Ms B Otto for friendly advice and helpful suggestions. We thank the Alomone Labs, Jerusalem, Israel for their professional advice on protein chemistry and a gift of okadaic acid. This research was supported by The Israel Science Foundation (Grant No. 550/01) to NM and, in part, by the Dead-Sea Works, Israel to NM, and an EMBO short-term fellowship to LY.
- Motor cells
- Potassium channel