News and Publications

Calcium-Dependent Kinases and Ion Pumps in Plants

J. Harper, B. Hong, V. Vitart, J.-F. Huang, J. Christodoulou, Y. Wang, W. Chazin*

* Department of Molecular Biology, TSRI

The central focus of our laboratory is calcium signal transduction. Two general questions are being addressed: how are calcium signals formed, and how are they "decoded"? We use a model plant called Arabidopsis. This model system is expected to be the first higher plant to have its genome sequenced, providing a complete data base to help identify all the components of its calcium signaling systems.

A key component of calcium signaling in plants is a unique family of calcium sensors termed calcium-dependent protein kinases (CDPKs). This kinase family has been detected only in plants and protists. CDPKs are defined by their unique structure: they contain, in a single polypeptide, both a kinase domain and a calmodulin-like regulatory domain. This "fused" structural arrangement makes CDPKs distinct from other calcium-regulated protein kinases found in animal systems, such as protein kinase C and myosin light-chain kinase.

Evidence indicates that CDPKs are activated by intramolecular binding between their calmodulin-like domain and an autoinhibitory domain. These kinases are the first example of a calmodulin-like protein that has its target-binding sequence within the same polypeptide. One of our specific objectives is to understand the structural basis of how this intramolecular binding activates the kinase.

We are also investigating 2 families of P-type ATPases that pump calcium ions across different membranes. Our long-term goal is to determine the role of calcium pumps in defining the magnitude and duration of a calcium signal (i.e., the information content). The rationale is that the "shape" of a calcium signal is regulated by the rates of influx and efflux. Because the pumps are the primary control for calcium efflux, they provide potential control points for altering a calcium signal. Our specific focus is on 2 different calcium pumps, both located in the plant endoplasmic reticulum. One of these pumps (ACA2) is of special interest for 2 reasons: it has a unique structural arrangement with a calmodulin-regulated autoinhibitor located in the N-terminal instead of the C-terminal end, and it is the first calmodulin-regulated pump found in the endoplasmic reticulum of any organism. Why the plant endoplasmic reticulum contains 2 different types of calcium pumps is not known.

Our studies on ion pumps extend beyond our primary focus on calcium. An important long-term goal is to determine how different P-type ATPases selectively transport ions other than calcium, such as protons, molybdenum, and manganese. Our initial focus is a molybdenum uptake pump, the first to be detected in any organism. The challenge is to understand the structural basis for how this pump transports a molybdenum ion through a series of heavy metal--binding motifs. Molybdenum is an essential micronutrient for plants and animals. In plants, it is essential for nitrogen fixation and assimilation. Little is known about how molybdenum and other heavy-metal micronutrients are absorbed and transported within a plant, despite their importance for the fundamental biochemical reactions that make life possible.

PUBLICATIONS

Harper, J.F., Hong, B., Hwang, I., Guo, H.Q., Stoddard, R., Huang, J.F., Palmgren, M.G., Sze, H. A novel calmodulin-regulated Ca²⁺-ATPase (ACA2) from Arabidopsis with an N-terminal autoinhibitory domain. J. Biol. Chem. 273:1099, 1998.

Harrington, G.N., Franceschi, V.R., Offler, C.E., Patrick, J.W., Tegeder, M., Frommer, W.B., Harper, J.F., Hitz, W.D. Cell-specific expression of three genes involved in plasma membrane sucrose transport in developing Vicia faba seed. Protoplasma 197:160, 1997.

Stratmann, T., Schmid, S.R., Harper, J.F., Kang, A.S. Bacterial expression and purification of recombinant Plasmodium yoelii circumsporozoite protein. Protein Express. Purif. 11:72, 1997.