TOUSLED is a nuclear serine/threonine protein kinase that requires a coiled-coil region for oligomerization and catalytic activity.
The TOUSLED (TSL) gene is essential for the proper morphogenesis of leaves and flowers in Arabidopsis thaliana. Protein sequence analysis predicts TSL is composed of a carboxyl-terminal protein kinase catalytic domain and a large amino-terminal regulatory domain. TSL fusion proteins, expressed in and purified from yeast, were used to demonstrate TSL protein kinase activity in vitro. TSL trans-autophosphorylates on serine and threonine residues, and phosphorylates exogenous substrates. Using the yeast two-hybrid system, TSL was found to oligomerize via its NH2-terminal domain. A deletion series indicates that a region containing two alpha-helical segments predicted to participate in a coiled-coil structure is essential for oligomerization. TSL localizes to the nucleus in plant cells through an essential NH2-terminal nuclear localization signal; however, this signal is not necessary for protein kinase activity. Finally, deletion mutants demonstrate a strict correlation between catalytic activity and the ability to oligomerize, arguing that activation of the protein kinase requires interaction between TSL molecules.
Roe
J L
JL
Department of Plant Biology, University of California, Berkeley, California 94720, USA. jlr@nature.berkeley.edu
Durfee
T
T
Zupan
J R
JR
Repetti
P P
PP
McLean
B G
BG
Zambryski
P C
PC
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology*; D029681:Arabidopsis Proteins; D015153:Blotting, Western; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011487:Protein Conformation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D017384:Sequence Deletion
D029681:Arabidopsis Proteins; D011494:Protein Kinases; C084661:tousled protein, Arabidopsis; D017346:Protein Serine-Threonine Kinases