ABSTRACT
Actin nucleation is the key rate limiting step in the process of actin polymerization, and tight regulation of this process is critical to ensure actin filaments form only at specific times and at defined regions of the cell. WH2 domains are short sequence motifs found in many different actin binding proteins including WASP family proteins which regulate the actin nucleating complex Arp2/3. In this study we reveal a phosphorylation site, Serine 554, within the WH2 domain of the yeast WASP homologue Las17. Both phosphorylation and a phospho-mimetic mutation reduce actin monomer binding affinity while an alanine mutation, generated to mimic the non-phosphorylated state, increases actin binding affinity. The effect of these mutations on the Las17-dependent process of endocytosis in vivo was analysed and leads us to propose that switching of Las17 phosphorylation states may allow progression through distinct phases of endocytosis from site assembly through to the final scission stage. While the study is focused on Las17, the sole WASP family protein in yeast, our results have broad implications for our understanding of how a key residue in this conserved motif can underpin the many different actin regulatory roles with which WH2 domains have been associated.
Subject(s)
Actins/metabolism , Endocytosis , Protein Domains , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae/metabolism , Wiskott-Aldrich Syndrome Protein/metabolism , Amino Acid Sequence , Mutation , Phosphorylation , Polymerization , Protein Binding , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae Proteins/genetics , Sequence Homology, Amino Acid , Wiskott-Aldrich Syndrome Protein/chemistry , Wiskott-Aldrich Syndrome Protein/geneticsABSTRACT
C4 and crassulacean acid metabolism (CAM) have evolved in the order Caryophyllales many times but neither C4 nor CAM have been recorded for the Basellaceae, a small family in the CAM-rich sub-order Portulacineae. 24 h gas exchange and day-night changes in titratable acidity were measured in leaves of Anredera baselloides exposed to wet-dry-wet cycles. While net CO2 uptake was restricted to the light period in well-watered plants, net CO2 fixation in the dark, accompanied by significant nocturnal increases in leaf acidity, developed in droughted plants. Plants reverted to solely C3 photosynthesis upon rewatering. The reversible induction of nocturnal net CO2 uptake by drought stress indicates that this species is able to exhibit CAM in a facultative manner. This is the first report of CAM in a member of the Basellaceae.
