The role of Glycogen Synthase Kinase 3/ SHAGGY-like Kinases in Plant Development

Animal glycogen synthase kinase 3 (GSK-3)/ SHAGGY kinases, are known for 20 years or more. On the other hand, information about plant GSK 3/SHAGGY-like kinases (GSKs) is fairly recent. In a review article published in the October 2002 issue of Trend in Plant Science (vol. 7:457-461), Claudia Jonak and Heribert Hirt at the Institute of Microbiology and Genetics, University of Vienna, Austria, discuss the present status of our knowledge about the involvement of various members of GSK family in multitude of developmental processes such as flower development, brassinosteroid signaling, NaCl stress and wound responses.

The authors begin their article by summarizing the various vital roles that GSK-3 plays in regulating different pysiological processes in animal systems. GSK-3, first isolated from animals, was found to inhibit glycogen synthase from catalyzing the last step of glycogen synthesis. It was soon revealed that GSK-3 is a part of Wnt pathway that is involved in regulation of cell proliferation in adult tissues and cell fate during embryogenesis. GSK-3 is active in the absence of Wnt. The active GSK-3 phosphorylates – beta-catenin/armadillo, a transcriptional regulator.

Kinases are crucial for signal transduction. Kinases add a phosphate group via the transfer of the terminal phosphate from ATP to an amino acid residue of its substrate. The function of these post-translational modifications is to alter the substrate’s activity, subcellular localization, binding properties or association with other proteins.

With the help of suitable diagrams, the authors explain how the function of mammalian glycogen synthase kinase 3 (GSK-3) is regulated. GSK –3 has two phosphate binding sites (a) a priming phosphate site and (b) an active phosphate site. In certain instances, binding of a substrate to the active phosphate site occurs in two steps. First, the substrate is primed (made ready) by undergoing phosphorylation – which means the substrate is positioned in order to bind to the active site of GSK-3; in the second step the priming phosphorylated substrate positions itself to the active site of GSK-3 and binds to it for phosphorylation. Protein kinase B (PKB) can phosphorylate, GSK-3 at the N-terminus serine 9. The phosphorylation twists the N-terminus of GSK-3 and it assumes the appearance of a substrate called pseudosubstrate. Such a twist blocks the pocket of priming phosphorylated site of GSK-3 and thus prevents binding of the priming phosphorylated substrate with GSK-3. In other instances, GSK-3 forms a protein complex including axin and beta-catenin, followed by their phosphorylation. Formation of a such a complex does not require priming phosphorylated site. Complex formation is inhibited by FRAT, when it binds to the GSK-3 protein. Interestingly, this inhibitory mechanism does not affect substrates that require priming a phosphorylation.

Structural similarities between animal and plant GSKs: Animals and plant GSKs share a highly conserved kinase domain. They are all characterized by the T-loop having a tyrosine residue (Tyr216). The priming phosphate-binding pocket in both plants and animal are located in equivalent positions, namely, at Arg94, Arg180 and Lys205. However, the authors point out that there is one major difference distinguishing the two kingdoms. Plant GSKs do not possess the N-terminal Ser-9, so characteristic of animal GSKs, that is phosphorylated by PKB.

Biological functions of plant GSKs:
Recent studies with plant kinases have shown that AtGSK1 regulate the high-salt response in a positive manner. Another enzyme of this family, namely WIG (wound-induced GSK) is involved in wound signaling. Furthermore, AtSK11 and AtSK12 have been shown to determine the correct flower patterning. Analysis of bin 2 mutant (brassinosteroid-insensitive 2) has revealed that BIN2 is involved in brassinosteroid (BR) signaling.

Stress responses: Results reported recently show that while NaCl- and abscisic acid induce AtGSK1 mRNA accumulation, KCl was found to be ineffective in causing its induction. Overproduction of AtGSK1 affects intracellular cation levels, and induced salt stress-responsive genes, thereby enhancing salt and drought tolerance.

Results of another set of experiments demonstrated that wounding induced the kinase activity of GSK-3 (WIG) in Medicago sativa. In unaffected leaves of this species the levels of active WIG were low and an injury of the plant led to activation of the kinase to heal the damaged tissue. Injury-dependent activation of WIG is a post-translational process, whereas one or more protein factors may be involved in keeping the WIG protein from functioning under normal conditions.

Development: AtSK11 and AtSK12 have been shown to be involved in orchestrating flower patterning. The function of these two genes has been investigated by altering their transcript levels by antisense technology. Reduction of the transcript levels resulted in an increase of flower mersistems and perianth numbers accompanied by a change in the gynoecium pattern. The authors also describe the putative role of a few more ASKs. For instance, ASKz  is expressed in all parts of the embryo, whereas expression of ASKh is confined only to the suspensor. ASKb and ASK  mRNA accumulate in pollen grains. The remaining ASK members have been shown to be expressed differently in different organs.

Hormone signaling: The authors describe the relationship between ASK and brassinosteroid signaling. Brassinosteroids are a group of polyhydroxy steroids. Of these steroids, brassinolide is the most biologically active in the bioassay system. These steroid compounds have been shown to take part in a variety of plant functions such as etiolation, vascular differentia- tion and reproductive development (male sterility). The steroid hormone, brassino- lide is perceived by the receptor kinase BRI1 (brassinosteroid-insensitive 1). This receptor has an extracellular domain consisting of (a) leucine- rich repeats (LRRs), (b) a transmembrane domain and (c) a cytoplasmic kinase domain. Brassinosteroid binds to the extracellular domain of the receptor either directly or indirectly. Activation of the cytoplasmic protein kinase triggers the brassinosteroid signaling cascade. The authors describe the results of recent studies that show that BIN2 (brassinosteroid insensitive 2) is an important regulatory components of the brassinosteroid pathway. It is located down- stream of BRI1. It acts as a negative regulator by suppressing the activity of BRI1 as demonstrated by the gain-of-function nature of the semidwarf and semidominant mutants.

The authors describe the following model to explain the mode of action of BIN2. The action of BIN2 depends upon whether a brassino- steroid signal is present or absent. In the absence of a signal, BIN2 phosphorylates and negatively regulates protein levels of BES1 (Brassinosteroid-EMS-suppressor) and its homolog BZR1 (brassinazole-resistant 1), two positive effectors of brassinosteriod signaling. When a signal is perceived by the BRI1 receptor kinase, BIN2 is inhibited. Inactivation of BIN2 follows with simultaneous accumulation of BES1 and BRZ1 in the nucleus.

Semidominant ASK-eta mutants have also been isolated as ucu1 (ultracurvata 1) mutants. These mutants, ucu1 showed modified leaf development. ucu1 mutants were dwarf with circinate leaves – a phenotype similar to brassinosteroid-deficient mutants.

Recent years witnessed a lot of research activity leading to the discovery of a greater number of GSK 3 genes in plants (ten in Arabidopsis thaliana) and their regulatory role in plant development. Of great significance is the realization that GSKs play a significant role in transcriptional regulation. With the progress of ongoing study, the other regulatory roles of GSKs as to how some of its members might direct the formation of cytoskeletal proteins or control processes such as determining specific functions of metabolic enzymes will soon be discovered. Tying up all loose ends, will give a picture of plant Glycogen Synthase kinase 3/ SHAGGY-like kinases.’

 

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