The phytohormone abscisic acid (ABA) regulates stress-responsive gene expression during vegetative growth. The ABA regulation of many genes is mediated by a subfamily of basic leucine zipper class transcription factors referred to as ABFs (i.e. ABF1-ABF4), whose transcriptional activity is induced by ABA. Here we show that a calcium-dependent protein kinase is involved in the ABA-dependent activation process. We carried out yeast two-hybrid screens to identify regulatory components of ABF4 function and isolated AtCPK32 as an ABF4-interacting protein. AtCPK32 has autophosphorylation activity and can phosphorylate ABF4 in vitro. Mutational analysis indicated that serine-110 of ABF4, which is highly conserved among ABF family members, may be phosphorylated by AtCPK32. The serine-110 residue is essential for ABF4-AtCPK32 interaction, and transient expression assay revealed that it is also required for the normal transcriptional function of ABF4. The expression patterns and subcellular localization of AtCPK32 are similar to those of ABF4. Furthermore, its overexpression affects both ABA sensitivity and the expression of a number of ABF4-regulated genes. Together, our data demonstrate that AtCPK32 is an ABA signaling component that regulates the ABA-responsive gene expression via ABF4.
Sesamum indicum L. was used as an important oil crop in the world. An efficient protocol for in vitro plant regeneration via adventitious shoot formation from deembryonated cotyledon explants isolated from mature seeds of sesame is developed. Optimal medium for direct adventitious shoot formation was Murashige and Skoog (MS) medium with 22.2 μM 6-benzylaminopurin (BA) and 5.7 μM indole-3-acetic acid (IAA). Abscisic acid (3.8 μM ABA) and AgNO 3 (29.4 μM) were effective in enhancing the frequency of adventitious shoot formation. Preculture of cotyledon explants on high sucrose concentration (6-9%) for 2 wk and subsequent transfer to 3% sucrose enhanced the frequency of adventitious shoot induction. Root formation from the adventitious shoots was easily achieved on MS medium containing 2.7 μM of !-naphthalene acetic acid (NAA). Regenerated plantlets were acclimatized on sand and peat moss (1:1), showing 95% survival with subsequent flowering and seed set. We established the high-frequency plant regeneration via adventitious shoot formation in S. indicum L.
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