Adenylate cyclase is the key enzyme solely synthesizing cAMP which participates in cell metabolism regulations and functions as an intracellular second messenger. However, the biological functions of plant ACs have not been elucidated clearly for their poor conservative sequences and low detectable cAMP. We performed a systematic study of plant ACs by using Chinese jujube, whose fruit exhibits the highest cAMP content among plants. Three novel ACs were identified from Chinese jujube, and two types of methods including in vitro and in vivo were used to certificate ZjAC1−3 which can catalyze the conversion of ATP into cAMP. The biological functions of significant accelerations of seed germination, root growth, and flowering were found via overexpression of these AC genes in Arabidopsis, and these functions of ACs were further demonstrated by treating the AC-overexpressing transgenic lines and wild type Arabidopsis with bithionol and dibutyryl-cAMP. At last, transcriptome data revealed that the underlying mechanism of the biological functions of ACs might be regulation of the key genes involved in the circadian rhythm pathway and the hormone signal transduction pathway. This research established a foundation for further investigating plant AC genes and provided strong evidence for cAMP serving as a signaling molecule in plants.
Adenylyl cyclase (AC) is the vital enzyme for generating 3′,5′-cyclic adenosine monophosphate, an important signaling molecule with profound nutritional and medicinal values. However, merely, a dozen of AC proteins have been reported in plants so far. Here, a protein annotated as triphosphate tunnel metalloenzyme (PbrTTM1) in pear, the important worldwide fruit plant, was firstly identified to possess AC activity with both in vivo and in vitro methods. It exhibited a relatively low AC activity but was capable of complementing AC functional deficiencies in the E. coli SP850 strain. Its protein conformation and potential catalytic mechanism were analyzed by means of biocomputing. The active site of PbrTTM1 is a closed tunnel constructed by nine antiparallel β-folds surrounded with seven helices. Inside the tunnel, the charged residues were possibly involved in the catalytic process by coordinating with divalent cation and ligand. The hydrolysis activity of PbrTTM1 was tested as well. Compared to the much higher capacity of hydrolyzing, the AC activity of PbrTTM1 tends to be a moonlight function. Through a comparison of protein structures in various plant TTMs, it is reasonable to speculate that many plant TTMs might possess AC activity as a form of moonlighting enzyme function.
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