Purple acid phosphatases comprise a family of binuclear metal-containing acid hydrolases, representatives of which have been found in animals, plants, and fungi. The goal of this study was to characterize purple acid phosphatases from sweet potato tubers and soybean seeds and to establish their relationship with the only well-characterized plant purple acid phosphatase, the FeIII-ZnII-containing red kidney bean enzyme. Metal analysis indicated the presence in the purified sweet potato enzyme of 1.0 g-atom of iron, 0.6-0.7 g-atom of manganese, and small amounts of zinc and copper. The soybean enzyme contained 0.8-0.9 g-atom of iron, 0.7-0.8 g-atom of zinc per subunit, and small amounts of manganese, copper, and magnesium. Both enzymes exhibited visible absorption maxima at 550-560 nm, with molar absorption coefficients of 3200 and 3300 M(-1) cm(-1), respectively, very similar to the red kidney bean enzyme. Substrate specificities were markedly different from those of the red kidney bean enzyme. A cloning strategy was developed based on N-terminal sequences of the sweet potato and soybean enzymes and short sequences around the conserved metal ligands of the mammalian and red kidney bean enzymes. Three sequences were obtained, one from soybean and two from sweet potato. All three showed extensive sequence identity (>66%) with red kidney bean purple acid phosphatase, and all of the metal ligands were conserved. The combined results establish that these enzymes are binuclear metalloenzymes: Fe-Mn in the sweet potato enzyme and Fe-Zn in soybean. The sweet potato enzyme is the first well-defined example of an Fe-Mn binuclear center in a protein.

/pdf/binuclear-metal-centers-in-plant-purple-acid-phosphatases-fe-4d1tnbo6ig.pdf

Binuclear Metal Centers in Plant Purple Acid Phosphatases: Fe-Mn in Sweet Potato

Climacteric and non-climacteric fruits are differentiated by its ripening process, in particular by the involvement of ethylene, high respiration rates and the nature of the process, being autocatalytic or not, respectively. Here, we will focus on the biosynthesis, metabolism and function of three compounds (auxin, salicylic acid and melatonin) sharing not only a common precursor (chorismate), but also regulatory functions in plants, and therefore in fruits. Aside from describing their biosynthesis in plants, with a particular emphasis on common precursors and points of metabolic diversion, we will discuss recent advances on their role in fruit ripening and the regulation of bioactive compounds accumulation, both in climacteric and non-climacteric fruits.

/pdf/biosynthesis-metabolism-and-function-of-auxin-salicylic-acid-51mq4xzwfp.pdf

Biosynthesis, Metabolism and Function of Auxin, Salicylic Acid and Melatonin in Climacteric and Non-climacteric Fruits

Papaya (Carica papaya L.) is a commercially important fruit crop. Various phytohormones, particularly ethylene and auxin, control papaya fruit ripening. However, little is known about the interaction between auxin and ethylene signaling during the fruit ripening process. In the present study, we determined that the interaction between the CpARF2 and CpEIL1 mediates the interaction between auxin and ethylene signaling to regulate fruit ripening in papaya. We identified the ethylene-induced auxin response factor CpARF2 and demonstrated that it is essential for fruit ripening in papaya. CpARF2 interacts with an important ethylene signal transcription factor CpEIL1, thus increasing the CpEIL1-mediated transcription of the fruit ripening-associated genes CpACS1, CpACO1, CpXTH12 and CpPE51. Moreover, CpEIL1 is ubiquitinated by CpEBF1 and is degraded through the 26S proteasome pathway. However, CpARF2 weakens the CpEBF1-CpEIL1 interaction and interferes with CpEBF1-mediated degradation of CpEIL1, promoting fruit ripening. Therefore, CpARF2 functions as an integrator in the auxin-ethylene interaction and regulates fruit ripening by stabilizing CpEIL1 protein and promoting the transcriptional activity of CpEIL1. To our knowledge, we have revealed a novel module of CpARF2/CpEIL1/CpEBF1 that fine-tune fruit ripening in papaya. Manipulating this mechanism could help growers tightly control papaya fruit ripening and prolong shelf life.

CpARF2 and CpEIL1 interact to mediate auxin-ethylene interaction and regulate fruit ripening in papaya.

There is growing evidence to suggest that epigenetic tags, especially DNA methylation, are critical regulators of fruit ripening. To examine whether this is the case in sweet pepper (Capsicum annuum) we conducted experiments at the transcriptional, epigenetic, and physiological levels. McrBC PCR, bisulfite sequencing, and real-time PCR demonstrated that DNA hypomethylation occurred in the upstream region of the transcription start site of some genes related to pepper ripening at the turning stage, which may be attributed to up-regulation of CaDML2-like and down-regulation of CaMET1-like1, CaMET1-like2, CaCMT2-like, and CaCMT4-like. Silencing of CaMET1-like1 by virus-induced gene silencing led to DNA hypomethylation, increased content of soluble solids, and accumulation of carotenoids in the fruit, which was accompanied by changes in expression of genes involved in capsanthin/capsorubin biosynthesis, cell wall degradation, and phytohormone metabolism and signaling. Endogenous ABA increased during fruit ripening, whereas endogenous IAA showed an opposite trend. No ethylene signal was detected during ripening. DNA hypomethylation repressed the expression of auxin and gibberellin biosynthesis genes as well as cytokinin degradation genes, but induced the expression of ABA biosynthesis genes. In mature-green pericarp, exogenous ABA induced expression of CaDML2-like but repressed that of CaCMT4-like. IAA treatment promoted the transcription of CaMET1-like1 and CaCMT3-like. Ethephon significantly up-regulated the expression of CaDML2-like. Treatment with GA3 and 6-BA showed indistinct effects on DNA methylation at the transcriptional level. On the basis of the results, a model is proposed that suggests a high likelihood of a role for DNA methylation in the regulation of ripening in the non-climacteric pepper fruit.

/pdf/dna-methylation-is-involved-in-the-regulation-of-pepper-51rqumbtnm.pdf

DNA methylation is involved in the regulation of pepper fruit ripening and interacts with phytohormones.

Although exogenous applications of gibberellins (GAs) delay tomato ripening, the regulatory mechanisms of GAs in the process have never been well recognized. Here, we report that the concentration of endogenous GAs is declined before the increase of ethylene production in mature-green to breaker stage fruits. We further demonstrate that reductions in GA levels via overexpression of a GA catabolism gene SlGA2ox1 specifically in fruit tissues lead to early ripening. Consistently, we have also observed that application of a GA biosynthetic inhibitor, prohexadione-calcium, at the mature-green stage accelerates fruit ripening, while exogenous GA3 application delays the process. Furthermore, we demonstrate that ethylene biosynthetic gene expressions and ethylene production are activated prematurely in GA-deficient fruits but delayed/reduced in exogenous GA3-treated WT fruits. We also show that the GA deficiency-mediated activation of ethylene biosynthesis is due to the activation of the ripening regulator genes RIN, NOR and CNR. In conclusion, our results demonstrate that GAs play a negative role in tomato fruit ripening.

Gibberellins Play a Role in Regulating Tomato Fruit Ripening.

Silencing of SlGH3-2 in tomato alters auxin and ethylene levels during fruit ripening and cause reduced lycopene accumulation in the transgenic fruits. While auxin’s role during fleshy fruit ripening is widely acknowledged to be important, the physiological functions of several ripening-induced genes, especially those involved in the maintenance of cellular auxin homeostasis, largely remain under-explored. In the present study, the updated inventory shows that 24 members constitute the Gretchen-Hagen 3 (GH3) gene family in tomato. Their characterization using an expression profiling approach revealed that SlGH3-2, a member of the group II IAA-amido synthetase, is strongly induced at the commencement of fruit ripening. Phylogenetic analysis and homology modeling revealed that SlGH3-2 is the closest homolog of pepper CcGH3 and grapevine VvGH3-1. Expression profiling revealed that the mRNA level of SlGH3-2 is inhibited in ripening mutants such as ripening-inhibitor (rin) and Never-ripe (Nr). Whereas both auxin and ethylene were found to act as positive regulators of its transcript accumulation. The fruits of 35S::SlGH3-2 RNAi lines exhibited prolonged shelf-life. Both ethylene production and lycopene accumulation were affected in the fruits of SlGH3-2 silenced lines. These observations were corroborated by the altered expression of key ethylene and carotenoid biosynthesis genes such as ACS2 and PSY1, respectively, in the RNAi lines. Additionally, the SlGH3-2 silenced line fruits had higher IAA and IBA levels at the ripening stages, and showed increased transcript accumulation of several known auxin-induced SlIAA and SlGH3 genes. Altogether, the present study suggests that SlGH3-2 influences fruit ripening in tomato via modulating ethylene and auxin crosstalk, especially during the early phase.

A ripening-induced SlGH3-2 gene regulates fruit ripening via adjusting auxin-ethylene levels in tomato (Solanum lycopersicum L.)

Identification, structure analysis, and transcript profiling of purple acid phosphatases under Pi deficiency in tomato (Solanum lycopersicum L.) and its wild relatives.

Identification, evolutionary profiling, and expression analysis of F-box superfamily genes under phosphate deficiency in tomato.

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A ripening-induced SlGH3-2 gene regulates fruit ripening via adjusting auxin-ethylene levels in tomato (Solanum lycopersicum L.)

Identification, structure analysis, and transcript profiling of purple acid phosphatases under Pi deficiency in tomato (Solanum lycopersicum L.) and its wild relatives.

Identification, evolutionary profiling, and expression analysis of F-box superfamily genes under phosphate deficiency in tomato.