Amino acid transporters are the main mediators of nitrogen distribution throughout the plant body, and are essential for sustaining growth and development. In this review, we summarize the current state of knowledge on the identity and biological functions of amino acid transporters in plants, and discuss the regulation of amino acid transporters in response to environmental stimuli. We focus on transporter function in amino acid assimilation and phloem loading and unloading, as well as on the molecular identity of amino acid exporters. Moreover, we discuss the effects of amino acid transport on carbon assimilation, as well as their cross-regulation, which is at the heart of sustainable agricultural production.
In the fleshy fruit of cucumbers (Cucumis sativus L.), the phloem flow is unloaded via an apoplasmic pathway, which requires protein carriers to export sugars derived from stachyose and raffinose into the apoplasm. However, transporter(s) involved in this process remain unidentified. Here, we report that a hexose transporter, CsSWEET7a (Sugar Will Eventually be Exported Transporter 7a), was highly expressed in cucumber sink tissues and localized to the plasma membrane in companion cells of the phloem. Its expression level increased gradually during fruit development. Down-regulation of CsSWEET7a by RNA interference (RNAi) resulted in smaller fruit size along with reduced soluble sugar levels and reduced allocation of 14C-labelled carbon to sink tissues. CsSWEET7a overexpression lines showed an opposite phenotype. Interestingly, genes encoding alkaline α-galactosidase (AGA) and sucrose synthase (SUS) were also differentially regulated in CsSWEET7a transgenic lines. Immunohistochemical analysis demonstrated that CsAGA2 co-localized with CsSWEET7a in companion cells, indicating cooperation between AGA and CsSWEET7a in fruit phloem unloading. Our findings indicated that CsSWEET7a is involved in sugar phloem unloading in cucumber fruit by removing hexoses from companion cells to the apoplasmic space to stimulate the raffinose family of oligosaccharides (RFOs) metabolism so that additional sugars can be unloaded to promote fruit growth. This study also provides a possible avenue towards improving fruit production in cucumber.
Sugars are necessary for plant growth and fruit development. Cucumber (Cucumis sativus L.) transports sugars, mainly raffinose family oligosaccharides (RFOs), in the vascular bundle. As the dominant sugars in cucumber fruit, glucose and fructose are derived from sucrose, which is the product of RFO hydrolysis by α-galactosidases. Here, we characterized the cucumber alkaline α-galactosidase 2 (CsAGA2) gene and found that CsAGA2 has undergone human selection during cucumber domestication. Further experiments showed that the expression of CsAGA2 increases gradually during fruit development, especially in fruit vasculature. In CsAGA2-RNA interference (RNAi) lines, fruit growth was delayed because of lower hexose production in the peduncle and fruit main vascular bundle (MVB). In contrast, CsAGA2-overexpressing (OE) plants displayed bigger fruits. Functional enrichment analysis of transcriptional data indicated that genes related to sugar metabolism, cell wall metabolism, and hormone signaling were significantly downregulated in the peduncle and fruit MVBs of CsAGA2-RNAi plants. Moreover, downregulation of CsAGA2 also caused negative feedback regulation on source leaves, which was shown by reduced photosynthetic efficiency, fewer plasmodesmata at the surface between mesophyll cell and intermediary cell or between intermediary cell and sieve element, and downregulated gene expression and enzyme activities related to phloem loading, as well as decreased sugar production and exportation from leaves and petioles. The opposite trend was observed in CsAGA2-OE lines. Overall, we conclude that CsAGA2 is essential for cucumber fruit set and development through mediation of sugar communication between sink strength and source activity.
BackgroundGuard cell protoplasts (GCPs) isolated from various plants have proven to be especially useful for studies of signal transduction pathways and plant development. But it is not easy to isolate highly purified preparations of large numbers of GCPs from plants. In this research, our focus is on a method to isolate large numbers of guard cells from tomato leaves. The protocols described yield millions of highly purified, viable GCPs, which are also suitable for studies on guard cell physiology.ResultsWe developed an efficient method for isolating GCPs from epidermal fragments of tomato leaves. The protocol requires a two-step digestion to isolate high-quality tomato GCPs. In this procedure, cellulysin (in method L) was replaced by cellulose “Onozuka” RS (in method S) in the first digestion step, which indicated that cellulase RS was more effective than cellulysin. Method S dramatically shortened the time required for obtaining high yields and high-quality GCPs. Moreover, according to the GCP yields, hydroponic plants were more effective than substrate-cultured plants.ConclusionsIn this paper, protocols for large-scale preparation of GCPs and mesophyll cell protoplasts were described, followed by some success examples of their use in biochemical and molecular approaches such as reverse-transcription polymerase chain reaction, real-time polymerase chain reaction and sodium dodecyl sulfate–polyacrylamide gel electrophoresis. The method was proved to be a more efficient GCP-isolating method, capable of providing high yields with better quality in less time.Electronic supplementary materialThe online version of this article (10.1186/s13007-018-0294-7) contains supplementary material, which is available to authorized users.
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