Identification And Functional Analysis Of Tonoplast Sugar Transporters During Cold-induced Sweetening In Potato

Potatoes are increasingly consumed in the form of processed foods such as French fries and crisps.Potatoes need to be kept at a low temperature after harvest to prevent sprouting during storage for round-the-year processing.However,reducing sugars(RS)are accumulated in tubers at low temperatures,and this phenomenon refers to as coldinduced sweetening(CIS).Cold-induced sweetening(CIS),which results in a dark color,bitter taste,and increased acrylamide content in processed products,is a serious problem in the industry of potato processing.This is because the reducing sugar(RS)accumulating in the vacuole undergoes a Maillard reaction during processing at a high temperature.However,the transporter protein responsible for sugar loading into the cell vacuole of the tubers during CIS has not been identified yet.According to the current studies,three tonoplast sugar transporters(TSTs),StTST1,StTST3.1,and StTST3.2,were isolated in the potato plants.The results obtained from q RT-PCR indicated that StTSTs possessed distinct expression patterns at the transcript level in various potato tissues.Subcellular localization analysis showed that all these StTSTs targeted the tonoplast.Compared with StTST3.1 and StTST3.2,the knockdown of StTST1 caused light-color chips,suggesting StTST1 plays a critical role in CIS.Further research demonstrated that the silence of StTST1 could prevent the accumulation of RS in cold-stored tubers.It also revealed that the acrylamide content in potato chips processed from StTST1-silenced tubers decreased dramatically compared to the unsilenced control after tubers were stored at 4°C for 60 d.In the same case,potato chips processed from StTST1-silenced tubers performed lighter color.Transcriptome analysis manifested that the suppression of StTST1 resulted in the redirection of starch metabolism in cold-stored tubers,likely due to the suppression of tonoplast sugar transporter activity,ultimately resulting in sucrose retention in the cytosol.Further studies established that the increased sucrose concentration in the StTST1-silenced tubers correlated with down-regulation of β-amylase StBAM1,which might be due to decreasing the ABA content and inhibiting the ABA-signaling pathway.Thus,the suppression of StTST1 could be a novel effective approach for improving the processing quality of tubers as well as decreasing the acrylamide content in processed products of potato.In addition,two tonoplast sugar transporter 3-type isoforms,StTST3.1 and StTST3.2,were also identified in potatoes.In between them,StTST3.2 was found as the expressed TST3-type isoform in tubers.The silencing of StTST3.2 caused the significantly lower content of RS in processed products of potato tubers at harvest or after storage at room temperature,suggesting StTST3.2 plays an essential role in the accumulation of RS in tubers.Accordingly,in the same case,potato chips processed from StTST3.2-silenced tubers showed lighter color and caused a dramatic decrease in acrylamide production compared to the unsilenced control.Thus,it was also demonstrated that the suppression of StTST3.2 has no significant effect on potato growth and development.As a result,the suppression of StTST3.2 could be another effective approach for improving processing quality and decreasing acrylamide content in processed products of potato tubers.It can be concluded that the suppression of StTST1 contributes to reducing the effects of CIS and can process light-colored chips from the cold-stored tubers.In contrast,the suppression of StTST3.2 only improves the processing quality of tubers after storage at room temperature.In addition,suppression of StTST1 and StTST3.2 reduces the risk of health hazards by decreasing the acrylamide content in the processed products of potato tubers.