亚砷酸钠抑制柑橘果实柠檬酸积累的机制

Mechanism of sodium arsenite inhibiting citrate accumulation in citrus fruits

以亚砷酸钠(NaAsO_(2))溶液(含6.16 g/L MgSO4·7H2O、1 mL/L Triton X–100、0.17 g/L NaAsO_(2))喷施处理‘山下红’温州蜜柑、‘橘湘早’温州蜜柑、‘大红甜橙’树体,离体处理‘冰糖橙’汁胞(MS培养基含1.0 mg/L NaAsO_(2))及愈伤组织(MS培养基含0.5、1.0 mg/L NaAsO_(2)),测定可溶性固形物(TSS)、苹果酸和柠檬酸的含量,分析CsAN1和CsPH8基因的表达及其启动子活性。结果表明:NaAsO_(2)可有效降低果实、汁胞和愈伤组织的柠檬酸含量;NaAsO_(2)处理后,‘山下红’‘橘湘早’和‘大红甜橙’果实柠檬酸含量分别较对照降低4.59%~61.57%、0.67%~14.92%、13.22%~54.35%;NaAsO_(2)处理60 d的‘橘湘早’果实中CsAN1和CsPH8的表达量分别下降73%和32%,在NaAsO_(2)处理21 d的‘冰糖橙’汁胞中的表达量分别降低55%和27%,在0.5 mg/L NaAsO_(2)处理20 d的‘冰糖橙’愈伤组织中分别降低73%和84%;启动子区域缺失试验结果表明,干旱信号可能作用在CsAN1和CsPH8启动子的ABRE元件上,并调控它们的表达,NaAsO_(2)处理后含有ABRE元件的启动子区段活性明显减弱,推测NaAsO_(2)抑制了柑橘中CsAN1和CsPH8启动子活性进而降低其表达,从而减弱了柠檬酸在果实中的积累。

In order to investigate the mechanism of sodium arsenite inhibiting citric acid accumulation in citrus fruits,sodium arsenite solution(containing 6.16 g/L MgSO4·7H2O,1 mL/L Triton X-100,0.17 g/L NaAsO_(2))was applied by spraying onto‘Yamashitabeni wase’(Citrus unshiu Marc),‘Juxiangzao’(C.unshiu Marc)and‘Dahongtiancheng’(C.sinensis Osbeck),as well as to detached juice sacs(MS medium containing 1.0 mg/L NaAsO_(2))and callus of‘Bingtangcheng’(C.sinensis Osbeck)(MS medium containing 0.5 and 1.0 mg/L NaAsO_(2)).The total soluble solid(TSS),citrate and malate contents,expression levels of CsAN1 and CsPH8 genes,and their promoter activities were analyzed.The results showed that NaAsO_(2)could reduce the citrate contents of fruit,juice sacs and callus tissues.After NaAsO_(2)treatment,the citrate contents of‘Yamashitabeni wase’,‘Juxiangzao’and‘Dahongtiancheng’decreased by 4.59 to 61.57%,0.67%to 14.92%and 13.22%to 54.35%,respectively,compared with CK group.CsAN1 and CsPH8 expression levels were significantly reduced under NaAsO_(2)treatment,with a 73%and 32%respectively decrease in‘Juxiangzao’fruit after 60 days of treatment,a 55%and 27%respectively decrease in‘Bingtangcheng’juice sacs after 21 days,and 73%and 84%respectively in the‘Bingtangcheng’calli callus treated with 0.5 mg/L NaAsO_(2)for 20 days.The experiment under the absence of promoter subregions indicated that drought signals might act on the ABRE elements in the CsAN1 and CsPH8 promoters to regulate their expression,while the activity of promoter segments containing ABRE elements was significantly reduced after NaAsO_(2)treatment.It is speculated that NaAsO_(2)inhibits the activity of CsAN1 and CsPH8 promoters in citrus,thereby reducing their expression and weakening the accumulation of citric acid in the fruit.