杧果NFU家族基因的克隆、鉴定及其对非生物胁迫的响应分析

Cloning and Identification of NFU Family Genes in Mango and Responsive Analysis to Abiotic Stresses

从二倍体杧果‘桂热82’中克隆和鉴定了4个Fe-S簇装配所需的NFU支架蛋白编码基因,命名为MiNFU1~MiNFU4。MiNFU2和MiNFU3拥有全部3个NFU蛋白典型的Motif基序,MiNFU1缺失Motif 3而MiNFU4缺失Motif 1和Motif 2;MiNFU1~MiNFU3拥有相似的三级结构,而MiNFU4的三级结构与其差异较大,极为独特;13种不同科属植物之间的NFU家族成员数目略有差异,拟南芥、盐芥和短柄草3种一年生草本植物基因组中含有更多的支架蛋白,而非功能性支架蛋白在杧果、草莓、桃、苹果、柑橘和葡萄等多年生果树作物中更易发生丢失;植物NFU同源蛋白在系统发育树上可以分为2个亚家族,且在遗传进化关系上差异较大,同为十字花科、禾本科或蔷薇科植物的NFU同源蛋白分别倾向于紧密聚在一起,杧果NFU蛋白与柑橘和番茄相应的同源蛋白紧密聚在一起;杧果NFU家族基因在不同组织/器官中的表达水平差异显著,MiNFU2在所有检测组织中的整体表达表达水平最高,其次是MiNFU4和MiNFU1,而MiNFU3整体表达水平最低,杧果NFU家族基因均在幼苗叶片中的表达量最高,其次是韧皮部和盛开期花朵,在果实(幼果和熟果)中的表达水平相对较低;杧果NFU家族基因在转录水平对缺铁、高铁毒害、NaCl、PEG和低温处理的响应差异明显,虽然MiNFU3的整体表达最低,但却不受5种非生物胁迫的影响,MiNFU1和MiNFU4对高铁处理最为敏感,其表达水平在‘桂热82’幼苗全身组织均受高铁处理的调控而显著增加。此外,NaCl处理显著增加MiNFU1在根部的表达量,缺铁和PEG处理倾向于降低根部响应的NFU基因的表达水平,而低温处理倾向于降低叶片中响应的NFU基因的表达水平。本研究为明确杧果Fe-S簇装配分子机制提供基因资源,并为解析热带作物果树铁素营养和铁代谢奠定理论基础。

Four NFU family genes that involved in Fe-S cluster assembly were isolated and identified from diploid mango‘Guire82’,which were entitled MiNFU1 to MiNFU4,respectively.MiNFU2 and MiNFU3 proteins had all three typical motifs of NFU family proteins,while MiNFU1 lacks motif 3 and MiNFU4 lacks motif 1 and motif 2.MiNFU1 to MiNFU3 had a similar tertiary structure,while the tertiary structure of MiNFU4 was quite unique and different.There are slight differences in NFU family members among13 different plant families and genera.Three annual herbs of Arabidopsis,Brachypodium,and Thellungiella contain more scaffold proteins,while non-functional scaffold proteins are more likely to be lost in perennial fruit crops,such as mango,strawberry,peach,apple,citrus and grape.According to the phylogenetic tree,plant NFU homologous proteins can be divided into two subfamilies,which have differences in genetic and evolutionary relationships.Among them,NFU homologous proteins from Cruciferae,Gramineae or Rosaceae tend to be closely clustered,respectively,and mango NFU proteins are closely clustered with the corresponding homologs of citrus and tomato.In addition,there were significant differences in the expression levels of NFU family genes in different tissues/organs of mango.The overall expression level of MiNFU2 was the highest in all tested tissues,followed by MiNFU4 and MiNFU1,while the overall expression level of MiNFU3 was the lowest.The expression levels of NFU family genes in seedling leaves were the highest,followed by phloem and full blooming flowers,while the expression levels of NFU family genes in fruits(young and mature fruits)was relatively low.Notably,the transcriptional responses of NFU family genes in mango seedlings to iron depletion,high iron toxicity,NaCl,PEG and low temperature treatments were significantly different.Although the overall expression of MiNFU3 was the lowest,it was not affected by all tested abiotic stresses.MiNFU1 and MiNFU4 were more sensitive to high iron treatment,whose expression levels were significantly increased throughout the whole plant under high iron treatment.In addition,NaCl treatment significantly increased the expression of MiNFU1 in roots.Iron deficiency and PEG treatment tended to reduce the expression levels of responsive NFU genes in roots,while low temperature treatment were prone to reduce the expression levels of responsive NFU genes in leaves.This study provides gene resources to elucidate the molecular mechanisms of Fe-S cluster assembly in mango,and lays a theoretical foundation to reveal Fe nutrition and metabolism in tropical fruit crops.