Genetic Effect Analysis Of Tip41in Wheat(Triticum Aestivum L.),a Gene Affecting Plant Development

Common wheat (Triticum aestivum L.), a hexaploid species of grass family (AABBDD), is characterized by its large and complex genome (40times of the rice genome) and high contents of repeat sequences (85%), which lead to great difficulty in genome sequencing and isolation of genes. Based on association analysis of Chinese wheat core collection, we found that a SSR locus (Xgwm212-5D) is associated with important traits significantly, such as Plant Height (PH), Thousand Kernel Weight (TKW), and Spikelet Length (SL). Through sequencing analysis of positive Bacterial Artificial Chromosome (BAC) clone which the length is-120kb, screened out by Xgwm212in Chinese Spring BAC library, we found that Xgwm212is located between the seventh exon and seventh intron of a gene which named as TaTip41. It belongs to a class of highly conserved gene family across species. It is reported that Tip41is a key member of TOR (Target of Rapamycin) signaling pathway, participating in the regulation of nitrogen uptaking and utilization in yeast. However, it is still not clear about its function in plants.Subcellular localization analysis showed that TaTIP41is located on cytoplasm, nucleus and cell membrane both in onion epidermal cells and wheat protoplasts. The highest expression of TaTip41was observed in floral organs despite of its constitutive expression. In comparison with wild type, the Arabidopsis attip41mutants showed later flowering, longer siliques and more seeds. TaTip41-overexpressing lines exhibit earlier flowering phenotype, suggesting that the Tip41gene may be a positive regulator of plant development. Yeast two hybrid assays showed that TIP41either from Arabidopsis or wheat, can interact with TAP46correspondingly. Moreover, the expression of Tip41and Tap46is negatively correlated, and attip41mutant demonstrates high sensitivity to N-starvation stress. These results implied that TIP41might participate in modulating the TOR signaling pathway by affecting TAP46. Interestingly, TIP41can interact with B55β (A regulatory subunit of PP2A) to promote the expression of some positive regulators involved in the flowering such as SOC1and FT, and stimutanously suppress the expression of negative regulators like FLC, which contributes to regulate flowering time in Arabidopsis and wheat indirectly.The TaTip41of genomic sequences and cDNA were isolated in this study, which were mapped to homoeologous group5chromosomes via nulli-tetrasomic and mapping population. Hapotyping in262wheat mini-core collections indicated that TaTip41has an effect on Plant Height (PH), Heading Date (HD), Maturing Date (MD), and Thousand Kernel Weight (TKW). We deduced from phenotypic variation in elected introgression lines (Qiangweimai/Zhengmai366) that the favourable allelic variation (Hap-5D-II) demonstrates obvious positive effect on thousand kernel weight and Grain Number (GN).Silencing TaTip41by RNA interfering technology causes about50%-75%decline of total expression in TO transgenic lines. The coming work will focus on phenotypic analysis of these transgenic plants for providing experimental evidence to clarify the function of Tip41. In this study, we discovered one novel gene associated plant development, TaTip41, which located on5DL chromosomes in T. aestivum. Meanwhile, we investigated the genetic effect of TaTip41in combination with gene function identification. Our experimental evidence supported that Tip41is an important gene might be involved in development and nutrition metabolism. This study may provide new strategy on important gene isolation in wheat.