Functional Analysis Of Cotton (Gossypium Hirsutum) BZR1 In Fiber Development

The cotton fibers are the most important natural sources of raw materials for the worldwide textile industry, studies about molecular mechanism in the development of fiber have a guiding significance to improve the cotton fiber quality. A large number of genes and proteins cooperate, which regulates the developmental processes in fiber. At the same time, this kind of activity is also affected by the endogenous hormones and environment. The plant hormones are a kind of small molecules, but have remarkable functions which affect the growth of plant. Brassinosteroids (BRs) are a kind of essential hormone for the developmental processes in fiber, which can significantly increase the length of fiber. BRs regulate a series of developmental and physiological activities in cotton fiber through a key transcription factor named BZR1. In this study, we separated a GhBZR1 gene by screening a yeast cDNA library and researched the functions of GhBZR1 in the development of fiber in cotton. Thus, the results of this study provide novel insights into the BR signalling related to fiber development of cotton.1. The protein sequence and phylogenetic analysis of GhBZR1The homology between GhBZR1 and GrBZR1 is extremely close, and GhBZR1 shares an intimate relation with GsBZR1 and CsBZR1 in the homology. The BZR1 protein sequences among different species in plant are quite conservative, which contain a DUF822 super family, suggesting that BZR1 play a key role in the growth and development of fiber. GhBZR1 has a closer relationship with CsBZR1 and JcBZRl over evolutionary history, however, compared with AtBZRl, they do not belong to the same branch. In addition, different species of BZR1 proteins and BZR2/BES1 proteins belong to different branches, which indicate that they appeared at the different time over the history of evolution.2. GhBZR1 promotes the development of fiber in cottonIn order to explore the functions of GhBZR1 in the growth of fiber, we got GhBZR1 transgenic cottons by using genetic transformation system. PCR was used to select the real transgenic plants. By observing the surface of 1DPA ovules, we found that compared with wild type, the fiber cells of over expression cottons grown rapidly, on the contrary, the phenotype showed that the development of the fiber cells in the suppressed expression plants was a slow process. Statistical analysis demonstrated that the mature fiber of the over expression lines elongated to a large extent compared with wild type, while the length of fiber in the suppressed expression cotton decreased sharply. All the results provided direct evidences that GhBZR1 can promote the development of fiber in cotton.3. The interaction between GhBZRl and other proteinsTo further explore the role of GhBZR1 in promoting the development of fiber, we screened the 9DPA fiber cDNA library by using the yeast two hybrid system to find out some proteins which can interact with GhBZRl and further improve the molecular mechanism of the growth in fiber. Since GhBZRl is a transcription factor that has transcriptional activation, we changed the length of gene based on the protein structure and constructed fusion plasmids. Through assaying the self transcriptional activation, we found that GhBZR1 A and GhBZR1 C had no self transcriptional activation, which can be used in the library screening experiment. We obtained 133 clones that contained longer than 750bp fragments through choosing GhBZRlC as a bait protein. The next step is sequencing. The experiment is still ongoing.4. The transcript profiling analysis of transgenic cottonsTo further study the role of BR in fiber growth and the function of GhBZR1 in the development of cotton, we used transcript profiling to analysis transgenic cottons. 10DPA fiber of wild type, over expression and suppressed expression cottons were collected respectively. We extracted the RNA then sent to the company. The results revealed that GhBZR1 is a key transcription factor in BR signaling pathway and its level of expression lead to the changes of expression in a series of genes in downstream BR signaling pathway, these genes involved in many aspects of fiber growth and development.We used transcript profiling to identify 1010 BR-regulated cotton genes. Functional studies of selected genes showed that they not only involved in the main physiological process in the fiber development, but also related to the plant hormones signaling transduction processes. They play a central role in the plant cell elongation, expansion and synthesis of cell wall that are the main processes in fiber growth, which indicated that GhBZR1 is a significant transcription factor that mediates a wide range of developmental and physiological processes in cotton by influencing the expression of BR-regulated genes. What is more, we found a extensive crosstalk between BR and other hormonal signaling pathways. The whole plant hormonal signaling transduction pathway is a large independent and interconnected network. GhBZR1 can not only regulate the expression of downstream genes, but also have a feedback to the expression levels of key components in BR signal pathway. In addition, other plant hormonal signaling transductions are also affected by GhBZRl, some of the GhBZR1 target genes play key roles in other hormonal signaling pathways, some of them are also other hormonal signals target genes.Our research provides a good theoretical guidance for the development of fiber, and improves the functions of BR throughout the developmental process in cotton, as well as shows direct evidences of the crosstalk between BR and other plant hormones.