Brassinosteroids is a steroid hormone in the plasma membrane of plants. It has been proved that brassinosteroids not only aids in the growth of plants but has a hand in photosynthesis, abiotic and biotic stressors and a wide range of physiological, genetic and developmental aspects of plants. The importance of the study of brassinosteroids and its regulators give an insight into the developmental process of plants and the way plants can be engineered in order to help solve many biological and social issues in the world. It also helps with identifying what inhibits and regulates various pathways in the plasma membrane. In an article entitled A Proteomics Study of Brassinosteroids Response in Aribidopsis It is said "BR-deficient or -insensitive mutant plants exhibit a wide range of phenotypes, which include dwarfism, curly and dark green leaves, delayed flowering and senescence, reduced fertility, photomorphogenesis in the dark, and altered vascular development" (Deng et. al.2007). This shows the importance of Brassinosteroids in plants and how it directly links with many physical attributes of plants. The research done and paper written by Xeula Wang and Joanne Chory entitled, Brassinosteroids Regulate Dissociation of BKl1, a Negative Regulator of BRl1 signaling, from the Plasma Membrane, highlights the importance of Brassinosteroids but goes in-depth and describes the dissociation of BKl1 and its effects on BRl1 signaling. The study of BRl1 aides in hypothesizing that BR1l interacting protein is a negative regulator of Brassinosteroids signaling.
Brassinosteroids have a major receptor in the plant Arabidopsis made of Serine and Threonine called BRl1. BRl1 has been used in many studies on mutant plants that have lost function to investigate the purpose and specificity of receptors. In order to identify the factors that regulate BRl1 activity the researchers directly bind Brassinolide, a highly active brassinosteroids, to the outer domain of BRl1 which in turn activated an organic polymer called a homo-oligomer. Phosphate is then added to the C-terminus of the BRl1 which in return enhances the attraction of BRl1 to its co-receptor, BAK1, this serves as the control. BRl1 lacking the C-terminus is then found to be a more extremely active but is unable to be fully activated showing that other factors is needed to regulate BRl1 activity.
The signaling activity between the plasma membrane and transcriptional responses were also researched. In order to investigate proteins that interact with BRl1 in yeast from Arabidopsis shoot meristems were searched for. Transthyretin-like protein which is a negative regulator and a protein named At5g42750 (also called BKl1), which has an unknown function were found. These two proteins were said to interact with the inactive kinase BRl1, leading to the exploration of BKl1 function in brassinosteroids signaling. Initially, plants with beta-glucuronidase gene from the BKl1 promoter are used to observe the expression of BKl1 during the developmental stage of the Arabidopsis. It was found that BKl1 was found in the leaves, roots, flowers and other structures of the Arabidopsis, thus suggesting that BKl1 and BRl1 are coexisting in the same tissues. Secondly, RNA interference lines were made to inhibit BKl1 RNA levels. In the article the results were said to show "Quantitative reverse transcription polymerase chain reaction analysis indicated that the transcript level of BKl1 was significantly reduced in many RNAi lines, compared with the control line. The RNAi lines had longer hypocotyls than the control line grown in short days…, and the levels of BKl1transcripts are negatively correlated with hypocotyl length… " (Xeulu Wang and Joanne Chory 2006) This suggests that the levels of BK1l represses the brassinosteroids thus repressing growth and other physiological developments associated with this steroid hormone. However over exposure of BKl1 with YFP (yellow fluorescent protein) tags resulted in the development of dwarf plants. These plants have smaller statures, rounder leaves and late blooming of the flowers in contrast with the wild type.
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To detect if the growth inhibition was caused by the overexposure to BKl1 with YFP was because of BRl1 signals, a downstream biomarker, BES1, was phosphorylated and tested by an immunoblot analysis procedure (a sensitive assessment that detects and characterizes proteins). A huge amount of BES-1 was in the wild type where as it was almost undetectable in the BKl1 over exposure line. The sample was also treated with 0.1 microM of brassinolide for one hour. This caused the accumulation of BES-1 in the wild type but much less n the BKl1-YFP line insinuating that BRl1 suppresses BKl1 overexposure.
The researchers also tried to determine where in the channel BKl1 function. To determine this, the sub cell of the BKl1 protein in the plants was analyzed. The BKl1-YFp was located in the plasma membrane and cytosol in the root tip cells. The BKl1-yfp line was then grown on two mediums. One containing a reducing agent that lowers the levels of endogenous brassinosteroids served as the control, which enhanced the fluorescent signal of BKl1-YFP and another containing brassinolide, which reduced the plasma membrane of BKl1-YFP and after several minutes it was completely diminished. This suggests that the Brassinosteroids alters the sub cell location of BKl1-YFP.
To confirm that the plasma membrane association of BKl1 inhibits BRl1 signaling, the researchers made plants with genes transferred from other species harboring BKl1-YFP protein and attaching it to the plasma membrane using a N-terminal myristoylation site. As the researchers believed the plants exhibited a great number of dwarf phenotypes in relation to other plants that have BKl1-YFP in similar levels.
The results of the research show that BKl1 function early in the pathway of the plasma membrane were brassinosteroids signal. Through the studies and research we have found that Bkl1 and BRl1 interact with each other directly and BRl1 signal is thus suppressed.BKl1 is strictly restricted to the plasma membrane and binds to BRl1 which keeps BRl1 from interacting with BAk-1, its co-receptor. This shows that Brassinosteroids Regulate Dissociation of BKl1 confirming the hypothesis of the researchers.
Work Cited
Zhiping Deng‡, Xin Zhang§, Wenqiang Tang‡, Juan A. Oses-Prieto§, Nagi Suzuki§, Joshua M. Gendron‡¶â€-, Huanjing Chen‡, Shenheng Guan§, Robert J. Chalkley§, T. Kaye Peterman**, Alma L. Burlingame§ and Zhi-Yong Wang. 2007. A Proteomics Study of Brassinosteroids Response in Arabidopsis. Molecular & Cellular Proteomics (internet). (cited 2010 September 13); 6, 2058-2071. Available from: http://www.mcponline.org/content/6/12/2058.full
Brassinosteroids Regulate Dissociation of BKI1, a Negative Regulator of BRI1 Signaling, from the Plasma Membrane Xuelu Wang and Joanne Chory Science 25 August 2006 313: 1118-1122; published online 20 July 200620 July 2006
