Conservation of noncoding microsatellites in plants: implication for gene regulation

BACKGROUND: Microsatellites are extremely common in plant genomes, and in particular, they are significantly enriched in the 5' noncoding regions. Although some 5' noncoding microsatellites involved in gene regulation have been described, the general properties of microsatellites as regulatory elements are still unknown. To address the question of microsatellites associated with regulatory elements, we have analyzed the conserved noncoding microsatellite sequences (CNMSs) in the 5' noncoding regions by inter- and intragenomic phylogenetic footprinting in the Arabidopsis and Brassica genomes. RESULTS: We identified 247 Arabidopsis-Brassica orthologous and 122 Arabidopsis paralogous CNMSs, representing 491 CT/GA and CTT/GAA repeats, which accounted for 10.6% of these types located in the 500-bp regions upstream of coding sequences in the Arabidopsis genome. Among these identified CNMSs, 18 microsatellites show high conservation in the regulatory regions of both orthologous and paralogous genes, and some of them also appear in the corresponding positions of more distant homologs in Arabidopsis, as well as in other plants. A computational scan of CNMSs for known cis-regulatory elements showed that light responsive elements were clustered in the region of CT/GA repeats, as well as salicylic acid responsive elements in the (CTT)(n)/(GAA)(n )sequences. Patterns of gene expression revealed that 70–80% of CNMS (CTT)(n)/(GAA)(n )associated genes were regulated by salicylic acid, which was consistent with the prediction of regulatory elements in silico. CONCLUSION: Our analyses showed that some noncoding microsatellites were conserved in plants and appeared to be ancient. These CNMSs served as regulatory elements involved in light and salicylic acid responses. Our findings might have implications in the common features of the over-represented microsatellites for gene regulation in plant-specific pathways

An L1 box binding protein, GbML1, interacts with GbMYB25 to control cotton fibre development

Transcription factors play key roles in plant development through their interaction with cis-elements and/or other transcription factors. A HD-Zip IV family transcription factor, Gossypium barbadense Meristem Layer 1 (GbML1) has been identified and characterized here. GbML1 specifically bound to the L1 box and the promoters of GbML1 and GbRDL1. GbML1 physically interacted with a key regulator of cotton fibre development, GbMYB25. Truncated and point mutation assays indicated the START–SAD domain was required for the binding to the C terminal domain (CTD) of GbMYB25. GbML1 overexpression in Arabidopsis increased the number of trichomes on stems and leaves and increased the accumulation of anthocyanin in leaves. Taken together, the L1 box binding protein, GbML1 was identified as the first partner for GbMYB25 and the role of START domain was discovered to be a protein binding domain in plants. Our findings will help the improvement of cotton fibre production and the understanding of the key role of HD-Zip family and MYB family in plants

Identification of Gene Modules Associated with Drought Response in Rice by Network-Based Analysis

Understanding the molecular mechanisms that underlie plant responses to drought stress is challenging due to the complex interplay of numerous different genes. Here, we used network-based gene clustering to uncover the relationships between drought-responsive genes from large microarray datasets. We identified 2,607 rice genes that showed significant changes in gene expression under drought stress; 1,392 genes were highly intercorrelated to form 15 gene modules. These drought-responsive gene modules are biologically plausible, with enrichments for genes in common functional categories, stress response changes, tissue-specific expression and transcription factor binding sites. We observed that a gene module (referred to as module 4) consisting of 134 genes was significantly associated with drought response in both drought-tolerant and drought-sensitive rice varieties. This module is enriched for genes involved in controlling the response of the plant to water and embryonic development, including a heat shock transcription factor as the key regulator in the expression of ABRE-containing genes. These results suggest that module 4 is highly conserved in the ABA-mediated drought response pathway in different rice varieties. Moreover, our study showed that many hub genes clustered in rice chromosomes had significant associations with QTLs for drought stress tolerance. The relationship between hub gene clusters and drought tolerance QTLs may provide a key to understand the genetic basis of drought tolerance in rice

Isolation and functional expression of a novel lipase gene isolated directly from oil-contaminated soil

A lipase gene SR1 encoding an extracellular lipase was isolated from oil-contaminated soil and expressed in Escherichia coli. The gene contained a 1845-bp reading frame and encoded a 615-amino-acid lipase protein. The mature part of the lipase was expressed with an N-terminal histidine tag in E. coli BL21, purified and characterized biochemically. The results showed that the purified lipase combines the properties of Pseudomonas chlororaphis and other Serratia lipases characterized so far. Its optimum pH and temperature for hydrolysis activity was pH 5.5-8.0 and 37°C respectively. The enzyme showed high preference for short chain substrates (556.3±2.8 U/µg for C10 fatty acid oil) and surprisingly it also displayed high activity for long-chain fatty acid. The deduced lipase SR1 protein is probably from Serratia, and is organized as a prepro-protein and belongs to the GXSXG lipase family

A Cotton Annexin Protein AnxGb6 Regulates Fiber Elongation through Its Interaction with Actin 1

<div><p>Annexins are assumed to be involved in regulating cotton fiber elongation, but direct evidence remains to be presented. Here we cloned six Annexin genes (<i>AnxGb</i>) abundantly expressed in fiber from sea-island cotton (<i>G. barbadense</i>). qRT-PCR results indicated that all six <i>G. barbadense</i> annexin genes were expressed in elongating cotton fibers, while only the expression of <i>AnxGb6</i> was cotton fiber-specific. Yeast two hybridization and BiFC analysis revealed that AnxGb6 homodimer interacted with a cotton fiber specific actin GbAct1. Ectopic-expressed <i>AnxGb6</i> in <i>Arabidopsis</i> enhanced its root elongation without increasing the root cell number. Ectopic <i>AnxGb6</i> expression resulted in more F-actin accumulation in the basal part of the root cell elongation zone. Analysis of <i>AnxGb6</i> expression in three cotton genotypes with different fiber length confirmed that <i>AnxGb6</i> expression was correlated to cotton fiber length, especially fiber elongation rate. Our results demonstrated that AnxGb6 was important for fiber elongation by potentially providing a domain for F-actin organization.</p></div

Induced <i>GbSBT1</i> expression following <i>V</i>. <i>dahliae</i> V991 infection.

<p>(A) Expression analysis of the <i>GbSBT1</i> gene in vegetative tissues (roots, stems, and leaves) via qPCR. The <i>Ubiquitin</i> gene is used as an internal control. At least three biological replicates were performed. (B) Expression analysis of <i>GbSBT1</i> in roots following <i>V</i>. <i>dahliae</i> strain V991 infection. The comparative CT method is adopted, and the expression is normalized. Each sample was repeated at least thrice. Error bars represent SE. Double asterisks represent a significant expression change of <i>GbSBT1</i> compared to uninfected condition (P < 0.01) in t-test.</p

Multiple sequence alignment analysis of GbSBT1 and other plant subtilases.

<p>GbSBT1 (KT336228, <i>Gossypium barbadense</i>), subtilase-like (XP_007017870, <i>Theobroma cacao</i>), SBT5.2 (NP_564107, <i>Arabidopsis thaliana</i>), SBT-like (AAM65424, <i>A</i>. <i>thaliana</i>). SP: signal peptide; Inhibitor I9: serine protease inhibitor domain; black triangles indicate conserved active sites; blank triangles indicate catalytic triads of serine protease; and blank box A indicates PA/protease or protease-like domain interface.</p

Comparison of F-actin organization in fiber cells between Pima-90, Coker312 and T586 Plants.

<p>A-C: Fiber cells at +3 DPA. A: Pima-90, B: Coker312, and C: T586. Note the length of fiber is different in the three varieties at the same stages. D: Fiber cells at +6 DPA and E: Fiber cells at +9 DPA. Bars: 10 µm in D and E.</p