Analysis of the PEBP gene family and identification of a novel FLOWERING LOCUS T orthologue in sugarcane.

Sugarcane (Saccharum spp.) is an important economic crop for both sugar and biomass, the yields of which are negatively affected by flowering. The molecular mechanisms controlling flowering in sugarcane are nevertheless poorly understood. RNA-seq data analysis and database searches have enabled a comprehensive description of the PEBP gene family in sugarcane. It is shown to consist of at least 13 FLOWERING LOCUS T (FT)-like genes, two MOTHER OF FT AND TFL (MFT)-like genes, and four TERMINAL FLOWER (TFL)-like genes. As expected, these genes all show very high homology to their corresponding genes in Sorghum, and also to FT-like, MFT-like, and TFL-like genes in maize, rice, and Arabidopsis. Functional analysis in Arabidopsis showed that the sugarcane ScFT3 gene can rescue the late flowering phenotype of the Arabidopsis ft-10 mutant, whereas ScFT5 cannot. High expression levels of ScFT3 in leaves of short day-induced sugarcane plants coincided with initial stages of floral induction in the shoot apical meristem as shown by histological analysis of meristem dissections. This suggests that ScFT3 is likely to play a role in floral induction in sugarcane; however, other sugarcane FT-like genes may also be involved in the flowering process.

Influence of Sugarcane Variety on Rhizosphere Microbiota Under Irrigated and Water-Limiting Conditions.

Drought is one of the main problems linked to climate change that is faced by agriculture, affecting various globally important crops, including sugarcane. Environmentally sustainable strategies have been sought to mitigate the effects of climate change on crops. Among them, the use of beneficial microorganisms offers a promising approach. However, it is still necessary to understand the mechanisms that regulate plant-microorganism interactions, in normal situations and under stress. In this work, the rhizosphere metagenomes of two sugarcane varieties, one resistant and the other susceptible to drought, were compared under normal conditions and under water-limiting conditions. The results showed that for the drought-resistant sugarcane variety, bacteria belonging to the order Sphingomonadales and the family Xanthomonadaceae presented increased activities in terms of mobility, colonization, and cell growth. In contrast, the rhizosphere associated with the drought-sensitive variety exhibited increases of bacteria belonging to the family Polyangiaceae, and the genus Streptomyces, with modifications in DNA metabolism and ribosome binding proteins. The results pointed to variation in the rhizosphere microbiota that was modulated by the host plant genotype, revealing potential bacterial candidates that could be recruited to assist plants during water-limiting conditions.

Diel oscillations in cell wall components and soluble sugars as a response to short-day in sugarcane (Saccharum sp.).

BACKGROUND: Sugarcane is a tropical crop that can accumulate high concentration of sucrose in the stem as a storage carbohydrate. For that reason, sugarcane accounts for approximately 75% of all the sugar produced in the world and has become the main sugar source to produce first-generation bioethanol in Brazil. Daily rhythms cause plants to adapt and coordinate their metabolism to achieve maximum photosynthesis and carbohydrate production throughout the day. Circadian rhythms arise from the interaction of an internal oscillator and external stimuli, whereas diel rhythms occur in response to a light-dark cycle. Diel signalling contributes to synchronizing circadian rhythms to photoperiods, and levels of carbohydrates oscillate in a diel fashion. Under regular photoperiods, they are synthesized during the daytime and consumed throughout the night as an energy reserve. However, short days can induce higher rates of synthesis during daytime and lower rates of consumption in the dark. Cell wall carbohydrates are also diurnally regulated, and it has been shown that celluloses, hemicelluloses and pectin are deposited/degraded at different times of the day. To assess the diel carbohydrate profile in young sugarcane plants, we measured soluble sugars and cell wall components along a time course in plants subjected either to a regular day or short day. RESULTS: Short-day influenced sucrose synthesis and cell wall components. In short-day a 44% increase in sucrose concentration was detected in the dark, but was stable during the day. Cellulose, hemicellulose and pectin also fluctuate within a 24 h interval when subjected to a short day. A 38% increase in leaf sheath cellulose was observed from the middle of the day to the first hour of the night. Leaf sheath pectin and hemicellulose also increased from the day to the night, while it decreased in leaves. CONCLUSIONS: The presented data show diurnal patterns of soluble sugar metabolism together with temporal regulation of cell wall metabolism for a short day, suggesting that diel signalling has a role in how sugarcane manages sugar accumulation and partitioning. Understanding cell wall synthesis/degradation dynamics may help to improve the yield of sugarcane.

Prospecting Plant Growth-Promoting Bacteria Isolated from the Rhizosphere of Sugarcane Under Drought Stress.

In the rhizosphere, the soil bacteria and the plants are closely related, with the plant-associated microbiota playing an important role in promoting plant growth under both normal and stress conditions. In this study, the cultivable bacteria in the sugarcane rhizosphere under different levels of drought stress were characterized and screened for plant growth activities. The results suggested that the microbial community associated with the sugarcane rhizosphere was strongly affected by drought, but some important genera of bacteria such as Arthrobacter, Pseudomonas, Microbacterium, and Bacillus remained present during the entire experiment, indicating the adaptability of these organisms and their importance in the rhizosphere community. Many isolates exhibited positive results for one or more plant growth activity, and they were also capable of growing under simulated drought stress, suggesting that the microorganisms isolated from the sugarcane rhizosphere could be explored for uses such as biofertilizers or biocontrol agents in agriculture.

ScGAI is a key regulator of culm development in sugarcane.

Sugarcane contributes more than 70% of sugar production and is the second largest feedstock for ethanol production globally. Since sugar accumulates in sugarcane culms, culm biomass and sucrose content are the most commercially important traits. Despite extensive breeding, progress in both cane yield and sugar content remains very slow in most countries. We hypothesize that manipulating the genetic elements controlling culm growth will alter source-sink regulation and help break down the yield barriers. In this study, we investigate the role of sugarcane ScGAI, an ortholog of SLR1/D8/RHT1/GAI, on culm development and source-sink regulation through a combination of molecular techniques and transgenic strategies. We show that ScGAI is a key molecular regulator of culm growth and development. Changing ScGAI activity created substantial culm growth and carbon allocation changes for structural molecules and storage. ScGAI regulates spatio-temporal growth of sugarcane culm and leaf by interacting with ScPIF3/PIF4 and ethylene signaling elements ScEIN3/ScEIL1, and its action appears to be regulated by SUMOylation in leaf but not in the culm. Collectively, the remarkable culm growth variation observed suggests that ScGAI could be used as an effective molecular breeding target for breaking the slow yield gain in sugarcane.

Genome-wide identification and characterization of tRNA-derived RNA fragments in land plants.

KEY MESSAGE: The manuscript by Alves et al. entitled "Genome-wide identification and characterization of tRNA-derived RNA fragments in land plants" describes the identification and characterization of tRNAderived sRNA fragments in plants. By combining bioinformatic analysis and genetic and molecular approaches, we show that tRF biogenesis does not rely on canonical microRNA/siRNA processing machinery (i.e., independent of DICER-LIKE proteins). Moreover, we provide evidences that the Arabidopsis S-like Ribonuclease 1 (RNS1) might be involved in the biogenesis of tRFs. Detailed analyses showed that plant tRFs are sorted into different types of ARGONAUTE proteins and that they have potential target candidate genes. Our work advances the understanding of the tRF biology in plants by providing evidences that plant and animal tRFs shared common features and raising the hypothesis that an interplay between tRFs and other sRNAs might be important to fine-tune gene expression and protein biosynthesis in plant cells. Small RNA (sRNA) fragments derived from tRNAs (3'-loop, 5'-loop, anti-codon loop), named tRFs, have been reported in several organisms, including humans and plants. Although they may interfere with gene expression, their biogenesis and biological functions in plants remain poorly understood. Here, we capitalized on small RNA sequencing data from distinct species such as Arabidopsis thaliana, Oryza sativa, and Physcomitrella patens to examine the diversity of plant tRFs and provide insight into their properties. In silico analyzes of 19 to 25-nt tRFs derived from 5' (tRF-5s) and 3'CCA (tRF-3s) tRNA loops in these three evolutionary distant species showed that they are conserved and their abundance did not correlate with the number of genomic copies of the parental tRNAs. Moreover, tRF-5 is the most abundant variant in all three species. In silico and in vivo expression analyses unraveled differential accumulation of tRFs in Arabidopsis tissues/organs, suggesting that they are not byproducts of tRNA degradation. We also verified that the biogenesis of most Arabidopsis 19-25 nt tRF-5s and tRF-3s is not primarily dependent on DICER-LIKE proteins, though they seem to be associated with ARGONAUTE proteins and have few potential targets. Finally, we provide evidence that Arabidopsis ribonuclease RNS1 might be involved in the processing and/or degradation of tRFs. Our data support the notion that an interplay between tRFs and other sRNAs might be important to fine tune gene expression and protein biosynthesis in plant cells.

Identification, classification and transcriptional profiles of dirigent domain-containing proteins in sugarcane.

Dirigent (DIR) proteins, encoded by DIR genes, are referred to as "dirigent" because they direct the outcome of the coupling of the monolignol coniferyl alcohol into (+) or (-) pinoresinol, the first intermediates in the enantiocomplementary pathways for lignan biosynthesis. DIR domain-containing or DIR-like proteins are, thus, termed for not having a clear characterization. A transcriptome- and genome-wide survey of DIR domain-containing proteins in sugarcane was carried out, in addition to phylogenetic, physicochemical and transcriptional analyses. A total of 120 non-redundant sequences containing the DIR domain were identified and classified into 64 groups according to phylogenetic and sequence alignment analyses. In silico analysis of transcript abundance showed that these sequences are expressed at low levels in leaves and genes in the same phylogenetic clade have similar expression patterns. Expression analysis of ShDIR1-like transcripts in the culm internodes of sugarcane demonstrates their abundance in mature internodes, their induction by nitrogen fertilization and their predominant expression in cells that have a lignified secondary cell wall, such as vascular bundles of young internodes and parenchymal cells of the pith of mature internodes. Due to the lack of information about the functional role of DIR in plants, a possible relationship is discussed between the ShDIR1-like transcriptional profile and cell wall development in parenchyma cells of sugarcane culm, which typically accumulates large amounts of sucrose. The number of genes encoding the DIR domain-containing proteins in sugarcane is intriguing and is an indication per se that these proteins may have an important metabolic role and thus deserve to be better studied.

Genome-Wide Survey of Genes Under Positive Selection in Avian Pathogenic Escherichia coli Strains.

The ability to obtain bacterial genomes from the same host has allowed for comparative studies that help in the understanding of the molecular evolution of specific pathotypes. Avian pathogenic Escherichia coli (APEC) is a group of extraintestinal strains responsible for causing colibacillosis in birds. APEC is also suggested to possess a role as a zoonotic agent. Despite its importance, APEC pathogenesis still has several cryptic pathogenic processes that need to be better understood. In this work, a genome-wide survey of eight APEC strains for genes with evidence of recombination revealed that ∼14% of the homologous groups evaluated present signs of recombination. Enrichment analyses revealed that nine Gene Ontology (GO) terms were significantly more represented in recombinant genes. Among these GO terms, several were noted to be ATP-related categories. The search for positive selection in these APEC genomes revealed 32 groups of homologous genes with evidence of positive selection. Among these groups, we found several related to cell metabolism, as well as several uncharacterized genes, beyond the well-known virulence factors ompC, lamB, waaW, waaL, and fliC. A GO term enrichment test showed a prevalence of terms related to bacterial cell contact with the external environment (e.g., viral entry into host cell, detection of virus, pore complex, bacterial-type flagellum filament C, and porin activity). Finally, the genes with evidence of positive selection were retrieved from genomes of non-APEC strains and tested as were done for APEC strains. The result revealed that none of the groups of genes presented evidence of positive selection, confirming that the analysis was effective in inferring positive selection for APEC and not for E. coli in general, which means that the study of the genes with evidence of positive selection identified in this study can contribute for the better understanding of APEC pathogenesis processes.

Sugarcane Serine Peptidase Inhibitors, Serine Peptidases, and Clp Protease System Subunits Associated with Sugarcane Borer (Diatraea saccharalis) Herbivory and Wounding.

Sugarcane's (Saccharum spp.) response to Diatraea saccharalis (F.) (Lepidoptera: (Crambidae) herbivory was investigated using a macroarray spotted with 248 sugarcane Expressed Sequence Tags (ESTs) encoding serine peptidase inhibitors, serine peptidases. and Clp protease system subunits. Our results showed that after nine hours of herbivory, 13 sugarcane genes were upregulated and nine were downregulated. Among the upregulated genes, nine were similar to serine peptidase inhibitors and four were similar to Bowman-Birk Inhibitors (BBIs). Phylogenetic analysis revealed that these sequences belong to a phylogenetic group of sugarcane BBIs that are potentially involved in plant defense against insect predation. The remaining four upregulated genes included serine peptidases and one homolog to the Arabidopsis AAA+ chaperone subunit ClpD, which is a member of the Clp protease system. Among the downregulated genes, five were homologous to serine peptidases and four were homologous to Arabidopsis Clp subunits (three homologous to Clp AAA+ chaperones and one to a ClpP-related ClpR subunit). Although the roles of serine peptidase inhibitors in plant defenses against herbivory have been extensively investigated, the roles of plant serine peptidases and the Clp protease system represent a new and underexplored field of study. The up- and downregulated D. saccharalis genes presented in this study may be candidate genes for the further investigation of the sugarcane response to herbivory.

Sugarcane genome sequencing by methylation filtration provides tools for genomic research in the genus Saccharum.

Many economically important crops have large and complex genomes that hamper their sequencing by standard methods such as whole genome shotgun (WGS). Large tracts of methylated repeats occur in plant genomes that are interspersed by hypomethylated gene-rich regions. Gene-enrichment strategies based on methylation profiles offer an alternative to sequencing repetitive genomes. Here, we have applied methyl filtration with McrBC endonuclease digestion to enrich for euchromatic regions in the sugarcane genome. To verify the efficiency of methylation filtration and the assembly quality of sequences submitted to gene-enrichment strategy, we have compared assemblies using methyl-filtered (MF) and unfiltered (UF) libraries. The use of methy filtration allowed a better assembly by filtering out 35% of the sugarcane genome and by producing 1.5× more scaffolds and 1.7× more assembled Mb in length compared with unfiltered dataset. The coverage of sorghum coding sequences (CDS) by MF scaffolds was at least 36% higher than by the use of UF scaffolds. Using MF technology, we increased by 134× the coverage of gene regions of the monoploid sugarcane genome. The MF reads assembled into scaffolds that covered all genes of the sugarcane bacterial artificial chromosomes (BACs), 97.2% of sugarcane expressed sequence tags (ESTs), 92.7% of sugarcane RNA-seq reads and 98.4% of sorghum protein sequences. Analysis of MF scaffolds from encoded enzymes of the sucrose/starch pathway discovered 291 single-nucleotide polymorphisms (SNPs) in the wild sugarcane species, S. spontaneum and S. officinarum. A large number of microRNA genes was also identified in the MF scaffolds. The information achieved by the MF dataset provides a valuable tool for genomic research in the genus Saccharum and for improvement of sugarcane as a biofuel crop.

Characterization of the core microbiota of the drainage and surrounding soil of a Brazilian copper mine.

The core microbiota of a neutral mine drainage and the surrounding high heavy metal content soil at a Brazilian copper mine were characterized by 16S rDNA pyrosequencing. The core microbiota of the drainage was dominated by the generalist genus Meiothermus. The soil samples contained a more heterogeneous bacterial community, with the presence of both generalist and specialist bacteria. Both environments supported mainly heterotrophic bacteria, including organisms resistant to heavy metals, although many of the bacterial groups identified remain poorly characterized. The results contribute to the understanding of bacterial communities in soils impacted by neutral mine drainage, for which information is scarce, and demonstrate that heavy metals can play an important role in shaping the microbial communities in mine environments.

Building the sugarcane genome for biotechnology and identifying evolutionary trends.

BACKGROUND: Sugarcane is the source of sugar in all tropical and subtropical countries and is becoming increasingly important for bio-based fuels. However, its large (10 Gb), polyploid, complex genome has hindered genome based breeding efforts. Here we release the largest and most diverse set of sugarcane genome sequences to date, as part of an on-going initiative to provide a sugarcane genomic information resource, with the ultimate goal of producing a gold standard genome. RESULTS: Three hundred and seventeen chiefly euchromatic BACs were sequenced. A reference set of one thousand four hundred manually-annotated protein-coding genes was generated. A small RNA collection and a RNA-seq library were used to explore expression patterns and the sRNA landscape. In the sucrose and starch metabolism pathway, 16 non-redundant enzyme-encoding genes were identified. One of the sucrose pathway genes, sucrose-6-phosphate phosphohydrolase, is duplicated in sugarcane and sorghum, but not in rice and maize. A diversity analysis of the s6pp duplication region revealed haplotype-structured sequence composition. Examination of hom(e)ologous loci indicate both sequence structural and sRNA landscape variation. A synteny analysis shows that the sugarcane genome has expanded relative to the sorghum genome, largely due to the presence of transposable elements and uncharacterized intergenic and intronic sequences. CONCLUSION: This release of sugarcane genomic sequences will advance our understanding of sugarcane genetics and contribute to the development of molecular tools for breeding purposes and gene discovery.

Lignification in sugarcane: biochemical characterization, gene discovery, and expression analysis in two genotypes contrasting for lignin content.

Sugarcane (Saccharum spp.) is currently one of the most efficient crops in the production of first-generation biofuels. However, the bagasse represents an additional abundant lignocellulosic resource that has the potential to increase the ethanol production per plant. To achieve a more efficient conversion of bagasse into ethanol, a better understanding of the main factors affecting biomass recalcitrance is needed. Because several studies have shown a negative effect of lignin on saccharification yield, the characterization of lignin biosynthesis, structure, and deposition in sugarcane is an important goal. Here, we present, to our knowledge, the first systematic study of lignin deposition during sugarcane stem development, using histological, biochemical, and transcriptional data derived from two sugarcane genotypes with contrasting lignin contents. Lignin amount and composition were determined in rind (outer) and pith (inner) tissues throughout stem development. In addition, the phenolic metabolome was analyzed by ultra-high-performance liquid chromatography-mass spectrometry, which allowed the identification of 35 compounds related to the phenylpropanoid pathway and monolignol biosynthesis. Furthermore, the Sugarcane EST Database was extensively surveyed to identify lignin biosynthetic gene homologs, and the expression of all identified genes during stem development was determined by quantitative reverse transcription-polymerase chain reaction. Our data provide, to our knowledge, the first in-depth characterization of lignin biosynthesis in sugarcane and form the baseline for the rational metabolic engineering of sugarcane feedstock for bioenergy purposes.

Differential gene expression in Acidithiobacillus ferrooxidans LR planktonic and attached cells in the presence of chalcopyrite.

Acidithiobacillus ferrooxidans is commonly used in bioleaching operations to recover copper from sulfide ores. It is commonly accepted that A. ferrooxidans attaches to mineral surfaces by means of extracellular polymeric substances (EPS), however the role of type IV pili and tight adherence genes in this process is poorly understood. Genes related to the formation of type IV pili and tight adherence were identified in the genome of the bacterium, and in this work, we show that A. ferrooxidans actively expresses these genes, as demonstrated by quantitative real-time PCR analysis using cells incubated with chalcopyrite for 2 h. Significant differences in gene expression were observed between planktonic and adhered cells, with the level of expression being much greater in planktonic cells. These results might indicate that planktonic cells can actively adhere to the substrate. A bioinformatics analysis of interaction networks of the tight adherence and type IV pilus assembly genes revealed a strong relationship between conjugation systems (tra operon) and regulatory systems (PilR, PilS).

Involvement of microRNA-related regulatory pathways in the glucose-mediated control of Arabidopsis early seedling development.

In plants, sugars such as glucose act as signalling molecules that promote changes in gene expression programmes that impact on growth and development. Recent evidence has revealed the potential importance of controlling mRNA decay in some aspects of glucose-mediated regulatory responses suggesting a role of microRNAs (miRNAs) in these responses. In order to get a better understanding of glucose-mediated development modulation involving miRNA-related regulatory pathways, early seedling development of mutants impaired in miRNA biogenesis (hyl1-2 and dcl1-11) and miRNA activity (ago1-25) was evaluated. All mutants exhibited a glucose hyposensitive phenotype from germination up to seedling establishment, indicating that miRNA regulatory pathways are involved in the glucose-mediated delay of early seedling development. The expression profile of 200 miRNA primary transcripts (pri-miRs) was evaluated by large-scale quantitative real-time PCR profiling, which revealed that 38 pri-miRs were regulated by glucose. For several of them, the corresponding mature miRNAs are known to participate directly or indirectly in plant development, and their accumulation was shown to be co-regulated with the pri-miR by glucose. Furthermore, the expression of several miRNA target genes was found to be deregulated in response to glucose in the miRNA machinery mutants ago1-25, dcl1-11, and hyl1-2. Also, in these mutants, glucose promoted misexpression of genes for the three abscisic acid signalling elements ABI3, ABI4, and ABI5. Thus, miRNA regulatory pathways play a role in the adjustments of growth and development triggered by glucose signalling.

Expression of SofLAC, a new laccase in sugarcane, restores lignin content but not S:G ratio of Arabidopsis lac17 mutant.

Lignin is a complex phenolic heteropolymer deposited in the secondarily thickened walls of specialized plant cells to provide strength for plants to stand upright and hydrophobicity to conducting cells for long-distance water transport. Although essential for plant growth and development, lignin is the major plant cell-wall component responsible for biomass recalcitrance to industrial processing. Peroxidases and laccases are generally thought to be responsible for lignin polymerization, but, given their broad substrate specificities and large gene families, specific isoforms involved in lignification are difficult to identify. This study used a combination of co-expression analysis, tissue/cell-type-specific expression analysis, and genetic complementation to correlate a sugarcane laccase gene, SofLAC, to the lignification process. A co-expression network constructed from 37 cDNA libraries showed that SofLAC was coordinately expressed with several phenylpropanoid biosynthesis genes. Tissue-specific expression analysis by quantitative RT-PCR showed that SofLAC was expressed preferentially in young internodes and that expression levels decrease with stem maturity. Cell-type-specific expression analysis by in situ hybridization demonstrated the localization of SofLAC mRNA in lignifying cell types, mainly in inner and outer portions of sclerenchymatic bundle sheaths. To investigate whether SofLAC is able to oxidize monolignols during lignification, the Arabidopsis lac17 mutant, which has reduced lignin levels, was complemented by expressing SofLAC under the control of the Arabidopsis AtLAC17 promoter. The expression of SofLAC restored the lignin content but not the lignin composition in complemented lac17 mutant lines. Taken together, these results suggest that SofLAC participates in lignification in sugarcane.

Global analysis of the sugarcane microtranscriptome reveals a unique composition of small RNAs associated with axillary bud outgrowth.

Axillary bud outgrowth determines shoot architecture and is under the control of endogenous hormones and a fine-tuned gene-expression network, which probably includes small RNAs (sRNAs). Although it is well known that sRNAs act broadly in plant development, our understanding about their roles in vegetative bud outgrowth remains limited. Moreover, the expression profiles of microRNAs (miRNAs) and their targets within axillary buds are largely unknown. Here, we employed sRNA next-generation sequencing as well as computational and gene-expression analysis to identify and quantify sRNAs and their targets in vegetative axillary buds of the biofuel crop sugarcane (Saccharum spp.). Computational analysis allowed the identification of 26 conserved miRNA families and two putative novel miRNAs, as well as a number of trans-acting small interfering RNAs. sRNAs associated with transposable elements and protein-encoding genes were similarly represented in both inactive and developing bud libraries. Conversely, sequencing and quantitative reverse transcription-PCR results revealed that specific miRNAs were differentially expressed in developing buds, and some correlated negatively with the expression of their targets at specific stages of axillary bud development. For instance, the expression patterns of miR159 and its target GAMYB suggested that they may play roles in regulating abscisic acid-signalling pathways during sugarcane bud outgrowth. Our work reveals, for the first time, differences in the composition and expression profiles of diverse sRNAs and targets between inactive and developing vegetative buds that, together with the endogenous balance of specific hormones, may be important in regulating axillary bud outgrowth.

Analysis of plant LTR-retrotransposons at the fine-scale family level reveals individual molecular patterns.

BACKGROUND: Sugarcane is an important crop worldwide for sugar production and increasingly, as a renewable energy source. Modern cultivars have polyploid, large complex genomes, with highly unequal contributions from ancestral genomes. Long Terminal Repeat retrotransposons (LTR-RTs) are the single largest components of most plant genomes and can substantially impact the genome in many ways. It is therefore crucial to understand their contribution to the genome and transcriptome, however a detailed study of LTR-RTs in sugarcane has not been previously carried out. RESULTS: Sixty complete LTR-RT elements were classified into 35 families within four Copia and three Gypsy lineages. Structurally, within lineages elements were similar, between lineages there were large size differences. FISH analysis resulted in the expected pattern of Gypsy/heterochromatin, Copia/euchromatin, but in two lineages there was localized clustering on some chromosomes. Analysis of related ESTs and RT-PCR showed transcriptional variation between tissues and families. Four distinct patterns were observed in sRNA mapping, the most unusual of which was that of Ale1, with very large numbers of 24nt sRNAs in the coding region. The results presented support the conclusion that distinct small RNA-regulated pathways in sugarcane target the lineages of LTR-RT elements. CONCLUSIONS: Individual LTR-RT sugarcane families have distinct structures, and transcriptional and regulatory signatures. Our results indicate that in sugarcane individual LTR-RT families have distinct behaviors and can potentially impact the genome in diverse ways. For instance, these transposable elements may affect nearby genes by generating a diverse set of small RNA's that trigger gene silencing mechanisms. There is also some evidence that ancestral genomes contribute significantly different element numbers from particular LTR-RT lineages to the modern sugarcane cultivar genome.

Gene expression modulation by heat stress in Acidithiobacillus ferrooxidans LR.

During bioleaching, Acidithiobacillus ferrooxidans is subjected to different types of stress, including heat stress, which affect bacterial growth. In this work, real time quantitative PCR was used to analyze the expression of heat shock genes, as well as genes that encode proteins related to several functional categories in A. ferrooxidans. Cells were submitted to long-term growth and heat shock, both at 40°C. The results showed that heat shock affected the expression levels of most genes investigated, whilst long-term growth at 40°C resulted in minor changes in gene expression, except for certain genes related to iron transport, which were strongly down-regulated, suggesting that the iron processing capability of A. ferrooxidans was affected by long-term growth at 40°C. A bioinformatic analysis of the genes' promoter regions indicated a putative transcriptional regulation by the σ(32) factor in 12 of the 31 genes investigated, suggesting the involvement of other regulatory mechanisms in the response of A. ferrooxidans to heat stress.