Altered levels of histone deacetylase OsHDT1 affect differential gene expression patterns in hybrid rice

Hybrids between different inbred varieties display novel patterns of gene expression resulted from parental variation in allelic nucleotide sequences. To study the function of chromatin regulators in hybrid gene expression, the histone deacetylase gene OsHDT1 whose expression displayed a circadian rhythm was over-expressed or inactivated by RNAi in an elite rice parent. Increased OsHDT1 expression did not affect plant growth in the parent but led to early flowering in the hybrid. Nonadditive up-regulation of key flowering time genes was found to be related to flowering time of the hybrid. Over-expression of OsHDT1 repressed the nonadditive expression of the key flowering repressors in the hybrid (i.e. OsGI and Hd1) inducing early flowering. Analysis of histone acetylation suggested that OsHDT1 over-expression might promote deacetylation on OsGI and Hd1 chromatin during the peak expression phase. High throughput differential gene expression analysis revealed that altered OsHDT1 levels affected nonadditive expression of many genes in the hybrid. These data demonstrate that nonadditive gene expression was involved in flowering time control in the hybrid rice and that OsHDT1 level was important for nonadditive or differential expression of many genes including the flowering time genes, suggesting that OsHDT1 may be involved in epigenetic control of parental genome interaction for differential gene expression.     

Influence of mitochondria on gene expression in a citrus cybrid

The production of cybrids, combining nucleus of a species with alien cytoplasmic organelles, is a valuable method used for improvement of various crops. Several citrus cybrids have been created by somatic hybridization. These genotypes are interesting models to analyze the impact of cytoplasmic genome change on nuclear genome expression. Herein, we report genome-wide gene expression analysis in leaves of a citrus cybrid between C. reticulata cv ‘Willowleaf mandarin’ and C. limon cv ‘Eureka lemon’ compared with its lemon parent, using a Citrus 20K cDNA microarray. Molecular analysis showed that this cybrid possesses nuclear and chloroplast genomes of Eureka lemon plus mitochondria from Willowleaf mandarin and, therefore, can be considered as a lemon bearing foreign mitochondria. Mandarin mitochondria influenced the expression of a large set of lemon nuclear genes causing an over-expression of 480 of them and repression of 39 genes. Quantitative real-time RT–PCR further confirmed the credibility of microarray data. Genes over-expressed in cybrid leaves are predominantly attributed to the functional category “cellular protein metabolism” whereas in the down-regulated none functional category was enriched. Overall, mitochondria replacement affected different nuclear genes including particularly genes predicted to be involved in mitochondrial retrograde signaling. Mitochondria regulate all cell structures even chloroplast status. These results suggest that nuclear gene expression is modulated with respect to new information received from the foreign organelle, with the final objective to suit specific needs to ensure better cell physiological balance.     

Ultrabithorax is a regulator of beta 3 tubulin expression in the Drosophila visceral mesoderm.

beta 3 tubulin expression accompanies the specification and differentiation of the Drosophila mesoderm. The genetic programs involved in these processes are largely unknown. Our previous studies on the regulation of the beta 3 tubulin gene have shown that upstream sequences guide the expression in the somatic musculature, while regulatory elements in the first intron are necessary for expression in the visceral musculature. To further analyse this mode of regulation, which reflects an early embryonic specification program, we undertook a more detailed analysis of the regulatory capabilities of the intron. The results reveal not only a certain degree of redundancy in the cis-acting elements, which act at different developmental stages in the same mesodermal derivatives, but they also demonstrate in the visceral mesoderm, which forms a continuous epithelium along the body axis of the embryo, an early action of regulators guiding gene expression along the anterior-posterior axis of the embryo: an enhancer element in the intron leads to expression in a subdomain restricted along the anterior-posterior axis. This pattern is altered in mutants in the homeotic gene Ultrabithorax (Ubx), whereas ectopic Ubx expression leads to activity of the enhancer in the entire visceral mesoderm. So this element is likely to be a target of homeotic genes, which would define the beta 3 tubulin gene as a realisator gene under the control of selector genes.     

Cloning and expression of a beta tubulin gene of Physarum polycephalum

A beta tubulin gene of Physarum polycephalum has been isolated from a genomic library in the phage EMBL4. Southern-blot hybridization to genomic DNA indicates that the cloned DNA is derived from the betB1 locus of the beta tubulin gene family. A tubulin-specific subfragment of the phage DNA was used as a hybridization probe to construct a restriction map of the betB1 locus. The probe consisted of the almost complete coding region of the 5' half of the tubulin gene, interrupted by one intron. The derived amino acid sequence of this subclone deviates from the protein sequence for Physarum amoebal beta tubulin (amino acids 4-207) in two of 207 amino acids. We used both recA and recBC sbcB bacterial host strains, which have been recommended for cloning of instability-conferring sequences of the Physarum genome, but were unable to subclone the 3' part of the gene from the phage DNA. Primer-extension analysis indicates that the betB gene is expressed in the vegetatively proliferating amoebal and plasmodial stages of the life cycle as well as in differentiating (sporulating) plasmodia.     

Changes of gamma-tubulin expression and distribution in the zebrafish (Danio rerio) ovary, oocyte and embryo

The tubulin gene family is important for individual zebrafish development from the oocyte through to hatching. This involves often rapid, complex changes in the gametes and embryonic cells that are reflected in underlying gene expression changes. Tubulin dynamics, i.e., the interchange of polymeric and soluble forms in zebrafish oogenesis and embryogenesis, is important for microtubule (MT) cellular functions. Nevertheless, our understanding of how tubulin gene expression changes during zebrafish development is not clear. Previous data showed that soluble alpha-tubulin and gamma-tubulin are associated with large molecular weight complexes (>2MDa) which are reduced by the blastula stage, with a concomitant decrease in soluble tubulin amount. Complexes (<2MDa) then increased in the gastrula with an increase in soluble tubulin. Microarray revealed similar patterns of tubulin gene product expression for zebrafish ovary and eggs while both differed from day 4 larva. In situ hybridization with gamma-tubulin oligonucleotide probes revealed diffuse label in oocytes, with a marked localization to the primordial blastodisc upon maturation. These findings, together with recent work on gamma-tubulin ring complexes in other species, suggest that gamma-tubulin (protein complexes) may be involved in regulating tubulin dynamics, thus is important for zebrafish oogenesis and embryogenesis.     

Study of β-III tubulin expression in human eye tissues during prenatal development

The expression of β-III tubulin, a marker protein of early neuronal cells, was studied by molecular genetic and immunochemical techniques. The study was performed with the eyes of human fetuses on weeks 8.5 to 27–28 of intrauterine development. Expression of β-III tubulin was detected immunochemically in the retina and lens fibers of fetuses at stages of 8.5 to 22–23 weeks. By means of PCR, a strong expression of the β-III tubulin gene was revealed in the retina of 9.5-week fetuses. Its level remained high until week 18, became slightly lower by week 24, and decreased to zero by week 27–28. The expression of this gene was also revealed in the lens of 9.5-week fetuses. Its level at stages of 15 to 24 weeks was very low, and no expression could be detected in 27-to 28-week fetuses. The results of PCR analysis were consistent with immunochemical data.     

水稻XIOsPR10基因表达调控的初步研究

为探索水稻PR10蛋白在水稻抗细菌性条斑病中的作用,构建了XIOsPR10基因的植物过量表达载体p1301-XIOsPR10及RNAi表达载体pDS1301-XIOsPR10,通过农杆菌介导分别转化水稻愈伤组织,获得了相应的再生植株.经GUS检测和PCR分析,证实XIOsPR10基因以及RNAi片段分别整合到水稻再生植株基因组中;半定量RT-PCR分析显示,过量表达植株中XIOsPR10基因的表达量高于对照,而RNAi转基因植株中XIOsPR10基因的表达被抑制.     

Effect of delta-opioid receptor over-expression on cortical expression of GABAA receptor alpha1-subunit in hypoxia

Recent studies show that both delta-opioid receptors (DOR) and GABA receptors play a neuroprotective role in the mature cortex. Since we have observed that DOR over-expression renders the cortex more tolerant to hypoxic stress, we asked whether DOR over-expression affects GABA receptors expression in the cortex under hypoxia. As the first step, we investigated the expression of GABAA receptor alpha1-subunit (GABAA Ralpha1, the most abundant alpha-subunit of GABA receptors in the adult brain) in the mouse cortex with transgenic DOR over-expression after hypoxia. The results showed that GABAA Ralpha1 expression was lower in the transgenic than wild-type cortex, suggesting that DOR overexpression induces an inhibitory effect on GABAA receptor expression. Hypoxia for 1-3 days significantly increased GABAA Ralpha1 expression in the wild-type cortex, which may be an adaptive strategy for protecting the cortex against hypoxic stress. Interestingly, such increase was not found in the transgenic cortex with DOR over-expression. This may represent an interactive regulation in the transgenic cortex to efficiently balance energy production and consumption for better adaptation to hypoxic environment. Since DOR over-expression increases cortical tolerance to hypoxia, an increase in GABA receptors expression (an energy-costing process) may not be necessary in the cortex with DOR over-expression.     

Clustering formal concepts to discover biologically relevant knowledge from gene expression data

The production of high-throughput gene expression data has generated a crucial need for bioinformatics tools to generate biologically interesting hypotheses. Whereas many tools are available for extracting global patterns, less attention has been focused on local pattern discovery. We propose here an original way to discover knowledge from gene expression data by means of the so-called formal concepts which hold in derived Boolean gene expression datasets. We first encoded the over-expression properties of genes in human cells using human SAGE data. It has given rise to a Boolean matrix from which we extracted the complete collection of formal concepts, i.e., all the largest sets of over-expressed genes associated to a largest set of biological situations in which their over-expression is observed. Complete collections of such patterns tend to be huge. Since their interpretation is a time-consuming task, we propose a new method to rapidly visualize clusters of formal concepts. This designates a reasonable number of Quasi-Synexpression-Groups (QSGs) for further analysis. The interest of our approach is illustrated using human SAGE data and interpreting one of the extracted QSGs. The assessment of its biological relevancy leads to the formulation of both previously proposed and new biological hypotheses.     

Cloning and expression of the tubulin genes in barley

Tubulins are encoded by small gene families in plants. Based on the barley EST collection, cDNAs for alpha-, beta-, and gamma-tubulins were selected. Five genes for alpha-tubulin, eight newly identified beta-tubulin sequences and one gamma-tubulin gene were found to be expressed in barley. In silico analysis of relative abundance of the distinct tubulin sequences among ESTs derived from different libraries revealed that the various tubulin genes differed in their level of expression, and to some extent were tissue specific.     

Ovarian tumor expression is dependent on the functions of the somatic sex regulatory genes transformer-2 and doublesex

The doublesex-dependent sex regulatory pathway in Drosophila controls major aspects of somatic sexual differentiation, but its expression is not required in the X/X germline. Nevertheless, mutations in doublesex and in the genes that directly regulate its expression, transformer and transformer-2, disrupt early stages of oogenic differentiation to produce gonads containing immature germ cells. This indicates a critical, but uncharacterized, set of soma-germline interactions essential for oogenesis. In this paper, we examined the effects of mutations in transformer-2 on the expression and function of the germline-specific ovarian tumor gene. We demonstrated that in transformer-2 mutants, there was a marked reduction in the activity of the ovarian tumor promoter in the mutant germline. In addition, the phenotypic effects on the arrested germline could be partially suppressed by the simultaneous over-expression of both ovarian tumor and a second germline gene, Sex-lethal. This differs from transformer mutations, in which the over-expression of ovarian tumor alone is sufficient for a similar improvement in germline differentiation. In contrast to transformer-2, doublesex activity was not required for ovarian tumor promoter activity and we found indirect evidence that the doublesex male-specific function might have a negative regulatory effect. These data indicate that the components of the genetic pathway regulating somatic sexual differentiation have specific and differential effects on germline gene activity.     

Inducible gene expression in transgenic Xenopus embryos

The amphibian Xenopus laevis has been successfully used for many years as a model system for studying vertebrate development. Because of technical limitations, however, molecular investigations have mainly concentrated on early stages. We have developed a straightforward method for stage-specific induction of gene expression in transgenic Xenopus embryos [1] [2]. This method is based on the Xenopus heat shock protein 70 (Xhsp70 [3]) promoter driving the expression of desired gene products. We found that ubiquitous expression of the transgene is induced upon relatively mild heat treatment. Green fluorescent protein (GFP) was used as a marker to monitor successful induction of gene expression in transgenic embryos. We used this method to study the stage specificity of Wnt signalling function. Transient ectopic Wnt-8 expression during early neurulation was sufficient to repress anterior head development and this capacity was restricted to early stages of neurulation. By transient over-expression at different stages of development, we show that frizzled-7 disrupted morphogenesis sequentially from anterior to posterior along the dorsal axis as development proceeds. These results demonstrate that this method for inducible gene expression in transgenic Xenopus embryos will be a very powerful tool for temporal analysis of gene function and for studying molecular mechanisms of vertebrate organogenesis.     

The tubulin genes of Trypanosoma cruzi

The organization of the alpha- and beta-tubulin genes in the genome of Trypanosoma cruzi have been analysed by Southern blotting using tubulin probes derived from Trypanosoma brucei. The tubulin array appears to be more complex in this organism than in other members of the same family. Some tubulin genes are tightly clustered in an alternating (alpha-beta)n array with a basic repeat unit length of 4.3 kb. However, other pairs of alternating alpha- and beta-tubulin sequences appear to be physically separated from the basic group. This finding indicates that the tubulin gene cluster present in T. cruzi is less perfectly conserved than in T. brucei. T. (Herpetosoma) rangeli is similar to T. (Schizotrypanum) cruzi in its tubulin gene organization whereas most of these genes are tandemly clustered in the genome of T. (Trypanozoon) evansi, with a basic repeat unit length of 3.6 kb as previously described for T. (Trypanozoon) brucei. Two overlapping recombinant clones containing T. cruzi tubulin sequences have been isolated from a genomic cosmid library of T. cruzi epimastigotes using the T. brucei tubulin probes. Partial sequencing of the T. cruzi beta-tubulin gene has confirmed its identity and shows more than 70% homology with the sea urchin, chicken and T. b. rhodesiense beta-tubulin reported gene sequences. Analysis of tubulin gene organization through the parasite life cycle does not show evidence of major rearrangements within the repeat unit. Several T. cruzi strains and cloned lines whilst sharing the 4.3-kb tubulin repeat unit, exhibited very variable tubulin gene organization with tubulin probes. These striking differences in the organization of this structural gene among T. cruzi strains and cloned lines suggest that the heterogeneity previously reported in parasite populations may be related to a very dynamic, diploid genome.