Repeated Cocaine Self-Administration Causes Multiple Changes in Rat Frontal Cortex Gene Expression

Repeated cocaine administration produces changes in gene expression that are thought to contribute to the behavioral alterations observed with cocaine abuse. This study focuses on gene expression changes in the frontal cortex, a component of reinforcement, sensory, associative, and executive circuitries. Changes in frontal cortex gene expression after repeated cocaine self-administration may lead to changes in the behaviors associated with this brain region. Rats self-administered cocaine for 10 days in a continuous access, discrete trial paradigm (averaging 100 mg/kg/day) and were examined for changes in relative frontal cortex mRNA abundance by cDNA hybridization arrays. Among the changes observed following array analysis, increased nerve-growth-factor–induced B (NGFI-B), adenylyl cyclase type VIII (AC VIII), and reduced cysteine-rich protein 2 (CRP2) mRNA were confirmed by quantitative RT-PCR. These changes share commonalities and exhibit differences with previous reports of gene expression changes in the frontal cortex after noncontingent cocaine administration.     

Clock Genes Influence Gene Expression in Growth Plate and Endochondral Ossification in Mice

We have previously shown transient promotion by parathyroid hormone of Period-1 (Per1) expression in cultured chondrocytes. Here we show the modulation by clock genes of chondrogenic differentiation through gene transactivation of the master regulator of chondrogenesis Indian hedgehog (IHH) in chondrocytes of the growth plate. Several clock genes were expressed with oscillatory rhythmicity in cultured chondrocytes and rib growth plate in mice, whereas chondrogenesis was markedly inhibited in stable transfectants of Per1 in chondrocytic ATDC5 cells and in rib growth plate chondrocytes from mice deficient of brain and muscle aryl hydrocarbon receptor nuclear translocator-like (BMAL1). Ihh promoter activity was regulated by different clock gene products, with clear circadian rhythmicity in expression profiles of Ihh in the growth plate. In BMAL1-null mice, a predominant decrease was seen in Ihh expression in the growth plate with a smaller body size than in wild-type mice. BMAL1 deficit led to disruption of the rhythmic expression profiles of both Per1 and Ihh in the growth plate. A clear rhythmicity was seen with Ihh expression in ATDC5 cells exposed to dexamethasone. In young mice defective of BMAL1 exclusively in chondrocytes, similar abnormalities were found in bone growth and Ihh expression. These results suggest that endochondral ossification is under the regulation of particular clock gene products expressed in chondrocytes during postnatal skeletogenesis through a mechanism relevant to the rhythmic Ihh expression.     

Alternate Bearing in Citrus: Changes in the Expression of Flowering Control Genes and in Global Gene Expression in ON- versus OFF-Crop Trees

Alternate bearing (AB) is the process in fruit trees by which cycles of heavy yield (ON crop) one year are followed by a light yield (OFF crop) the next. Heavy yield usually reduces flowering intensity the following year. Despite its agricultural importance, how the developing crop influences the following year''s return bloom and yield is not fully understood. It might be assumed that an ‘AB signal’ is generated in the fruit, or in another organ that senses fruit presence, and moves into the bud to determine its fate—flowering or vegetative growth. The bud then responds to fruit presence by altering regulatory and metabolic pathways. Determining these pathways, and when they are altered, might indicate the nature of this putative AB signal. We studied bud morphology, the expression of flowering control genes, and global gene expression in ON- and OFF-crop buds. In May, shortly after flowering and fruit set, OFF-crop buds were already significantly longer than ON-crop buds. The number of differentially expressed genes was higher in May than at the other tested time points. Processes differentially expressed between ON- and OFF-crop trees included key metabolic and regulatory pathways, such as photosynthesis and secondary metabolism. The expression of genes of trehalose metabolism and flavonoid metabolism was validated by nCounter technology, and the latter was confirmed by metabolomic analysis. Among genes induced in OFF-crop trees was one homologous to SQUAMOSA PROMOTER BINDING-LIKE (SPL), which controls juvenile-to-adult and annual phase transitions, regulated by miR156. The expression pattern of SPL-like, miR156 and other flowering control genes suggested that fruit load affects bud fate, and therefore development and metabolism, a relatively long time before the flowering induction period. Results shed light on some of the metabolic and regulatory processes that are altered in ON and OFF buds.     

Intrachromosomal Gene Conversion and the Maintenance of Sequence Homogeneity among Repeated Genes

Intrachromosomal gene conversion is the non-reciprocal transfer of information between a pair of repeated genes on a single chromosome. This process produces eventual sequence homogeneity within a family of repeated genes. An evolutionary model for a single chromosome lineage was formulated and analyzed. Expressions were derived for the fixation probability, mean time to fixation or loss, and mean conditional fixation time for a variant repeat with an arbitrary initial frequency. It was shown that a small conversional advantage or disadvantage for the variant repeat (higher or lower probability of producing two variant genes by conversion than two wild-type genes) can have a dramatic effect on the probability of fixation. The results imply that intrachromosomal gene conversion can act sufficiently rapidly to be an important mechanism for maintaining sequence homogeneity among repeated genes.     

From Description to Causality: Mechanisms of Gene Expression Signatures in Cancer

Global gene expression profiles of thousands of cancer samples have been completed, giving rise to hundreds of gene expression signatures (GES). Although many expression signatures show promise in predicting patient prognosis or response to therapies, the usefulness of the signatures in understanding the underlying mechanisms of cancer has not been fully exploited. While “reverse genomic” methods can test specific hypotheses of gene regulation, they fare less well in deciphering novel or combinatorial mechanisms of gene regulation. Recently we described SLAMS (stepwise linkage analysis of microarray signatures), a novel method that can prospectively identify genetic regulators of gene expression signatures in cancer. Applying SLAMS on a poor-prognosis wound signature in human breast cancer, we identified CSN5-mediated ubiquitination of MYC as a novel mechanism to activate a biological program favoring metastasis.     

From Genes to Flower Patterns and Evolution: Dynamic Models of Gene Regulatory Networks

Genes and proteins form complex dynamical systems or gene regulatory networks (GRN) that can reach several steady states (attractors). These may be associated with distinct cell types. In plants, the ABC combinatorial model establishes the necessary gene combinations for floral organ cell specification. We have developed dynamic gene regulatory network (GRN) models to understand how the combinatorial selection of gene activity is established during floral organ primordia specification as a result of the concerted action of ABC and non-ABC genes. Our analyses have shown that the floral organ specification GRN reaches six attractors with gene configurations observed in primordial cell types during early stages of flower development and four that correspond to regions of the inflorescence meristem. This suggests that it is the overall GRN dynamics rather than precise signals that underlie the ABC model. Furthermore, our analyses suggest that the steady states of the GRN are robust to random alterations of the logical functions that define the gene interactions. Here we have updated the GRN model and have systematically altered the outputs of all the logical functions and addressed in which cases the original attractors are recovered. We then reduced the original three-state GRN to a two-state (Boolean) GRN and performed the same systematic perturbation analysis. Interestingly, the Boolean GRN reaches the same number and type of attractors as reached by the three-state GRN, and it responds to perturbations in a qualitatively identical manner as the original GRN. These results suggest that a Boolean model is sufficient to capture the dynamical features of the floral network and provide additional support for the robustness of the floral GRN. These findings further support that the GRN model provides a dynamical explanation for the ABC model and that the floral GRN robustness could be behind the widespread conservation of the floral plan among eudicotyledoneous plants. Other aspects of evolution of flower organ arrangement and ABC gene expression patterns are discussed in the context of the approach proposed here. álvaro Chaos, Max Aldana and Elena Alvarez-Buylla contributed equally to this work.     

Increasing Maternal or Post-Weaning Folic Acid Alters Gene Expression and Moderately Changes Behavior in the Offspring

Background

Studies have indicated that altered maternal micronutrients and vitamins influence the development of newborns and altered nutrient exposure throughout the lifetime may have potential health effects and increased susceptibility to chronic diseases. In recent years, folic acid (FA) exposure has significantly increased as a result of mandatory FA fortification and supplementation during pregnancy. Since FA modulates DNA methylation and affects gene expression, we investigated whether the amount of FA ingested during gestation alters gene expression in the newborn cerebral hemisphere, and if the increased exposure to FA during gestation and throughout the lifetime alters behavior in C57BL/6J mice.

Methods

Dams were fed FA either at 0.4 mg or 4 mg/kg diet throughout the pregnancy and the resulting pups were maintained on the diet throughout experimentation. Newborn pups brain cerebral hemispheres were used for microarray analysis. To confirm alteration of several genes, quantitative RT-PCR (qRT-PCR) and Western blot analyses were performed. In addition, various behavior assessments were conducted on neonatal and adult offspring.

Results

Results from microarray analysis suggest that the higher dose of FA supplementation during gestation alters the expression of a number of genes in the newborns’ cerebral hemispheres, including many involved in development. QRT-PCR confirmed alterations of nine genes including down-regulation of Cpn2, Htr4, Zfp353, Vgll2 and up-regulation of Xist, Nkx6-3, Leprel1, Nfix, Slc17a7. The alterations in the expression of Slc17a7 and Vgll2 were confirmed at the protein level. Pups exposed to the higher dose of FA exhibited increased ultrasonic vocalizations, greater anxiety-like behavior and hyperactivity. These findings suggest that although FA plays a significant role in mammalian cellular machinery, there may be a loss of benefit from higher amounts of FA. Unregulated high FA supplementation during pregnancy and throughout the life course may have lasting effects, with alterations in brain development resulting in changes in behavior.     

Expression Analysis of Proline Metabolism-related Genes From Halophyte Arabis stelleri under Osmotic Stress Conditions

Arabis stelleri var.japonica evidenced stronger osmotic stress tolerance than Arabidopsis thaliana.Using an A.thaliana microarray chip,we determined changes in the expression of approximately 2 800genes between A.stelleri plants treated with 0.2 M mannitol versus mock-treated plants.The most significant changes in the gene expression patterns were in genes defining cellular components or in genes associated with the endomembrane system,stimulus response,stress response,chemical stimulus response,and defense response.The expression patterns of three de novo proline biosynthesis enzymes were evaluated in A.stelleri var.japonica seedlings treated with 0.2 M mannitol,0.2 M sorbitol,and 0.2 M NaCl.The expression of Δ1-pyrroline-5-carboxylate synthetase was not affected by NaCl stress but was similarly induced by mannitol and sorbitol.The proline dehydrogenase gene,which is known to be repressed by dehydration stress and induced by free L-proline,was induced at an early stage by mannitol treatment,but the level of proline dehydrogenase was increased later by treatment with both mannitol and NaCl.The level of free L-proline accumulation increased progressively in response to treatments with mannitol,sorbitol,and NaCl.Mannitol induced L-proline accumulation more rapidly than NaCl or sorbitol.These findings demonstrate that the osmotic tolerance of the novel halophyte,Arabis stelleri,is associated with the accumulation of L-proline.     

Hashimoto's Thyroiditis: From Genes to the Disease

Hashimoto's thyroiditis (HT) is the most prevalent autoimmune thyroid disorder. Intrathyroidal lymphocytic infiltration is followed by a gradual destruction of the thyroid gland which may lead to subclinical or overt hypothyroidism. Biochemical markers of the disease are thyroid peroxidase and/or thyroglobulin autoantibodies in the serum which are present with a higher prevalence in females than in males and increase with age. Although exact mechanisms of aetiology and pathogenesis of the disorder are not completely understood, a strong genetic susceptibility to the disease has been confirmed predominantly by family and twin studies. Several genes were shown to be associated with the disease occurrence, progression, and severity. Genes for human leukocyte antigen, cytotoxic T lymphocyte antigen-4, protein tyrosine phosphatase nonreceptor-type 22, thyroglobulin, vitamin D receptor, and cytokines are considered to be of utmost importance. Amongst endogenous factors for the disease development, the attention is focused predominantly on female sex, pregnancy with postpartum period and fetal microchimerism. Environmental factors influencing HT development are iodine intake, drugs, infections and different chemicals. Disturbed self-tolerance accompanied by the increased antigen presentation is a prerequisite for the HT occurrence, whereas proper interaction of thyroid cells, antigen presenting cells, and T cells are necessary for the initiation of thyroid autoimmunity. Secreted cytokines lead predominantly to T-helper type 1 (Th1) response as well as to Th 17 response which has only recently been implicated. Final outcome of HT is thyroid destruction which is mostly a consequence of the apoptotic processes combined with T-cell mediated cytotoxicity.     

基因芯片技术研究柽柳NaHCO3胁迫下基因的表达

运用基因芯片技术研究了NaHCO3胁迫下柽柳(Tamarix androssowii)基因的表达.将Cy5和Cy3两种荧光染料分别标记在NaHCO3处理和对照的柽柳cDNA上,将两种荧光探针混合,与载有柽柳基因的高密度芯片进行杂交并用芯片扫描系统进行扫描,通过Cy5与Cy3信号强度比值的计算研究基因的差异表达.共获得了89个差异表达的基因,其中,27个下调表达,62个上调表达.BlastX分析表明这些基因按功能可以分为光合作用、活性氧清除、渗透调节、信号传导与表达调控、代谢、发育相关、核糖体蛋白、蛋白质的分解与再生、转运类蛋白、水通道蛋白等几大类别.同时,发现了一些与盐胁迫相关的功能未知基因或未有任何功能信息的基因,这些基因可能在柽柳抗盐过程中具有重要作用.揭示了柽柳的抗盐胁迫涉及的几种重要途径,并获得了NaHCO3胁迫前后柽柳基因表达谱.     

Interrupted Genes in Extremophilic Archaea: Mechanisms of Gene Expression in Early Organisms

Extremophilic Archaea populate biotopes previously considered inaccessible for life. This feature, and the possibility that they are the extant forms of life closest to the last common ancestor, make these organisms excellent candidates for the study of evolution on Earth and stimulate the exobiological research in planets previously considered totally inhospitable. Among the other aspects of the physiology of these organisms, the study of the molecular genetics of extremophilic Archaea can give hints on how the genetic information is transmitted and propagated in ancient forms of life. We review here the expression of interrupted genes in a recently discovered nanoarchaeon and the mechanisms of reprogrammed genetic decoding in Archaea. Presented at: National Workshop on Astrobiology: Search for Life in the Solar System, Capri, Italy, 26 to 28 October, 2005.     

With Reference to Reference Genes: A Systematic Review of Endogenous Controls in Gene Expression Studies

The choice of reference genes that are stably expressed amongst treatment groups is a crucial step in real-time quantitative PCR gene expression studies. Recent guidelines have specified that a minimum of two validated reference genes should be used for normalisation. However, a quantitative review of the literature showed that the average number of reference genes used across all studies was 1.2. Thus, the vast majority of studies continue to use a single gene, with β-actin (ACTB) and/or glyceraldehyde 3-phosphate dehydrogenase (GAPDH) being commonly selected in studies of vertebrate gene expression. Few studies (15%) tested a panel of potential reference genes for stability of expression before using them to normalise data. Amongst studies specifically testing reference gene stability, few found ACTB or GAPDH to be optimal, whereby these genes were significantly less likely to be chosen when larger panels of potential reference genes were screened. Fewer reference genes were tested for stability in non-model organisms, presumably owing to a dearth of available primers in less well characterised species. Furthermore, the experimental conditions under which real-time quantitative PCR analyses were conducted had a large influence on the choice of reference genes, whereby different studies of rat brain tissue showed different reference genes to be the most stable. These results highlight the importance of validating the choice of normalising reference genes before conducting gene expression studies.