YY1 negatively regulates mouse myelin proteolipid protein (Plp1) gene expression in oligodendroglial cells

YY1 (Yin and Yang 1) is a multifunctional, ubiquitously expressed, zinc finger protein that can act as a transcriptional activator, repressor, or initiator element binding protein. Previous studies have shown that YY1 modulates the activity of reporter genes driven by the myelin PLP (proteolipid protein) (PLP1/Plp1) promoter. However, it is known that Plp1 intron 1 DNA contains regulatory elements that are required for the dramatic increase in gene activity, coincident with the active myelination period of CNS (central nervous system) development. The intron in mouse contains multiple prospective YY1 target sites including one within a positive regulatory module called the ASE (anti-silencer/enhancer) element. Results presented here demonstrate that YY1 has a negative effect on the activity of a Plp1-lacZ fusion gene [PLP(+)Z] in an immature oligodendroglial cell line (Oli-neu) that is mediated through sequences present in Plp1 intron 1 DNA. Yet YY1 does not bind to its alleged site in the ASE (even though the protein is capable of recognizing a target site in the promoter), indicating that the down-regulation of PLP(+)Z activity by YY1 in Oli-neu cells does not occur through a direct interaction of YY1 with the ASE sequence. Previous studies with Yy1 conditional knockout mice have demonstrated that YY1 is essential for the differentiation of oligodendrocyte progenitors. Nevertheless, the current study suggests that YY1 functions as a repressor (not an activator) of Plp1 gene expression in immature oligodendrocytes. Perhaps YY1 functions to keep the levels of PLP in check in immature cells before vast quantities of the protein are needed in mature myelinating oligodendrocytes

Acarbose, 17‐α‐estradiol, and nordihydroguaiaretic acid extend mouse lifespan preferentially in males

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/106661/1/acel12170.pd

Does age matter? The impact of rodent age on study outcomes

Rodent models produce data which underpin biomedical research and non-clinical drug trials, but translation from rodents into successful clinical outcomes is often lacking. There is a growing body of evidence showing that improving experimental design is key to improving the predictive nature of rodent studies and reducing the number of animals used in research. Age, one important factor in experimental design, is often poorly reported and can be overlooked. The authors conducted a survey to assess the age used for a range of models, and the reasoning for age choice. From 297 respondents providing 611 responses, researchers reported using rodents most often in the 6–20 week age range regardless of the biology being studied. The age referred to as ‘adult’ by respondents varied between six and 20 weeks. Practical reasons for the choice of rodent age were frequently given, with increased cost associated with using older animals and maintenance of historical data comparability being two important limiting factors. These results highlight that choice of age is inconsistent across the research community and often not based on the development or cellular ageing of the system being studied. This could potentially result in decreased scientific validity and increased experimental variability. In some cases the use of older animals may be beneficial. Increased scientific rigour in the choice of the age of rodent may increase the translation of rodent models to humans

The Proteolipid Protein Promoter Drives Expression outside of the Oligodendrocyte Lineage during Embryonic and Early Postnatal Development

The proteolipid protein (Plp) gene promoter is responsible for driving expression of one of the major components of myelin – PLP and its splice variant DM-20. Both products are classically thought to express predominantly in oligodendrocytes. However, accumulating evidence suggests Plp expression is more widespread than previously thought. In an attempt to create a mouse model for inducing oligodendrocyte-specific gene deletions, we have generated transgenic mice expressing a Cre recombinase cDNA under control of the mouse Plp promoter. We demonstrate Plp promoter driven Cre expression is restricted predominantly to mature oligodendrocytes of the central nervous system (CNS) at postnatal day 28. However, crosses into the Rosa26LacZ and mT/mG reporter mouse lines reveal robust and widespread Cre activity in neuronal tissues at E15.5 and E10.5 that is not strictly oligodendrocyte lineage specific. By P28, all CNS tissues examined displayed high levels of reporter gene expression well outside of defined white matter zones. Importantly, our study reinforces the emerging idea that Plp promoter activity is not restricted to the myelinating cell lineage, but rather, has widespread activity both during embryonic and early postnatal development in the CNS. Specificity of the promoter to the oligodendrocyte cell lineage, as shown through the use of a tamoxifen inducible Plp-CreERt line, occurs only at later postnatal stages. Understanding the temporal shift in Plp driven expression is of consequence when designing experimental models to study oligodendrocyte biology

Fold change and p-value cutoffs significantly alter microarray interpretations

<p>Abstract</p> <p>Background</p> <p>As context is important to gene expression, so is the preprocessing of microarray to transcriptomics. Microarray data suffers from several normalization and significance problems. Arbitrary fold change (FC) cut-offs of >2 and significance p-values of <0.02 lead data collection to look only at genes which vary wildly amongst other genes. Therefore, questions arise as to whether the biology or the statistical cutoff are more important within the interpretation. In this paper, we reanalyzed a zebrafish (<it>D. rerio</it>) microarray data set using GeneSpring and different differential gene expression cut-offs and found the data interpretation was drastically different. Furthermore, despite the advances in microarray technology, the array captures a large portion of genes known but yet still leaving large voids in the number of genes assayed, such as leptin a pleiotropic hormone directly related to hypoxia-induced angiogenesis.</p> <p>Results</p> <p>The data strongly suggests that the number of differentially expressed genes is more up-regulated than down-regulated, with many genes indicating conserved signalling to previously known functions. Recapitulated data from Marques et al. (2008) was similar but surprisingly different with some genes showing unexpected signalling which may be a product of tissue (heart) or that the intended response was transient.</p> <p>Conclusions</p> <p>Our analyses suggest that based on the chosen statistical or fold change cut-off; microarray analysis can provide essentially more than one answer, implying data interpretation as more of an art than a science, with follow up gene expression studies a must. Furthermore, gene chip annotation and development needs to maintain pace with not only new genomes being sequenced but also novel genes that are crucial to the overall gene chips interpretation.</p

Vesicoureteral Reflux and Other Urinary Tract Malformations in Mice Compound Heterozygous for Pax2 and Emx2

Congenital anomalies of the kidney and urinary tract (CAKUT) are the most common cause of chronic kidney disease in children. This disease group includes a spectrum of urinary tract defects including vesicoureteral reflux, duplex kidneys and other developmental defects that can be found alone or in combination. To identify new regulators of CAKUT, we tested the genetic cooperativity between several key regulators of urogenital system development in mice. We found a high incidence of urinary tract anomalies in Pax2;Emx2 compound heterozygous mice that are not found in single heterozygous mice. Pax2+/−;Emx2+/− mice harbor duplex systems associated with urinary tract obstruction, bifid ureter and a high penetrance of vesicoureteral reflux. Remarkably, most compound heterozygous mice refluxed at low intravesical pressure. Early analysis of Pax2+/−;Emx2+/− embryos point to ureter budding defects as the primary cause of urinary tract anomalies. We additionally establish Pax2 as a direct regulator of Emx2 expression in the Wolffian duct. Together, these results identify a haploinsufficient genetic combination resulting in CAKUT-like phenotype, including a high sensitivity to vesicoureteral reflux. As both genes are located on human chromosome 10q, which is lost in a proportion of VUR patients, these findings may help understand VUR and CAKUT in humans

The PlcR Virulence Regulon of Bacillus cereus

PlcR is a Bacillus cereus transcriptional regulator, which activates gene expression by binding to a nucleotidic sequence called the ‘PlcR box’. To build a list of all genes included in the PlcR regulon, a consensus sequence was identified by directed mutagenesis. The reference strain ATCC14579 sequenced genome was searched for occurrences of this consensus sequence to produce a virtual regulon. PlcR control of these genes was confirmed by comparing gene expression in the reference strain and its isogenic Δ-plcR strain using DNA microarrays, lacZ fusions and proteomics methods. The resulting list included 45 genes controlled by 28 PlcR boxes. Forty of the PlcR controlled proteins were exported, of which 22 were secreted in the extracellular medium and 18 were bound or attached to cell wall structures (membrane or peptidoglycan layer). The functions of these proteins were related to food supply (phospholipases, proteases, toxins), cell protection (bacteriocins, toxins, transporters, cell wall biogenesis) and environment-sensing (two-component sensors, chemotaxis proteins, GGDEF family regulators). Four genes coded for cytoplasmic regulators. The PlcR regulon appears to integrate a large range of environmental signals, including food deprivation and self cell-density, and regulate the transcription of genes designed to overcome obstacles that hinder B. cereus growth within the host: food supply, host barriers, host immune defenses, and competition with other bacterial species. PlcR appears to be a key component in the efficient adaptation of B. cereus to its host environment

Development of a framework for genotyping bovine-derived Cryptosporidium parvum, using a multilocus fragment typing tool

Background: There is a need for an integrated genotyping approach for C. parvum; no sufficiently discriminatory scheme to date has been fully validated or widely adopted by veterinary or public health researchers. Multilocus fragment typing (MLFT) can provide good differentiation and is relatively quick and cheap to perform. A MLFT tool was assessed in terms of its typeability, specificity, precision (repeatability and reproducibility), accuracy and ability to genotypically discriminate bovine-derived Cryptosporidium parvum. Methods: With the aim of working towards a consensus, six markers were selected for inclusion based on their successful application in previous studies: MM5, MM18, MM19, TP14, MS1 and MS9. Alleles were assigned according to the fragment sizes of repeat regions amplified, as determined by capillary electrophoresis. In addition, a region of the GP60 gene was amplified and sequenced to determine gp60 subtype and this was added to the allelic profiles of the 6 markers to determine the multilocus genotype (MLG). The MLFT tool was applied to 140 C. parvum samples collected in two cross-sectional studies of UK calves, conducted in Cheshire in 2004 (principally dairy animals) and Aberdeenshire/Caithness in 2011 (beef animals). Results: Typeability was 84 %. The primers did not amplify tested non-parvum species frequently detected in cattle. In terms of repeatability, within- and between-run fragment sizes showed little variability. Between laboratories, fragment sizes differed but allele calling was reproducible. The MLFT had good discriminatory ability (Simpson’s Index of Diversity, SID, was 0.92), compared to gp60 sequencing alone (SID 0.44). Some markers were more informative than others, with MS1 and MS9 proving monoallelic in tested samples. Conclusions: Further inter-laboratory trials are now warranted with the inclusion of human-derived C. parvum samples, allowing progress towards an integrated, standardised typing scheme to enable source attribution and to determine the role of livestock in future outbreaks of human C. parvum

SLEPR: A Sample-Level Enrichment-Based Pathway Ranking Method — Seeking Biological Themes through Pathway-Level Consistency

Analysis of microarray and other high throughput data often involves identification of genes consistently up or down-regulated across samples as the first step in extraction of biological meaning. This gene-level paradigm can be limited as a result of valid sample fluctuations and biological complexities. In this report, we describe a novel method, SLEPR, which eliminates this limitation by relying on pathway-level consistencies. Our method first selects the sample-level differentiated genes from each individual sample, capturing genes missed by other analysis methods, ascertains the enrichment levels of associated pathways from each of those lists, and then ranks annotated pathways based on the consistency of enrichment levels of individual samples from both sample classes. As a proof of concept, we have used this method to analyze three public microarray datasets with a direct comparison with the GSEA method, one of the most popular pathway-level analysis methods in the field. We found that our method was able to reproduce the earlier observations with significant improvements in depth of coverage for validated or expected biological themes, but also produced additional insights that make biological sense. This new method extends existing analyses approaches and facilitates integration of different types of HTP data

Translational control of depression-like behavior via phosphorylation of eukaryotic translation initiation factor 4E

Translation of mRNA into protein has a fundamental role in neurodevelopment, plasticity, and memory formation; however, its contribution in the pathophysiology of depressive disorders is not fully understood. We investigated the involvement of MNK1/2 (MAPK-interacting serine/threonine-protein kinase 1 and 2) and their target, eIF4E (eukaryotic initiation factor 4E), in depression-like behavior in mice. Mice carrying a mutation in eIF4E for the MNK1/2 phosphorylation site (Ser209Ala, Eif4e ki/ki), the Mnk1/2 double knockout mice (Mnk1/2 -/-), or mice treated with the MNK1/2 inhibitor, cercosporamide, displayed anxiety-and depression-like behaviors, impaired serotonin-induced excitatory synaptic activity in the prefrontal cortex, and diminished firing of the dorsal raphe neurons. In Eif4e ki/ki mice, brain IκBα, was decreased, while the NF-κB target, TNFα was elevated. TNFα inhibition in Eif4e ki/ki mice rescued, whereas TNFα administration to wild-type mice mimicked the depression-like behaviors and 5-HT synaptic deficits. We conclude that eIF4E phosphorylation modulates depression-like behavior through regulation of inflammatory responses