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Background: Vocal learning is a rare, convergent trait that is fundamental to both human speech and birdsong. The Forkhead Box P2 (FoxP2) transcription factor appears necessary for both types of learned signals, as human mutations in FoxP2 result in speech deficits, and disrupting its expression in zebra finches impairs male-specific song learning. In juvenile and adult male finches, striatal FoxP2 mRNA and protein decline acutely within song-dedicated neurons during singing, indicating that its transcriptional targets are also behaviorally regulated. The identities of these targets in songbirds, and whether they differ across sex, development and/or behavioral conditions, are largely unknown., Results: Here we used chromatin immunoprecipitation followed by sequencing (ChIP-Seq) to identify genomic sites bound by FoxP2 in male and female, juvenile and adult, and singing and non-singing birds. Our results suggest robust FoxP2 binding concentrated in putative promoter regions of genes. The number of genes likely to be bound by FoxP2 varied across conditions, suggesting specialized roles of the candidate targets related to sex, age, and behavioral state. We validated these binding targets both bioinformatically, with comparisons to previous studies and biochemically, with immunohistochemistry using an antibody for a putative target gene. Gene ontology analyses revealed enrichment for human speech- and language-related functions in males only, consistent with the sexual dimorphism of song learning in this species. Fewer such targets were found in juveniles relative to adults, suggesting an expansion of this regulatory network with maturation. The fewest speech-related targets were found in the singing condition, consistent with the well-documented singing-driven down-regulation of FoxP2 in the songbird striatum., Conclusions: Overall, these data provide an initial catalog of the regulatory landscape of FoxP2 in an avian vocal learner, offering dozens of target genes for future study and providing insight into the molecular underpinnings of vocal learning., Competing Interests: Competing Interests The authors declare that they have no competing interests.

Abstract Background Gene activity is largely controlled by transcriptional regulation through the action of transcription factors and other regulators. QsMYB1 is a member of the R2R3-MYB transcription factor family related to secondary growth, and in particular, with the cork development process. In order to identify the putative gene targets of QsMYB1 across the cork oak genome we developed a ChIP-Seq strategy. Results Results provide direct evidence that QsMY1B targets genes encoding for enzymes involved in the lignin and suberin pathways as well as gene encoding for ABCG transporters and LTPs implicated in the transport of monomeric suberin units across the cellular membrane. These results highlight the role of QsMYB1 as a regulator of lignin and suberin biosynthesis, transport and assembly. Conclusion To our knowledge, this work constitutes the first ChIP-Seq experiment performed in cork oak, a non-model plant species with a long-life cycle, and these results will contribute to deepen the knowledge about the molecular mechanisms of cork formation and differentiation.

Background: Zfp819, a member of the Krüppel-associated box (KRAB) family, encodes a spermatogenic cell-specific transcription factor. Zfp819-overexpression induces apoptosis and inhibits proliferation in somatic cell lines. Results: In the present study, we examined the cellular effects of Zfp819 in a male germ cell line (GC-2 cells). Overexpression of Zfp819 demonstrated an increase in the number of apoptotic cells, leading to inhibition of proliferation in GC-2 cells. We further investigated genes regulated by ZFP819 using microarray analysis and chromatin-immunoprecipitation combined with microarray analysis (ChIP-chip) in GC-2 cells. We identified 118 downregulated genes in Zfp819-overexpressing GC-2 cells using microarray analysis. ChIP-chip assay revealed that 1011 promoter sites (corresponding to 262 genes) were specifically enriched in GC-2 cells transfected with Zfp819. Two genes (trinucleotide repeat containing 6b and annexin A11) were commonly found when we compared the data between microarray and ChIP-chip analyses. Consistent with these results, Zfp819 overexpression significantly reduced the transcript levels of the two genes by binding to their promoter regions. Tissue distribution analysis indicated that both genes were predominantly expressed in testis. It has been reported that these two genes function in apoptosis. Conclusion: Collectively, our study provides inclusive information on germ cell-specific gene regulation by ZFP819, which is involved in apoptosis, to maintain the integrity of spermatogenesis. [ABSTRACT FROM AUTHOR]

Abstract: SLC22A18, a poly-specific organic cation transporter, is paternally imprinted in humans and mice. It shows loss-of-heterozygosity in childhood and adult tumors, and gain-of-imprinting in hepatocarcinomas and breast cancers. Despite the importance of this gene, its transcriptional regulation has not been studied, and the promoter has not yet been characterized. We therefore set out to identify the potential cis-regulatory elements including the promoter of this gene. The luciferase reporter assay in human cells indicated that a region from −120 bp to +78 bp is required for the core promoter activity. No consensus TATA or CAAT boxes were found in this region, but two Sp1 binding sites were conserved in human, chimpanzee, mouse and rat. Mutational analysis of the two Sp1 sites suggested their requirement for the promoter activity. Chromatin-immunoprecipitation showed binding of Sp1 to the promoter region in vivo. Overexpression of Sp1 in Drosophila Sp1-null SL2 cells suggested that Sp1 is the transactivator of the promoter. The human core promoter was functional in mouse 3T3 and monkey COS7 cells. We found a CpG island which spanned the core promoter and exon 1. COBRA technique did not reveal promoter methylation in 10 normal oral tissues, 14 oral tumors, and two human cell lines HuH7 and A549. This study provides the first insight into the mechanism that controls expression of this imprinted tumor suppressor gene. A COBRA-based assay has been developed to look for promoter methylation in different cancers. The present data will help to understand the regulation of this gene and its role in tumorigenesis. [Copyright &y& Elsevier]

Abstract: The E2F-associated phosphoprotein (EAPP) is a ubiquitous nuclear protein that interacts with the activating members of the E2F family of transcription factors and increases the activity of several cell-cycle regulated promoters in an E2F-dependent manner. Our previous studies also showed that EAPP levels are elevated in most transformed human cells. To examine the molecular basis of this increase of EAPP we isolated and studied the nucleotide sequence at the 5′ end of the EAPP gene. In silico analysis revealed a TATA-less promoter with several putative binding sites for transcription factors, the most probable ones being Sp1, Sp3 and Egr-1. We could confirm the binding of these factors in vitro by electrophoretic mobility shift assays, supershift experiments and competition assays. Additionally we could validate the binding in vivo by chromatin-immunoprecipitation assays. To analyse the function of these transcription factors in the expression of EAPP, we performed reporter-assays with the promoter and truncations thereof. We found that Sp1 and Egr-1 stimulate the EAPP promoter, whereas Sp3 acts as a repressor that could even overcome the positive effect of the activators. Increasing the amounts of Sp3 also caused a strong reduction of EAPP, but the overexpression of Sp1 or Egr-1 resulted in only marginally higher EAPP levels. Our results suggest that the elevated EAPP levels in transformed cells can be caused by reduced Sp3 activity, but higher Sp1 activity might also play a role. [Copyright &y& Elsevier]

Background: Gene activity is largely controlled by transcriptional regulation through the action of transcription factors and other regulators. QsMYB1 is a member of the R2R3-MYB transcription factor family related to secondary growth, and in particular, with the cork development process. In order to identify the putative gene targets of QsMYB1 across the cork oak genome we developed a ChIP-Seq strategy. Results: Results provide direct evidence that QsMY1B targets genes encoding for enzymes involved in the lignin and suberin pathways as well as gene encoding for ABCG transporters and LTPs implicated in the transport of monomeric suberin units across the cellular membrane. These results highlight the role of QsMYB1 as a regulator of lignin and suberin biosynthesis, transport and assembly. Conclusion: To our knowledge, this work constitutes the first ChIP-Seq experiment performed in cork oak, a non-model plant species with a long-life cycle, and these results will contribute to deepen the knowledge about the molecular mechanisms of cork formation and differentiation. [ABSTRACT FROM AUTHOR]

Environmental factors affect self-renewal of stem cells by modulating the components of self-renewal networks. Heat shock, an environmental factor, induces heat shock factors (HSFs), which up-regulate stress response-related genes. However, the link of heat shock to self-renewal of stem cells has not been elucidated yet. Here, we present the direct link of heat shock to a core stem cell regulator, OCT4, in the self-renewal network through SAPK/JNK and HSF1 pathway. We first showed that heat shock initiated differentiation of human embryonic stem cells (hESCs). Gene expression analysis revealed that heat shock increased the expression of many genes involved in cellular processes related to differentiation of stem cells. We then examined the effects of HSFs induced by heat shock on core self-renewal factors. Among HSFs, heat shock induced mainly HSF1 in hESCs. The HSF1 repressed the expression of OCT4, leading to the differentiation of hESCs and the above differentiation-related gene expression change. We further examined the effects of the upstream MAP (mitogen-activated protein) kinases of HSF1 on the repression of OCT4 expression by HSF1. Among the MAP kinases, SAPK/JNK controlled predominantly the repression of the OCT4 expression by HSF1. The direct link of heat shock to the core self-renewal regulator through SAPK/JNK and HSF1 provides a fundamental basis for understanding the effect of heat and other stresses involving activation of HSF1 on the self-renewal program and further controlling differentiation of hESCs in a broad spectrum of stem cell applications using these stresses., (© 2013.)

14-3-3 proteins regulate numerous cellular processes through interaction with a variety of proteins, and have been identified as HNF1α binding partner by mass spectrometry analysis in our previous study. In the present study, the interaction between 14-3-3ζ and HNF1α has been further validated by in vivo and in vitro assays. Moreover, we have found that overexpression of 14-3-3ζ potentiated the transcriptional activity of HNF1α in cultured cells, and silencing of 14-3-3ζ by RNA interference in HepG2 cells specifically affected the HNF1α-dependent gene expression. Furthermore, we have demonstrated that 14-3-3ζ is recruited to endogenous HNF1α responsive promoters and enhances HNF1α binding to its cognate DNA sequences. In addition, we have also provided evidence that the association between HNF1α and 14-3-3ζ is phosphorylation-dependent. Taken together, these results suggest that 14-3-3ζ may be an endogenous physiologic regulator of HNF1α., (Copyright © 2013 Elsevier B.V. All rights reserved.)