Circulating transcripts in maternal blood reflect a molecular signature of early-onset preeclampsia.

Circulating RNA (C-RNA) is continually released into the bloodstream from tissues throughout the body, offering an opportunity to noninvasively monitor all aspects of pregnancy health from conception to birth. We asked whether C-RNA analysis could robustly detect aberrations in patients diagnosed with preeclampsia (PE), a prevalent and potentially fatal pregnancy complication. As an initial examination, we sequenced the circulating transcriptome from 40 pregnancies at the time of severe, early-onset PE diagnosis and 73 gestational age-matched controls. Differential expression analysis identified 30 transcripts with gene ontology annotations and tissue expression patterns consistent with the placental dysfunction, impaired fetal development, and maternal immune and cardiovascular system dysregulation characteristic of PE. Furthermore, machine learning identified combinations of 49 C-RNA transcripts that classified an independent cohort of patients (early-onset PE, n = 12; control, n = 12) with 85 to 89% accuracy. C-RNA may thus hold promise for improving the diagnosis and identification of at-risk pregnancies.

The Role of Transcription Factors and Nuclear Pore Proteins in Controlling the Spatial Organization of the Yeast Genome.

Loss of nuclear pore complex (NPC) proteins, transcription factors (TFs), histone modification enzymes, Mediator, and factors involved in mRNA export disrupts the physical interaction of chromosomal sites with NPCs. Conditional inactivation and ectopic tethering experiments support a direct role for the TFs Gcn4 and Nup2 in mediating interaction with the NPC but suggest an indirect role for factors involved in mRNA export or transcription. A conserved "positioning domain" within Gcn4 controls interaction with the NPC and inter-chromosomal clustering and promotes transcription of target genes. Such a function may be quite common; a comprehensive screen reveals that tethering of most yeast TFs is sufficient to promote targeting to the NPC. While some TFs require Nup100, others do not, suggesting two distinct targeting mechanisms. These results highlight an important and underappreciated function of TFs in controlling the spatial organization of the yeast genome through interaction with the NPC.

Transcription factors dynamically control the spatial organization of the yeast genome.

In yeast, inducible genes such as INO1, PRM1 and HIS4 reposition from the nucleoplasm to nuclear periphery upon activation. This leads to a physical interaction with nuclear pore complex (NPC), interchromosomal clustering, and stronger transcription. Repositioning to the nuclear periphery is controlled by cis-acting transcription factor (TF) binding sites located within the promoters of these genes and the TFs that bind to them. Such elements are both necessary and sufficient to control positioning of genes to the nuclear periphery. We have identified 4 TFs capable of controlling the regulated positioning of genes to the nuclear periphery in budding yeast under different conditions: Put3, Cbf1, Gcn4 and Ste12. In each case, we have defined the molecular basis of regulated relocalization to the nuclear periphery. Put3- and Cbf1-mediated targeting to nuclear periphery is regulated through local recruitment of Rpd3(L) histone deacetylase complex by transcriptional repressors. Rpd3(L), through its histone deacetylase activity, prevents TF-mediated gene positioning by blocking TF binding. Many yeast transcriptional repressors were capable of blocking Put3-mediated recruitment; 11 of these required Rpd3. Thus, it is a general function of transcription repressors to regulate TF-mediated recruitment. However, Ste12 and Gcn4-mediated recruitment is regulated independently of Rpd3(L) and transcriptional repressors. Ste12-mediated recruitment is regulated by phosphorylation of an inhibitor called Dig2, and Gcn4-mediated gene targeting is up-regulated by increasing Gcn4 protein levels. The ability to control spatial position of genes in yeast represents a novel function for TFs and different regulatory strategies provide dynamic control of the yeast genome through different time scales.

Set1/COMPASS and Mediator are repurposed to promote epigenetic transcriptional memory.

In yeast and humans, previous experiences can lead to epigenetic transcriptional memory: repressed genes that exhibit mitotically heritable changes in chromatin structure and promoter recruitment of poised RNA polymerase II preinitiation complex (RNAPII PIC), which enhances future reactivation. Here, we show that INO1 memory in yeast is initiated by binding of the Sfl1 transcription factor to the cis-acting Memory Recruitment Sequence, targeting INO1 to the nuclear periphery. Memory requires a remodeled form of the Set1/COMPASS methyltransferase lacking Spp1, which dimethylates histone H3 lysine 4 (H3K4me2). H3K4me2 recruits the SET3C complex, which plays an essential role in maintaining this mark. Finally, while active INO1 is associated with Cdk8(-) Mediator, during memory, Cdk8(+) Mediator recruits poised RNAPII PIC lacking the Kin28 CTD kinase. Aspects of this mechanism are generalizable to yeast and conserved in human cells. Thus, COMPASS and Mediator are repurposed to promote epigenetic transcriptional poising by a highly conserved mechanism.

Strategies to regulate transcription factor-mediated gene positioning and interchromosomal clustering at the nuclear periphery.

In budding yeast, targeting of active genes to the nuclear pore complex (NPC) and interchromosomal clustering is mediated by transcription factor (TF) binding sites in the gene promoters. For example, the binding sites for the TFs Put3, Ste12, and Gcn4 are necessary and sufficient to promote positioning at the nuclear periphery and interchromosomal clustering. However, in all three cases, gene positioning and interchromosomal clustering are regulated. Under uninducing conditions, local recruitment of the Rpd3(L) histone deacetylase by transcriptional repressors blocks Put3 DNA binding. This is a general function of yeast repressors: 16 of 21 repressors blocked Put3-mediated subnuclear positioning; 11 of these required Rpd3. In contrast, Ste12-mediated gene positioning is regulated independently of DNA binding by mitogen-activated protein kinase phosphorylation of the Dig2 inhibitor, and Gcn4-dependent targeting is up-regulated by increasing Gcn4 protein levels. These different regulatory strategies provide either qualitative switch-like control or quantitative control of gene positioning over different time scales.

A new direction for gene looping.

Upon binding to a promoter, RNA polymerase II can synthesize either a coding mRNA or a divergently transcribed noncoding RNA. In a recent issue of Science, Tan-Wong et al. (2012) find that intragenic looping increases the proper orientation of RNA polymerase II, reducing the production of divergent noncoding transcripts.