A comparative study of S/MAR prediction tools

BACKGROUND: S/MARs are regions of the DNA that are attached to the nuclear matrix. These regions are known to affect substantially the expression of genes. The computer prediction of S/MARs is a highly significant task which could contribute to our understanding of chromatin organisation in eukaryotic cells, the number and distribution of boundary elements, and the understanding of gene regulation in eukaryotic cells. However, while a number of S/MAR predictors have been proposed, their accuracy has so far not come under scrutiny. RESULTS: We have selected S/MARs with sufficient experimental evidence and used these to evaluate existing methods of S/MAR prediction. Our main results are: 1.) all existing methods have little predictive power, 2.) a simple rule based on AT-percentage is generally competitive with other methods, 3.) in practice, the different methods will usually identify different sub-sequences as S/MARs, 4.) more research on the H-Rule would be valuable. CONCLUSION: A new insight is needed to design a method which will predict S/MARs well. Our data, including the control data, has been deposited as additional material and this may help later researchers test new predictors

Single-Cell Transcriptional Analysis of Neuronal Progenitors

AbstractThe extraordinary cellular heterogeneity of the mammalian nervous system has largely hindered the molecular analysis of neuronal identity and diversity. In order to uncover mechanisms involved in neuronal differentiation and diversification, we have monitored the expression profiles of individual neurons and progenitor cells collected from dissociated tissue or captured from intact slices. We demonstrate that this technique provides a sensitive and reproducible representation of the single-cell transcriptome. In the olfactory system, hundreds of transcriptional differences were identified between olfactory progenitors and mature sensory neurons, enabling us to define the large variety of signaling pathways expressed by individual progenitors at a precise developmental stage. Finally, we show that regional differences in gene expression can be predicted from transcriptional analysis of single neuronal precursors isolated by laser capture from defined areas of the developing brain

A hierarchical model of transcriptional dynamics allows robust estimation of transcription rates in populations of single cells with variable gene copy number

Motivation: cis-regulatory DNA sequence elements, such as enhancers and silencers, function to control the spatial and temporal expression of their target genes. Although the overall levels of gene expression in large cell populations seem to be precisely controlled, transcription of individual genes in single cells is extremely variable in real time. It is, therefore, important to understand how these cis-regulatory elements function to dynamically control transcription at single-cell resolution. Recently, statistical methods have been proposed to back calculate the rates involved in mRNA transcription using parameter estimation of a mathematical model of transcription and translation. However, a major complication in these approaches is that some of the parameters, particularly those corresponding to the gene copy number and transcription rate, cannot be distinguished; therefore, these methods cannot be used when the copy number is unknown. Results: Here, we develop a hierarchical Bayesian model to estimate biokinetic parameters from live cell enhancer–promoter reporter measurements performed on a population of single cells. This allows us to investigate transcriptional dynamics when the copy number is variable across the population. We validate our method using synthetic data and then apply it to quantify the function of two known developmental enhancers in real time and in single cells

Conserved cis-regulatory modules control robustness in Msx1 expression at single cell resolution

The process of transcription is highly stochastic leading to cell-to-cell variations and noise in gene expression levels. However, key essential genes have to be precisely expressed at the correct amount and time to ensure proper cellular development and function. Studies in yeast and bacterial systems have shown that gene expression noise decreases as mean expression levels increase, a relationship that is controlled by promoter DNA sequence. However, the function of distal cis-regulatory modules (CRMs), an evolutionary novelty of metazoans, in controlling transcriptional robustness and variability is poorly understood. In this study, we used live cell imaging of transfected reporters combined with a mathematical modelling and statistical inference scheme to quantify the function of conserved Msx1 CRMs and promoters in modulating single-cell real-time transcription rates in C2C12 mouse myoblasts. The results show that the mean expression–noise relationship is solely promoter controlled for this key pluripotency regulator. In addition, we demonstrate that CRMs modulate single-cell basal promoter rate distributions in a graded manner across a population of cells. This extends the rheostatic model of CRM action to provide a more detailed understanding of CRM function at single-cell resolution. We also identify a novel CRM transcriptional filter function that acts to reduce intracellular variability in transcription rates and show that this can be phylogenetically separable from rate modulating CRM activities. These results are important for understanding how the expression of key vertebrate developmental transcription factors is precisely controlled both within and between individual cells

Journal club. A genomic systems biologist muses on how shared DNA mistakes reveal shared cellular ancestry

The way an organism or cancer develops often depends on how cells relate to each other. This is because ancestral cells make molecular decisions that affect the regulation of genes in their offspring. Revealing complex lineage relationships usually requires sophisticated methods of molecular embryology that are available for only a few 'model' species and not for humans. I recently stumbled upon a string of creative papers from two competing labs that prepare the ground for change

Novel cis-regulatory modules control expression of the hairy and enhancer of split-1 (HES1) transcription factor in myoblasts

The expression profile of a gene is controlled by DNA sequences called cis-regulatory modules (CRMs). CRMs can function over large genomic distances and can be located many kilobases away from their target promoters. hes1 is a key developmental gene that is overexpressed in certain cancers and is a primary target of NOTCH signaling. Despite this, analysis of hes1 transcriptional control has been limited solely to its promoter. Here, we identify seven conserved DNA sequence blocks, representing the hes1 promoter and six novel CRMs, within 57 kb upstream of the mouse hes1 gene. We identify 12 binding sites for the RBP-Jκ NOTCH effector and a single M-CAT motif within these regions. We validate RBP-Jκ and TEAD family occupancy in cells in culture and test the response of each of these CRMs to active NOTCH. We show that two regions, CRM5 and CRM7, function as enhancers, and four can repress transcription. A pair of RBP-Jκ motifs arranged in a tail-tail configuration in CRM5 and the M-CAT motif in CRM7 are necessary for enhancer function. Furthermore, these enhancers are occupied by transcriptional co-activators and loop onto the hes1 promoter within the endogenous hes1 locus. This work demonstrates the power of combining computational genomics and experimental methodologies to identify novel CRMs and characterize their function. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc

Loss of Sex Discrimination and Male-Male Aggression in Mice Deficient for TRP2

The mouse vomeronasal organ (VNO) is thought to mediate social behaviors and neuroendocrine changes elicited by pheromonal cues. The molecular mechanisms underlying the sensory response to pheromones and the behavioral repertoire induced through the VNO are not fully characterized. Using the tools of mouse genetics and multielectrode recording, we demonstrate that the sensory activation of VNO neurons requires TRP2, a putative ion channel of the transient receptor potential family that is expressed exclusively in these neurons. Moreover, we show that male mice deficient in TRP2 expression fail to display male-male aggression, and they initiate sexual and courtship behaviors toward both males and females. Our study suggests that, in the mouse, sensory activation of the VNO is essential for sex discrimination of conspecifics and thus ensures gender-specific behavior

Characterization of an anti-apoptotic glycoprotein encoded by Kaposi’s sarcoma-associated herpesvirus which resembles a spliced variant of human survivin

We have investigated the expression and function of a novel protein encoded by open reading frame (ORF) K7 of Kaposi’s sarcoma-associated herpesvirus (KSHV). Computational analyses revealed that K7 is structurally related to survivin-ΔEx3, a splice variant of human survivin that protects cells from apoptosis by an undefined mechanism. Both K7 and survivin-ΔEx3 contain a mitochondrial-targeting sequence, an N-terminal region of a BIR (baculovirus IAP repeat) domain and a putative BH2 (Bcl-2 homology)-like domain. These suggested that K7 is a new viral anti-apoptotic protein and survivin-ΔEx3 is its likely cellular homologue. We show that K7 is a glycoprotein, which can inhibit apoptosis and anchor to intracellular membranes where Bcl-2 resides. K7 does not associate with Bax, but does bind to Bcl-2 via its putative BH2 domain. In addition, K7 binds to active caspase-3 via its BIR domain and thus inhibits the activity of caspase-3. The BH2 domain of K7 is crucial for the inhibition of caspase-3 activity and is therefore essential for its anti-apoptotic function. Furthermore, K7 bridges Bcl-2 and activated caspase-3 into a protein complex. K7 therefore appears to be an adaptor protein and part of an anti-apoptotic complex that presents effector caspases to Bcl-2, enabling Bcl-2 to inhibit caspase activity. These data also suggest that survivin-ΔEx3 might function by a similar mechanism to that of K7. We denote K7 as vIAP (viral inhibitor-of-apoptosis protein)

Neuropilin-2 Mediates Axonal Fasciculation, Zonal Segregation, but Not Axonal Convergence, of Primary Accessory Olfactory Neurons

AbstractThe mechanisms that underlie axonal pathfinding of vomeronasal neurons from the vomeronasal organ (VNO) in the periphery to select glomeruli in the accessory olfactory bulb (AOB) are not well understood. Neuropilin-2, a receptor for secreted semaphorins, is expressed in V1R- and V3R-expressing, but not V2R-expressing, postnatal vomeronasal neurons. Analysis of the vomeronasal nerve in neuropilin-2 (npn-2) mutant mice reveals pathfinding defects at multiple choice points. Vomeronasal sensory axons are severely defasciculated and a subset innervates the main olfactory bulb (MOB). While most axons of V1R-expressing neurons reach the AOB and converge into distinct glomeruli in stereotypic locations, they are no longer restricted to their normal anterior AOB target zone. Thus, Npn-2 and candidate pheromone receptors play distinct and complementary roles in promoting the wiring and patterning of sensory neurons in the accessory olfactory system