Regulation Of Gene Expression In Cells Research Articles

A DEAD-box helicase drives the partitioning of a pro-differentiation NAB protein into nuclear foci

How cells regulate gene expression in a precise spatiotemporal manner during organismal development is a fundamental question in biology. Although the role of transcriptional condensates in gene regulation has been established, little is known about the function and regulation of these molecular assemblies in the context of animal development and physiology. Here we show that the evolutionarily conserved DEAD-box helicase DDX-23 controls cell fate in Caenorhabditis elegans by binding to and facilitating the condensation of MAB-10, the C. elegans homolog of mammalian NGFI-A-binding (NAB) protein. MAB-10 is a transcriptional cofactor that functions with the early growth response (EGR) protein LIN-29 to regulate the transcription of genes required for exiting the cell cycle, terminal differentiation, and the larval-to-adult transition. We suggest that DEAD-box helicase proteins function more generally during animal development to control the condensation of NAB proteins important in cell identity and that this mechanism is evolutionarily conserved. In mammals, such a mechanism might underlie terminal cell differentiation and when dysregulated might promote cancerous growth.

Ancient transcriptional regulators can easily evolve new pair-wise cooperativity

Cells regulate gene expression by the specific binding of transcription regulators to cis-regulatory sequences. Pair-wise cooperativity between regulators-whereby two different regulators physically interact and bind DNA in a cooperative manner-is common and permits complex modes of gene regulation. Over evolutionary timescales, the formation of new combinations of regulators represents a major source of phenotypic novelty, facilitating new network structures. How functional, pair-wise cooperative interactions arise between regulators is poorly understood, despite the abundance of examples in extant species. Here, we explore a protein-protein interaction between two ancient transcriptional regulators-the homeodomain protein Matα2 and the MADS box protein Mcm1-that was gained approximately 200 million y ago in a clade of ascomycete yeasts that includes Saccharomyces cerevisiae. By combining deep mutational scanning with a functional selection for cooperative gene expression, we tested millions of possible alternative evolutionary solutions to this interaction interface. The artificially evolved, functional solutions are highly degenerate, with diverse amino acid chemistries permitted at all positions but with widespread epistasis limiting success. Nonetheless, approximately ~45% of the random sequences sampled function as well or better in controlling gene expression than the naturally evolved sequence. From these variants (which are unconstrained by historical contingency), we discern structural rules and epistatic constraints governing the emergence of cooperativity between these two transcriptional regulators. This work provides a mechanistic basis for long-standing observations of transcription network plasticity and highlights the importance of epistasis in the evolution of new protein-protein interactions.

Do small RNAs unlock the below ground microbiome-plant interaction mystery?

Over the past few decades, regulatory RNAs, such as small RNAs (sRNAs), have received increasing attention in the context of host-microbe interactions due to their diverse roles in controlling various biological processes in eukaryotes. In addition, studies have identified an increasing number of sRNAs with novel functions across a wide range of bacteria. What is not well understood is why cells regulate gene expression through post-transcriptional mechanisms rather than at the initiation of transcription. The finding of a multitude of sRNAs and their identified associated targets has allowed further investigation into the role of sRNAs in mediating gene regulation. These foundational data allow for further development of hypotheses concerning how a precise control of gene activity is accomplished through the combination of transcriptional and post-transcriptional regulation. Recently, sRNAs have been reported to participate in interkingdom communication and signalling where sRNAs originating from one kingdom are able to target or control gene expression in another kingdom. For example, small RNAs of fungal pathogens that silence plant genes and vice-versa plant sRNAs that mediate bacterial gene expression. However, there is currently a lack of evidence regarding sRNA-based inter-kingdom signalling across more than two interacting organisms. A habitat that provides an excellent opportunity to investigate interconnectivity is the plant rhizosphere, a multifaceted ecosystem where plants and associated soil microbes are known to interact. In this paper, we discuss how the interconnectivity of bacteria, fungi, and plants within the rhizosphere may be mediated by bacterial sRNAs with a particular focus on disease suppressive and non-suppressive soils. We discuss the potential roles sRNAs may play in the below-ground world and identify potential areas of future research, particularly in reference to the regulation of plant immunity genes by bacterial and fungal communities in disease-suppressive and non-disease-suppressive soils.

Disruption of CRTC1 and CRTC2 in Sim1 cells strongly increases high-fat diet intake in female mice but has a modest impact on male mice.

cAMP responsive element binding protein (CREB)-regulated transcription coactivators (CRTCs) regulate gene transcription in response to an increase in intracellular cAMP or Ca2+ levels. To date, three isoforms of CRTC have been identified in mammals. All CRTCs are widely expressed in various regions of the brain. Numerous studies have shown the importance of CREB and CRTC in energy homeostasis. In the brain, the paraventricular nucleus of the hypothalamus (PVH) plays a critical role in energy metabolism, and CRTC1 and CRTC2 are highly expressed in PVH neuronal cells. The single-minded homolog 1 gene (Sim1) is densely expressed in PVH neurons and in some areas of the amygdala neurons. To determine the role of CRTCs in PVH on energy metabolism, we generated mice that lacked CRTC1 and CRTC2 in Sim1 cells using Sim-1 cre mice. We found that Sim1 cell-specific CRTC1 and CRTC2 double-knockout mice were sensitive to high-fat diet (HFD)-induced obesity. Sim1 cell-specific CRTC1 and CRTC2 double knockout mice showed hyperphagia specifically for the HFD, but not for the normal chow diet, increased fat mass, and no change in energy expenditure. Interestingly, these phenotypes were stronger in female mice than in male mice, and a weak phenotype was observed in the normal chow diet. The lack of CRTC1 and CRTC2 in Sim1 cells changed the mRNA levels of some neuropeptides that regulate energy metabolism in female mice fed an HFD. Taken together, our findings suggest that CRTCs in Sim1 cells regulate gene expression and suppress excessive fat intake, especially in female mice.

A global chromatin compaction pathway that represses germline gene expression during starvation.

While much is known about how transcription is controlled at individual genes, comparatively little is known about how cells regulate gene expression on a genome-wide level. Here, we identify a molecular pathway in the C. elegans germline that controls transcription globally in response to nutritional stress. We report that when embryos hatch into L1 larvae, they sense the nutritional status of their environment, and if food is unavailable, they repress gene expression via a global chromatin compaction (GCC) pathway. GCC is triggered by the energy-sensing kinase AMPK and is mediated by a novel mechanism that involves the topoisomerase II/condensin II axis acting upstream of heterochromatin assembly. When the GCC pathway is inactivated, then transcription persists during starvation. These results define a new mode of whole-genome control of transcription.

Regulating gene expression to achieve temporal precision

Cellular response to a stimulus is often triggered by accumulation of an appropriate gene product up to a critical threshold. How cells regulate gene expression to achieve precision in timing of such responses is a question of interest. Earlier work has shown that for a stable gene product, a constant rate of accumulation provides minimum noise in the time of response, provided that initial gene product distribution is degenerate. Here, we show that this strategy is no longer optimal if the initial gene product level is drawn from a non-degenerate distribution. We also discuss biological relevance of this finding.

Functional characterization of long noncoding RNAs.

Mounting evidence suggests that long noncoding RNAs (lncRNAs) are essential regulators of gene expression. Although few lncRNAs have been the subject of detailed molecular and functional characterization, it is believed that lncRNAs play an important role in tissue homeostasis and development. In fact, gene expression profiling studies reveal lncRNAs are developmentally regulated in a tissue-type and cell-type specific manner. Such findings have brought significant attention to their potential contribution to disease cause. The current review summarizes recent studies of lncRNAs in the heart. lncRNA discovery has largely been driven by the implementation of next generation sequencing technologies. To date, such technologies have contributed to the identification of tens of thousands of distinct lncRNAs in humans -- accounting for a large majority of all RNA sequences transcribed across the human genome. Although the functions of these lncRNAs remain largely unknown, gain-of-function and loss-of-function studies (in vivo and in vitro) have uncovered a number of mechanisms by which lncRNAs regulate gene expression and protein function. Such mechanisms have been stratified according to three major functional categories: RNA sponges (RNA-mediated sequestration of free miRNAs; e.g. H19, MEG3, and MALAT1); transcription-modulating lncRNAs (RNA influences regulatory factor recruitment by binding to histone modifiers or transcription factors; e.g. CAIF, MANTIS, and NEAT1); and translation-modulating lncRNAs (RNA modifies protein function via directly interacting with a protein itself or binding partners; e.g. Airn, CCRR, and ZFAS1). Recent studies strongly suggest that lncRNAs function via binding to macromolecules (e.g. genomic DNA, miRNAs, or proteins). Thus, lncRNAs constitute an additional mode by which cells regulate gene expression.

MicroRNAs and small interfering RNAs as tools for the directed regulation of cellular processes for cancer therapy

MicroRNAs and small interfering RNAs (siRNAs) belong to an extensive class of small non-coding RNAs and play an important role in gene expression regulation in cells. It is shown that changes in the amount or activity of these molecules may lead to the development of various diseases, including cancer. This made it possible to consider them as promising diagnostic and prognostic markers, as well as tools for the directed regulation of protein synthesis in the cell and targets for therapy. This review summarizes the basic knowledge about the biogenesis, distribution and the mechanisms of action of microRNA and siRNA, as well as currently used ways of target genes expression management with their help. Possible methods of these molecules delivery into the cell in vitro and in vivo are considered.

Genomic Decoding of Neuronal Depolarization by Stimulus-Specific NPAS4 Heterodimers.

Cells regulate gene expression in response to salient external stimuli. In neurons, depolarization leads to the expression of inducible transcription factors (ITFs) that direct subsequent gene regulation. Depolarization encodes both a neuron's action potential (AP) output and synaptic inputs, via excitatory postsynaptic potentials (EPSPs). However, it is unclear if distinct types of electrical activity can be transformed by an ITF into distinct modes of genomic regulation. Here, we show that APs and EPSPs in mouse hippocampal neurons trigger two spatially segregated and molecularly distinct induction mechanisms that lead to the expression of the ITF NPAS4. These two pathways culminate in the formation of stimulus-specific NPAS4 heterodimers that exhibit distinct DNA binding patterns. Thus, NPAS4 differentially communicates increases in a neuron's spiking output and synaptic inputs to the nucleus, enabling gene regulation to be tailored to the type of depolarizing activity along the somato-dendritic axis of a neuron.

Adaptive Responses of Shewanella decolorationis to Toxic Organic Extracellular Electron Acceptor Azo Dyes in Anaerobic Respiration.

Bacterial anaerobic respiration using an extracellular electron acceptor plays a predominant role in global biogeochemical cycles. However, the mechanisms of bacterial adaptation to the toxic organic pollutant as the extracellular electron acceptor during anaerobic respiration are not clear, which limits our ability to optimize the strategies for the bioremediation of a contaminated environment. Here, we report the physiological characteristics and the global gene expression of an ecologically successful bacterium, Shewanella decolorationis S12, when using a typical toxic organic pollutant, amaranth, as the extracellular electron acceptor. Our results revealed that filamentous shift (the cells stretched to fiber-like shapes as long as 18 μm) occurred under amaranth stress. Persistent stress led to a higher filamentous cell rate and decolorization ability in subcultural cells compared to parental strains. In addition, the expression of genes involved in cell division, the chemotaxis system, energy conservation, damage repair, and material transport in filamentous cells was significantly stimulated. The detailed roles of some genes with significantly elevated expressions in filamentous cells, such as the outer membrane porin genes ompA and ompW, the cytochrome c genes arpC and arpD, the global regulatory factor gene rpoS, and the methyl-accepting chemotaxis proteins genes SHD_2793 and SHD_0015, were identified by site-directed mutagenesis. Finally, a conceptual model was proposed to help deepen our insights into both the bacterial survival strategy when toxic organics were present and the mechanisms by which these toxic organics were biodegraded as the extracellular electron acceptors.IMPORTANCE Keeping toxic organic pollutants (TOPs) in tolerable levels is a huge challenge for bacteria in extremely unfavorable environments since TOPs could serve as energy substitutes but also as survival stresses when they are beyond some thresholds. This study focused on the underlying adaptive mechanisms of ecologically successful bacterium Shewanella decolorationis S12 when exposed to amaranth, a typical toxic organic pollutant, as the extracellular electron acceptor. Our results suggest that filamentous shift is a flexible and valid way to solve the dilemma between the energy resource and toxic stress. Filamentous cells regulate gene expression to enhance their degradation and detoxification capabilities, resulting in a strong viability. These novel adaptive responses to TOPs are believed to be an evolutionary achievement to succeed in harsh habitats and thus have great potential to be applied to environment engineering or synthetic biology if we could picture every unknown node in this pathway.

Challenges of Decoding Transcription Factor Dynamics in Terms of Gene Regulation.

Technological advances are continually improving our ability to obtain more accurate views about the inner workings of biological systems. One such rapidly evolving area is single cell biology, and in particular gene expression and its regulation by transcription factors in response to intrinsic and extrinsic factors. Regarding the study of transcription factors, we discuss some of the promises and pitfalls associated with investigating how individual cells regulate gene expression through modulation of transcription factor activities. Specifically, we discuss four leading experimental approaches, the data that can be obtained from each, and important considerations that investigators should be aware of when drawing conclusions from such data.

Epigenetics: Making your mark on DNA.

Understanding the biological roles of modifications to DNA, RNA and proteins is critical to revealing how cells regulate gene expression in development and disease. Two papers now present a combination of new tools and discoveries that could enable biologists and chemical biologists to better study epigenetic regulation in mammals.

Abstract 4244: Detection and characterization of extrachromosomal circular DNA in human plasma

Abstract Background: eccDNA (extrachromosomal circular DNA) has been reported in most eukaryotes, including in human cells. However, little is known about the eccDNA profiles in human circulating system such as blood. Method: We extracted plasma cell free DNA (cfDNA) from three patients with advanced stage cancers. For each patient, we tested 10ng, 1ng and 0.1ng cfDNA, each with a technical replicate. The cfDNA samples were subjected to ATP-dependent DNase digestion and whole genome amplification (WGA). An additional 2ng cfDNA from each patient skipped the DNase digestion and went directly to WGA step. We sequenced the amplified DNAs using the Illumina 100bp PE read protocol. We analyzed sequencing data using a custom split reads-based strategy to retrieve circular DNA molecules. We further analyzed eccDNAs for their biofeatures using functional annotation tools. Results: We received sequence reads at ~14.8 million (11.4~22.4) for each sample. When read depth≥ 3 as a cutoff, we detected a total number of 2542, 1578, and 57 unique eccDNAs in three patients while negative controls showed <=5 eccDNAs. For non-DNase digested cfDNAs, we detected only 3 eccDNAs. By comparing read numbers mapped to mitochondria and human genome, we found that mitochondrial sequence reads increased significantly from 0.34-1.1% before digestion to 24.1-50.1% after digestion. Numbers of detected eccDNAs were similar between technical replicates and proportional to initial input cfDNA. However, eccDNAs detected also showed few overlaps between technical replicates. Size distribution ranged from 31bp to 19740bp (median 1801-1962bp). To test if eccDNAs were generated in specific genomic regions and if eccDNA-involved genes were enriched in specific pathways, we performed enrichment analysis and observed higher GC content in smaller eccDNAs (<500 bp, 48%) than larger ones (>500bp, 41%) (p<0.001). When compared to random distribution, eccDNAs were more likely located at CpG islands (1.36-1.53 fold enrichment). The eccDNAs were also enriched in exons, 5’UTR, 3’UTR and DNase hypersensitive sites (enrichment folds from 1.39 to 2.3). Using DAVID annotation tool, we observed significant enrichment of the eccDNA-related genes in several categories including the Coiled coil, phosphoprotein, fibronectin, suggesting a potential involvement in gene expression regulation and cell adhesion events. Using GREAT (Genomic Regions Enrichment of Annotations Tool), we found that the eccDNAs were involved in genes associated with invasive cancer cell gene expression regulation (p<0.001). Conclusion: Human plasma contains an abundant number of eccDNAs. Distribution of these eccDNAs is nonrandom event and is likely enriched in genomic regions with known functional consequences and genes that play a role in gene regulations. Further characterization of circulating eccDNAs in peripheral blood will facilitate understanding of their molecular mechanisms and potential clinical utilities. Citation Format: Jing Zhu, Meijun Du, Peng Zhang, Liang Wang. Detection and characterization of extrachromosomal circular DNA in human plasma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4244. doi:10.1158/1538-7445.AM2017-4244

Cyanobacteria Maintain Constant Protein Concentration despite Genome Copy-Number Variation

The cyanobacterium Synechococcus elongatus PCC 7942 has multiple copies of its single chromosome, and the copy number varies in individual cells, providing an ideal system to study the effect of genome copy-number variation on cell size and gene expression. Using single-cell fluorescence imaging, we found that protein concentration remained constant across individual cells regardless of genome copy number. Cell volume and the total protein amount from a single gene were both positively, linearly correlated with genome copy number, suggesting that changes in cell volume play an important role in buffering genome copy-number variance. This study provides a quantitative examination of gene expression regulation in cells with variable genome copies and sheds light on the compensation mechanisms for variance in genome copy number.

Biology, wet and dry

Sarah Teichmann's work on how cells regulate gene expression and build protein complexes recently won her a European Molecular Biology Organization Gold Medal. At 40, Teichmann holds a joint appointment with the European Bioinformatics Institute and the Wellcome Trust Sanger Institute in Hinxton, U.K. She leads a systems biology group of 17 researchers that uses both computational methods and lab experimentation. Science Careers asked Teichmann how she combines the two approaches. This interview has been edited for clarity and brevity. Q: What came first, informatics or biology? A: Biology, but informatics has always been at the heart of my research. I felt inspired by computational biology when, as an undergraduate, I read a 1992 commentary by Cyrus Chothia where he made a simple calculation of the likely number of protein families. I felt excited about discovering general principles in biology and became convinced that computational biology would provide the necessary tools. By the time I started looking for a Ph.D. in the mid-1990s, data sets had become large enough to gain a global view of the protein structure universe and of protein and gene sequences. So I went to work with Chothia at the Medical Research Council Laboratory of Molecular Biology in Cambridge, exploring protein families and the domain organization of proteins in the first completely sequenced genomes. ![Figure][1] ILLUSTRATION: ROBERT NEUBECKER > “What unifies both wet and dry work is the conceptual part of the science.” Q: Was computational biology a risky career choice? A: Yes. But I never looked back, even though at one point I came to feel that computational biology and bioinformatics were viewed as eccentric and unorthodox. My Ph.D. mentor exuded such unwavering optimism and confidence, however, that it made his lab a great place to work. Altogether, during my Ph.D., I published 10 papers. Q: What are the pros and cons of combining computational and experimental approaches? A: The beauty of a “wet-dry” group is that you have the ability to interpret your own experimental data computationally and to test computational predictions experimentally. The challenge, however, is maintaining the right balance and harnessing the potential of both sides of the group. Q: How did you learn these two approaches? A: During my first degree in the natural sciences at the University of Cambridge, I took math, physics, chemistry, and biology at a high level. I built on this through short courses and learning by doing. Today, I'd recommend more structured courses in algorithms and programming. Q: How do you feel about experimental work? A: I discovered early on that my fine motor skills are not up to some of the very tricky handling needed to be a top experimentalist in molecular biology. I have huge respect and gratitude for people who generate data through wet-lab experiments. What unifies both wet and dry work is the conceptual part of the science, which has to be logical and original, and both require exceptional dedication. Q: Is there anything you wish you had done differently? A: I made career choices mostly based on the package and colleagues I would have. In retrospect, it is also important to consider questions such as whether the scientific strategy and values of an institute are aligned with your own. I've also learned to shed my introverted nature and appreciate how important it is to talk to people often and openly. Q: Has it been hard to maintain a good work-life balance? A: Yes. I have two young daughters, and I believe that our society is a long way from empowering both men and women to balance their professional and family lives. There should be financial and cultural support for both men and women to work shorter hours for several months after the birth of an infant. Ultimately, fair and equal treatment would decrease unconscious bias in all of us. [1]: pending:yes

Readers, writers, and erasers: chromatin as the whiteboard of heart disease.

Dynamic packaging of DNA into strings of nucleosomes is a major mechanism whereby eukaryotic cells regulate gene expression. Intricate control of nucleosomal structure and assembly governs access of RNA polymerase II to DNA and consequent RNA synthesis. As part of this, post-translational modifications of histone proteins are central to the regulation of chromatin structure, playing vital roles in regulating the activation and repression of gene transcription. In the heart, dynamic homeostasis of histone modification-driven by the actions of modifiers and recruitment of downstream effectors-is a fundamental regulator of the transcriptional reprogramming that occurs in the setting of disease-related stress. Here, we examine the growing evidence for histone modification as a key mechanism governing pathological growth and remodeling of the myocardium.

Transfer RNA comes of age.

Transfer RNA comes of age.

Transcriptome wide annotation of eukaryotic RNase III reactivity and degradation signals.

Detection and validation of the RNA degradation signals controlling transcriptome stability are essential steps for understanding how cells regulate gene expression. Here we present complete genomic and biochemical annotations of the signals required for RNA degradation by the dsRNA specific ribonuclease III (Rnt1p) and examine its impact on transcriptome expression. Rnt1p cleavage signals are randomly distributed in the yeast genome, and encompass a wide variety of sequences, indicating that transcriptome stability is not determined by the recurrence of a fixed cleavage motif. Instead, RNA reactivity is defined by the sequence and structural context in which the cleavage sites are located. Reactive signals are often associated with transiently expressed genes, and their impact on RNA expression is linked to growth conditions. Together, the data suggest that Rnt1p reactivity is triggered by malleable RNA degradation signals that permit dynamic response to changes in growth conditions.

Functional Impact of RNA editing and ADARs on regulation of gene expression: perspectives from deep sequencing studies.

Cells regulate gene expression at multiple levels leading to a balance between robustness and complexity within their proteome. One core molecular step contributing to this important balance during metazoan gene expression is RNA editing, such as the co-transcriptional recoding of RNA transcripts catalyzed by the adenosine deaminse acting on RNA (ADAR) family of enzymes. Understanding of the adenosine-to-inosine RNA editing process has been broadened considerably by the next generation sequencing (NGS) technology, which allows for in-depth demarcation of an RNA editome at nucleotide resolution. However, critical issues remain unresolved with regard to how RNA editing cooperates with other transcript-associated events to underpin regulated gene expression. Here we review the growing body of evidence, provided by recent NGS-based studies, that links RNA editing to other mechanisms of post-transcriptional RNA processing and gene expression regulation including alternative splicing, transcript stability and localization, and the biogenesis and function of microRNAs (miRNAs). We also discuss the possibility that systematic integration of NGS data may be employed to establish the rules of an “RNA editing code”, which may give us new insights into the functional consequences of RNA editing.

Divergent Protein Motifs Direct Elongation Factor P-Mediated Translational Regulation in Salmonella enterica and Escherichia coli

ABSTRACTElongation factor P (EF-P) is a universally conserved bacterial translation factor homologous to eukaryotic/archaeal initiation factor 5A. In Salmonella, deletion of the efp gene results in pleiotropic phenotypes, including increased susceptibility to numerous cellular stressors. Only a limited number of proteins are affected by the loss of EF-P, and it has recently been determined that EF-P plays a critical role in rescuing ribosomes stalled at PPP and PPG peptide sequences. Here we present an unbiased in vivo investigation of the specific targets of EF-P by employing stable isotope labeling of amino acids in cell culture (SILAC) to compare the proteomes of wild-type and efp mutant Salmonella. We found that metabolic and motility genes are prominent among the subset of proteins with decreased production in the Δefp mutant. Furthermore, particular tripeptide motifs are statistically overrepresented among the proteins downregulated in efp mutant strains. These include both PPP and PPG but also additional motifs, such as APP and YIRYIR, which were confirmed to induce EF-P dependence by a translational fusion assay. Notably, we found that many proteins containing polyproline motifs are not misregulated in an EF-P-deficient background, suggesting that the factors that govern EF-P-mediated regulation are complex. Finally, we analyzed the specific region of the PoxB protein that is modulated by EF-P and found that mutation of any residue within a specific GSCGPG sequence eliminates the requirement for EF-P. This work expands the known repertoire of EF-P target motifs and implicates factors beyond polyproline motifs that are required for EF-P-mediated regulation.