Structures of SF3b1 reveal a dynamic Achilles heel of spliceosome assembly: Implications for cancer-associated abnormalities and drug discovery Publication date: Available online 9 November 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Debanjana Maji, Alan M. Grossfield, Clara L. Kielkopf Abstract The pre-mRNA splicing factor SF3b1 exhibits recurrent mutations among hematologic malignancies and cancers, and consequently is a major therapeutic target of clinically-advanced spliceosome inhibitors. In this review, we highlight and rigorously analyze emerging views of SF3b1 conformational transitions, including the human SF3b particle either in isolation or bound to spliceosome inhibitors, and human or yeast spliceosome assemblies. Among spliceosome states characterized to date, an SF3b1 α-helical superhelix significantly closes to surround a U2 small nuclear RNA duplex with the pre-mRNA branch point sequence. The SF3b1 torus is locally unwound at an active site adenosine, whereas protein cofactors appear to stabilize overall closure in the spliceosome. Network analyses demonstrates that the natural SF3b1 dynamics mimic its conformational change in the spliceosome, raising the possibility of conformational selection underpinning spliceosome assembly. These dynamic SF3b1 conformations have consequences for gatekeeping of spliceosome assembly and therapeutic targeting of its cancer-associated dysfunction.

Small Drosophila zinc finger C2H2 protein with an N-terminal zinc finger-associated domain demonstrates the architecture functions Publication date: Available online 6 November 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Oksana Maksimenko, Olga Kyrchanova, Natalia Klimenko, Nikolay Zolotarev, Anna Elizarova, Artem Bonchuk, Pavel Georgiev Abstract Recently, the concept has arisen that a special class of architectural proteins exists, which are responsible not only for global chromosome architecture but also for the local regulation of enhancer–promoter interactions. Here, we describe a new architectural protein, with a total size of only 375 aa, which contains an N-terminal zinc finger-associated domain (ZAD) and a cluster of five zinc finger C2H2 domains at the C-terminus. This new protein, named ZAD and Architectural Function 1 protein (ZAF1 protein), is weakly and ubiquitously expressed, with the highest expression levels observed in oocytes and embryos. The cluster of C2H2 domains recognizes a specific 15-bp consensus site, located predominantly in promoters, near transcription start sites. The expression of ZAF1 by a tissue-specific promoter led to the complete blocking of the eye enhancer when clusters of ZAF1 binding sites flanked the eye enhancer in transgenic lines, suggesting that the loop formed by the ZAF1 protein leads to insulation. The ZAF1 protein also supported long-range interactions between the yeast GAL4 activator and the white promoter in transgenic Drosophila lines. A mutant protein lacking the ZAD failed to block the eye enhancer or to support distance interactions in transgenic lines. Taken together, these results suggest that ZAF1 is a minimal architectural protein that can be used to create a convenient model for studying the mechanisms of distance interactions.

Splicing in the pathogenesis, diagnosis and treatment of ciliopathies Publication date: Available online 4 November 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Gabrielle Wheway, Jenny Lord, Diana Baralle Abstract Primary cilia are essential signalling organelles found on the apical surface of epithelial cells, where they coordinate chemosensation, mechanosensation and light sensation. Motile cilia play a central role in establishing fluid flow in the respiratory tract, reproductive tract, brain ventricles and ear. Genetic defects affecting the structure or function of cilia can lead to a broad range of developmental and degenerative diseases known as ciliopathies. Splicing contributes to the pathogenesis, diagnosis and treatment of ciliopathies. Tissue-specific alternative splicing contributes to the tissue-specific manifestation of ciliopathy phenotypes, for example the retinal-specific effects of some genetic defects, due to specific transcript expression in the highly specialised ciliated cells of the retina, the photoreceptor cells. Ciliopathies can arise both as a result of genetic variants in spliceosomal proteins, or as a result of variants affecting splicing of specific cilia genes. Here we discuss the opportunities and challenges in diagnosing ciliopathies using RNA sequence analysis and the potential for treating ciliopathies in a relatively mutation-neutral way by targeting splicing. This article is part of a Special Issue entitled: RNA structure and splicing regulation edited by Francisco Baralle, Ravindra Singh and Stefan Stamm.

Oxoglutarate dehydrogenase and acetyl-CoA acyltransferase 2 selectively associate with H2A.Z-occupied promoters and are required for histone modifications Publication date: Available online 1 November 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Sujung Choi, Jessica Pfleger, Yong Heui Jeon, Zhi Yang, Minzhen He, Hyewon Shin, Danish Sayed, Sophie Astrof, Maha Abdellatif Abstract Histone H2A.Z plays an essential role in regulating transcriptional rates and memory. Interestingly, H2A.Z-bound nucleosomes are located in both transcriptionally active and inactive promotors, with no clear understanding of the mechanisms via which it differentially regulates transcription. We hypothesized that its functions are mediated through recruitment of regulatory proteins to promoters. Using rapid chromatin immunoprecipitation-mass spectrometry, we uncovered the association of H2A.Z-bound chromatin with the metabolic enzymes, oxoglutarate dehydrogenase (OGDH) and acetyl-CoA acyltransferase 2 (ACAA2). Recombinant green florescence fusion proteins, combined with mutations of predicted nuclear localization signals, confirmed their nuclear localization and chromatin binding. Conclusively, chromatin immunoprecipitation-deep sequencing, confirmed the predominant association of OGDH and ACAA2 with H2A.Z-occupied transcription start sites and enhancers, the former of which we confirmed is conserved in both mouse and human tissue. Furthermore, H2A.Z-deficient human HAP1 cells exhibited reduced chromatin-bound metabolic enzymes, accompanied with reduced posttranslational histone modifications, including acetylation and succinylation. Specifically, knockdown of OGDH diminished H4 succinylation. Thus, the data reveal that select metabolic enzymes are assembled at active, H2A.Z-occupied, promoters, for potential site-directed production of metabolic intermediates that are required for histone modifications.

Intronic RNA: Ad‘junk’ mediator of post-transcriptional gene regulation Publication date: Available online 1 November 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Christopher R. Neil, William G. Fairbrother Abstract RNA splicing, the process through which intervening segments of noncoding RNA (introns) are excised from pre-mRNAs to allow for the formation of a mature mRNA product, has long been appreciated for its capacity to add complexity to eukaryotic proteomes. However, evidence suggests that the utility of this process extends beyond protein output and provides cells with a dynamic tool for gene regulation. In this review, we aim to highlight the role that intronic RNA plays in mediating specific splicing outcomes in pre-mRNA processing, as well as explore an emerging class of stable intronic sequences that have been observed to act in gene expression control. Building from underlying flexibility in both sequence and structure, intronic RNA provides mechanisms for post-transcriptional gene regulation that are amenable to the tissue and condition specific needs of eukaryotic cells. This article is part of a Special Issue entitled: RNA structure and splicing regulation edited by Francisco Baralle, Ravindra Singh and Stefan Stamm.

Differential chamber-specific expression and regulation of long non-coding RNAs during cardiac development Publication date: Available online 1 November 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Carlos García-Padilla, Jorge N. Domínguez, Amelia E. Aránega, Diego Franco Abstract Cardiovascular development is governed by a complex interplay between inducting signals such as Bmps and Fgfs leading to activation of cardiac specific transcription factors such as Nkx2.5, Mef2c and Srf that orchestrate the initial steps of cardiogenesis. Over the last decade we have witnessed the discovery of novel layers of gene regulation, i.e. post-transcriptional regulation exerted by non-coding RNAs. The function role of small non coding RNAs has been widely demonstrated, e.g. miR-1 knockout display several cardiovascular abnormalities during embryogenesis. More recently long non-coding RNAs have been also reported to modulate gene expression and function in the developing heart, as exemplified by the embryonic lethal phenotypes of Fendrr and Braveheart knock out mice, respectively. In this study, we investigated the differential expression profile during cardiogenesis of previously reported lncRNAs in heart development. Our data revealed that Braveheart, Fendrr, Carmen display a preferential adult expression while Miat, Alien, H19 preferentially display chamber-specific expression at embryonic stages. We also demonstrated that these lncRNAs are differentially regulated by Nkx2.5, Srf and Mef2c, Pitx2 > Wnt > miRNA signaling pathway and angiotensin II and thyroid hormone administration. Importantly isoform-specific expression and distinct nuclear vs cytoplasmic localization of Braveheart, Carmen and Fendrr during chamber morphogenesis is observed, suggesting distinct functional roles of these lncRNAs in atrial and ventricular chambers. Furthermore, we demonstrate by in situ hybridization a dynamic epicardial, myocardial and endocardial expression of H19 during cardiac development. Overall our data support novel roles of these lncRNAs in different temporal and tissue-restricted fashion during cardiogenesis.

Inference of plant gene regulatory networks using data-driven methods: A practical overview Publication date: Available online 31 October 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Shubhada R. Kulkarni, Klaas Vandepoele Abstract Transcriptional regulation is a complex and dynamic process that plays a vital role in plant growth and development. A key component in the regulation of genes is transcription factors (TFs), which coordinate the transcriptional control of gene activity. A gene regulatory network (GRN) is a collection of regulatory interactions between TFs and their target genes. The accurate delineation of GRNs offers a significant contribution to our understanding about how plant cells are organized and function, and how individual genes are regulated in various conditions, organs or cell types. During the past decade, important progress has been made in the identification of GRNs using experimental and computational approaches. However, a detailed overview of available platforms supporting the analysis of GRNs in plants is missing. Here, we review current databases, platforms and tools that perform data-driven analyses of gene regulation in Arabidopsis. The platforms are categorized into two sections, 1) promoter motif analysis tools that use motif mapping approaches to find TF motifs in the regulatory sequences of genes of interest and 2) network analysis tools that identify potential regulators for a set of input genes using a range of data types in order to generate GRNs. We discuss the diverse datasets integrated and highlight the strengths and caveats of different platforms. Finally, we shed light on the limitations of the above approaches and discuss future perspectives, including the need for integrative approaches to unravel complex GRNs in plants.

Gene regulatory network inference resources: A practical overview Publication date: Available online 31 October 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): D. Mercatelli, L. Scalambra, L. Triboli, F. Ray, F.M. Giorgi Abstract Transcriptional regulation is a fundamental molecular mechanism involved in almost every aspect of life, from homeostasis to development, from metabolism to behavior, from reaction to stimuli to disease progression. In recent years, the concept of Gene Regulatory Networks (GRNs) has grown popular as an effective applied biology approach for describing the complex and highly dynamic set of transcriptional interactions, due to its easy-to-interpret features. Since cataloguing, predicting and understanding every GRN connection in all species and cellular contexts remains a great challenge for biology, researchers have developed numerous tools and methods to infer regulatory processes. In this review, we catalogue these methods in six major areas, based on the dominant underlying information leveraged to infer GRNs: Coexpression, Sequence Motifs, Chromatin Immunoprecipitation (ChIP), Orthology, Literature and Protein-Protein Interaction (PPI) specifically focused on transcriptional complexes. The methods described here cover a wide range of user-friendliness: from web tools that require no prior computational expertise to command line programs and algorithms for large scale GRN inferences. Each method for GRN inference described herein effectively illustrates a type of transcriptional relationship, with many methods being complementary to others. While a truly holistic approach for inferring and displaying GRNs remains one of the greatest challenges in the field of systems biology, we believe that the integration of multiple methods described herein provides an effective means with which experimental and computational biologists alike may obtain the most complete pictures of transcriptional relationships. This article is part of a Special Issue entitled: Transcriptional Profiles and Regulatory Gene Networks edited by Dr. Dr. Federico Manuel Giorgi and Dr. Shaun Mahony.

Merging 1D and 3D genomic information: Challenges in modelling and validation Publication date: Available online 28 October 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Alessandra Merlotti, Angelo Rosa, Daniel Remondini Abstract Genome organization in eukaryotes during interphase stems from the delicate balance between non-random correlations present in the DNA polynucleotide linear sequence and the physico/chemical reactions which shape continuously the form and structure of DNA and chromatin inside the nucleus of the cell. It is now clear that these mechanisms have a key role in important processes like gene regulation, yet the detailed ways they act simultaneously and, eventually, come to influence each other even across very different length-scales remain largely unexplored. In this paper, we recapitulate some of the main results concerning gene regulatory and physical mechanisms, in relation to the information encoded in the 1D sequence and the 3D folding structure of DNA. In particular, we stress how reciprocal crossfeeding between 1D and 3D models may provide original insight into how these complex processes work and influence each other. This article is part of a Special Issue entitled: Transcriptional Profiles and Regulatory Gene Networks edited by Dr. Dr. Federico Manuel Giorgi and Dr. Shaun Mahony. Graphical Abstract

TDP-43 regulates transcription at protein-coding genes and Alu retrotransposons Publication date: Available online 23 October 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Andrés A. Morera, Nasiha S. Ahmed, Jacob C. Schwartz Abstract The 43-kDa transactive response DNA-binding protein (TDP-43) is an example of an RNA-binding protein that regulates RNA metabolism at multiple levels from transcription and splicing to translation. Its role in post-transcriptional RNA processing has been a primary focus of recent research, but its role in regulating transcription has been studied for only a few human genes. We characterized the effects of TDP-43 on transcription genome-wide and found that TDP-43 broadly affects transcription of protein-coding and noncoding RNA genes. Among protein-coding genes, the effects of TDP-43 were greatest for genes <30 thousand base pairs in length. Surprisingly, we found that the loss of TDP-43 resulted in increased evidence for transcription activity near repetitive Alu elements found within expressed genes. The highest densities of affected Alu elements were found in the shorter genes, whose transcription was most affected by TDP-43. Thus, in addition to its role in post-transcriptional RNA processing, TDP-43 plays a critical role in maintaining the transcriptional stability of protein-coding genes and transposable DNA elements.