Non-coding RNA regulatory networks Publication date: Available online 4 September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Simona Panni, Ruth C. Lovering, Pablo Porras, Sandra Orchard Abstract It is well established that the vast majority of human RNA transcripts do not encode for proteins and that non-coding RNAs regulate cell physiology and shape cellular functions. A subset of them is involved in gene regulation at different levels, from epigenetic gene silencing to post-transcriptional regulation of mRNA stability. Notably, the aberrant expression of many non-coding RNAs has been associated with aggressive pathologies. Rapid advances in network biology indicates that the robustness of cellular processes is the result of specific properties of biological networks such as scale-free degree distribution and hierarchical modularity, suggesting that regulatory network analyses could provide new insights on gene regulation and dysfunction mechanisms. In this study we present an overview of public repositories where non-coding RNA-regulatory interactions are collected and annotated, we discuss unresolved questions for data integration and we recall existing resources to build and analyse networks.

The how and why of lncRNA function: An innate immune perspective Publication date: Available online 2 September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms Author(s): Elektra K. Robinson, Sergio Covarrubias, Susan Carpenter Abstract Next-generation sequencing has provided a more complete picture of the composition of the human transcriptome indicating that much of the “blueprint” is a vastness of poorly understood non-protein-coding transcripts. This includes a newly identified class of genes called long noncoding RNAs (lncRNAs). The lack of sequence conservation for lncRNAs across species meant that their biological importance was initially met with some skepticism. LncRNAs mediate their functions through interactions with proteins, RNA, DNA, or a combination of these. Their functions can often be dictated by their localization, sequence, and/or secondary structure. Here we provide a review of the approaches typically adopted to study the complexity of these genes with an emphasis on recent discoveries within the innate immune field. Finally, we discuss the challenges, as well as the emergence of new technologies that will continue to move this field forward and provide greater insight into the biological importance of this class of genes. This article is part of a Special Issue entitled: ncRNA in control of gene expression edited by Kotb Abdelmohsen.

An interaction between MKL1, BRG1, and C/EBPβ mediates palmitate induced CRP transcription in hepatocytes Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Zhiwen Fan, Nan Li, Zheng Xu, Jiahao Wu, Xiangshan Fan, Yong Xu Abstract Non-alcoholic steatohepatitis (NASH) is one of the most predominant disorders in metabolic syndrome. Induction of pro-inflammatory mediators in hepatocytes exposed to free fatty acids represents a hallmark event during NASH pathogenesis. C-reactive protein (CRP) is a prototypical pro-inflammatory mediator. In the present study, we investigated the mechanism by which megakaryocytic leukemia 1 (MKL1) mediates palmitate (PA) induced CRP transcription in hepatocytes. We report that over-expression of MKL1, but not MKL2, activated the CRP promoter whereas depletion or inhibition of MKL1 repressed the CRP promoter. MKL1 potentiated the induction of the CRP promoter activity by PA treatment. Importantly, MKL1 knockdown by siRNA or pharmaceutical inhibition by CCG-1423 attenuated the induction of endogenous CRP expression in hepatocytes. Similarly, primary hepatocytes isolated from wild type (WT) mice produced more CRP than those isolated from MKL1 deficient (KO) mice when stimulated with PA. Mechanistically, the sequence-specific transcription factor CCAAT-enhancer-binding protein (C/EBPβ) interacted with MKL1 and recruited MKL1 to activate CRP transcription. Reciprocally, MKL1 modulated C/EBPβ activity by recruiting the chromatin remodeling protein BRG1 to the CRP promoter to alter histone modifications. In conclusion, our data delineate a novel epigenetic mechanism underlying augmented hepatic inflammation during NASH pathogenesis.

Tetrapeptide 60–63 of human ribosomal protein uS3 is crucial for translation initiation Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Elena Babaylova, Alexey Malygin, Alexander Gopanenko, Dmitri Graifer, Galina Karpova Abstract Conserved ribosomal protein uS3 contains a decapeptide fragment in positions 55–64 (human numbering), which has a very specific ability to cross-link to various RNA derivatives bearing aldehyde groups, likely provided by K62. It has been shown that during translation in the cell-free protein-synthesizing system, uS3 becomes accessible for such cross-linking only after eIF3j leaves the mRNA binding channel of the 40S ribosomal subunit. We studied the functional role of K62 and its nearest neighbors in the ribosomal assembly and translation with the use of HEK293T-derived cell cultures capable of producing FLAG-tagged uS3 (uS3FLAG) or its mutant form with amino acid residues at positions 60–63 replaced with alanines. Analysis of polysome profiles from the respective cells and cytosol lysates showed that the mutation significantly affected the uS3 ability to participate in the assembly of 40S subunits, but it was not essential for their maturation and did not prevent the binding of mRNAs to 40S subunits during translation initiation. The most striking effect of the replacement of amino acid residues in the above uS3 positions was that it almost completely deprived the 40S subunits of their ability to form 80S ribosomes, suggesting that the 48S pre-initiation complexes assembled on these subunits were defective in the binding of 60S subunits. Thus, our results revealed the previously unknown crucial role of the uS3 tetrapeptide 60GEKG63 in translation initiation related to maintaining the proper structure of the 48S complex, most likely via the prevention of premature mRNA loading into the ribosomal channel.

Plasticity at the DNA recognition site of the MeCP2 mCG-binding domain Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Ming Lei, Wolfram Tempel, Sizhuo Chen, Ke Liu, Jinrong Min Abstract MeCP2 is an abundant protein, involved in transcriptional repression by binding to CG and non-CG methylated DNA. However, MeCP2 might also function as a transcription activator as MeCP2 is found bound to sparsely methylated promoters of actively expressed genes. Furthermore, Attachment Region Binding Protein (ARBP), the chicken ortholog of MeCP2, has been reported to bind to Matrix/scaffold attachment regions (MARs/SARs) DNA with an unmethylated 5′-CAC/GTG-3′ consensus sequence. In our previous study, although we have systemically measured the binding abilities of MBDs to unmethylated CAC/GTG DNA and the complex structures reveal that the MBD2-MBD (MBD of MBD2) binds to the unmethylated CAC/GTG DNA by recognizing the complementary GTG trinucleotide, how the MeCP2-MBD (MBD of MeCP2) recognizes the unmethylated CAC/GTG DNA, especially the MARs DNA, is still unclear. In this study, we investigated the binding characteristics of MeCP2 in recognizing unmethylated 5′-CAC/GTG-3′ motif containing DNA by binding and structural studies. We found that MeCP2-MBD binds to MARs DNA with a comparable binding affinity to mCG DNA, and the MeCP2-CAC/GTG complex structure revealed that MeCP2 residues R111 and R133 form base-specific interactions with the GTG motif. For comparison, we also determined crystal structures of the MeCP2-MBD bound to mCG and mCAC/GTG DNA, respectively. Together, these crystal structures illustrate the adaptability of the MeCP2-MBD toward the GTG motif as well as the mCG DNA, and also provide structural basis of a biological role of MeCP2 as a transcription activator and its disease implications in Rett syndrome. Graphical abstract

Small changes, big implications: The impact of m6A RNA methylation on gene expression in pluripotency and development Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Adam M. Heck, Carol J. Wilusz Abstract In order to maintain a state of self-renewal, yet retain the ability to rapidly differentiate in response to external signals, pluripotent cells exert tight control over gene expression at many levels. Recent studies have suggested that N6-methyladenosine (m6A) RNA methylation, one of the most abundant post-transcriptional modifications, is important for both pluripotency and differentiation. In this review, we summarize the current state of the m6A field, with emphasis on the impact of writers, erasers and readers of m6A on RNA metabolism and stem cell biology.

TDP-43 and NOVA-1 RNA-binding proteins as competitive splicing regulators of the schizophrenia-associated TNIK gene Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Valentina Gumina, Claudia Colombrita, Claudia Fallini, Patrizia Bossolasco, Anna Maria Maraschi, John E. Landers, Vincenzo Silani, Antonia Ratti Abstract The RNA-binding protein TDP-43, associated to amyotrophic lateral sclerosis and frontotemporal dementia, regulates the alternative splicing of several genes, including the skipping of TNIK exon 15. TNIK, a genetic risk factor for schizophrenia and causative for intellectual disability, encodes for a Ser/Thr kinase regulating negatively F-actin dynamics. Here we show that in the human adult nervous system TNIK exon 15 is mostly included compared to the other tissues and that, during neuronal differentiation of human induced pluripotent stem cells and of human neuroblastoma cells, TNIK exon 15 inclusion increases independently of TDP-43 protein content. By studying the possible molecular interplay of TDP-43 with brain-specific splicing factors, we found that the neuronal NOVA-1 protein competitively inhibits both TDP-43 and hnRNPA2/B1 skipping activity on TNIK by means of a RNA-dependent interaction and that this competitive mechanism is common to other TDP-43 RNA targets. We also show that the TNIK protein isoforms including/excluding exon 15 differently regulate cell spreading in non-neuronal cells and neuritogenesis in primary cortical neurons. Our data suggest a complex regulation between the ubiquitous TDP-43 and the neuron-specific NOVA-1 splicing factors in the brain that may help better understand the pathobiology of both neurodegenerative diseases and schizophrenia.

HOXA2 activity regulation by cytoplasmic relocation, protein stabilization and post-translational modification Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Noémie Deneyer, Laure Bridoux, Céline Bombled, Tamara Pringels, Isabelle Bergiers, Sébastien Pyr dit Ruys, Didier Vertommen, Jean-Claude Twizere, René Rezsohazy Abstract HOX proteins are homeodomain transcription factors critically involved in patterning animal embryos and controlling organogenesis. While the functions of HOX proteins and the processes under their control begin to be well documented, the modalities of HOX protein activity regulation remain poorly understood. Here we show that HOXA2 interacts with PPP1CB, a catalytic subunit of the Ser/Thr PP1 phosphatase complex. This interaction co-localizes in the cytoplasm with a previously described HOXA2 interactor, KPC2, which belongs to the KPC E3 ubiquitin ligase complex. We provide evidence that HOXA2, PPP1CB and KPC2 define a molecularly and functionally interacting complex. Collectively, our experiments support that PPP1CB and KPC2 together inhibit the activity of HOXA2 by activating its nuclear export, but favored HOXA2 de-ubiquitination and stabilization thereby establishing a store of HOXA2 in the cytoplasm.

A genome-wide transcriptional study reveals that iron deficiency inhibits the yeast TORC1 pathway Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Antonia María Romero, Lucía Ramos-Alonso, Sandra Montellá-Manuel, José García-Martínez, María Ángeles de la Torre-Ruiz, José Enrique Pérez-Ortín, María Teresa Martínez-Pastor, Sergi Puig Abstract Iron is an essential micronutrient that participates as a cofactor in a broad range of metabolic processes including mitochondrial respiration, DNA replication, protein translation and lipid biosynthesis. Adaptation to iron deficiency requires the global reorganization of cellular metabolism directed to optimize iron utilization. The budding yeast Saccharomyces cerevisiae has been widely used to characterize the responses of eukaryotic microorganisms to iron depletion. In this report, we used a genomic approach to investigate the contribution of transcription rates to the modulation of mRNA levels during adaptation of yeast cells to iron starvation. We reveal that a decrease in the activity of all RNA polymerases contributes to the down-regulation of many mRNAs, tRNAs and rRNAs. Opposite to the general expression pattern, many genes including components of the iron deficiency response, the mitochondrial retrograde pathway and the general stress response display a remarkable increase in both transcription rates and mRNA levels upon iron limitation, whereas genes encoding ribosomal proteins or implicated in ribosome biogenesis exhibit a pronounced fall. This expression profile is consistent with an activation of the environmental stress response. The phosphorylation stage of multiple regulatory factors strongly suggests that the conserved nutrient signaling pathway TORC1 is inhibited during the progress of iron deficiency. These results suggest an intricate crosstalk between iron metabolism and the TORC1 pathway that should be considered in many disorders.

The interplay of chromatin and transcription factors during cell fate transitions in development and reprogramming Publication date: September 2019 Source: Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, Volume 1862, Issue 9 Author(s): Georgina Peñalosa-Ruiz, Ann Rose Bright, Klaas W. Mulder, Gert Jan C. Veenstra Abstract Reprogramming to induced pluripotency through expression of OCT4, SOX2, KLF4, MYC (OSKM) factors is often considered the dedifferentiation of somatic cells. This would suggest that reprogramming represents the reversal of embryonic differentiation. Indeed, molecular events involving the activity of the pluripotency network occur in opposite directions. However, reprogramming and development substantially differ as OSKM bind to accessible regulatory elements in the genome of somatic cells due to their overexpression, including regulatory elements never bound by these factors during normal differentiation. In addition, rewiring the transcriptional network back to pluripotency involves overcoming molecular barriers that protect or stabilize the somatic identity, whereas extrinsic and intrinsic cues will drive differentiation in an energetically favorable landscape in the embryo. This review focuses on how cell fate transitions in reprogramming and development are differentially governed by interactions between transcription factors and chromatin. We also discuss how these interactions shape chromatin architecture and the transcriptional output. Major technological advances have resulted in a better understanding of both differentiation and reprogramming, which is essential to exploit reprogramming regimes for regenerative medicine. Graphical abstract