Dcp2 C-terminal Cis-Binding Elements Control Selective Targeting of the Decapping Enzyme by Forming Distinct Decapping Complexes

A single Dcp1-Dcp2 decapping enzyme targets diverse classes of yeast mRNAs for decapping-dependent 5′ to 3′ decay, but the molecular mechanisms controlling selective mRNA targeting by the enzyme remain elusive. Through extensive genetic analyses we uncover cis-regulatory elements in the Dcp2 C-terminal domain that control selective targeting of the decapping enzyme by forming distinct decapping complexes. Two Upf1-binding motifs target the decapping enzyme to NMD substrates, and a single Edc3-binding motif targets both Edc3 and Dhh1 substrates. Pat1-binding leucine-rich motifs target Edc3 and Dhh1 substrates under selective conditions. Although it functions as a unique targeting component of specific complexes, Edc3 is a common component of multiple complexes. Xrn1 also has a specific Dcp2 binding site, allowing it to be directly recruited to decapping complexes. Collectively, our results demonstrate that Upf1, Edc3, and Pat1 function as regulatory subunits of the holo-decapping enzyme, controlling both its targeting specificity and enzymatic activation.

Download Full-text

Structural basis for selective targeting of Rac subfamily GTPases by a bacterial effector protein

10.1101/2020.06.29.167221 ◽

2020 ◽

Author(s):

Nikolaus Balthasar Dietz ◽

Markus Huber ◽

Isabel Sorg ◽

Arnaud Goepfert ◽

Alexander Harms ◽

...

Keyword(s):

Electrostatic Interactions ◽

Molecular Mechanisms ◽

Effector Proteins ◽

Structural Basis ◽

Effector Protein ◽

Binding Motif ◽

Selective Targeting ◽

Rho Family Gtpases ◽

Rho Family ◽

Specific Pair

SummaryRas-homology (Rho) family GTPases are conserved molecular switches controlling fundamental cellular activities in eukaryotic cells. As such, they are targeted by numerous bacterial toxins and effector proteins, which have been intensively investigated regarding their biochemical activities and discrete target spectra; however, molecular mechanisms of target selectivity have remained elusive. Here, we report a bacterial effector protein that targets all four Rac subfamily members of Rho family GTPases, but none of the closely related Cdc42 or RhoA subfamilies. This exquisite target selectivity of the FIC domain AMP-transferase Bep1 from Bartonella rochalimae is based on electrostatic interactions with a subfamily-specific pair of residues in the nucleotide-binding motif and the Rho insert helix. Residue substitutions at the identified positions in Cdc42 facilitate modification by Bep1, while corresponding Cdc42-like substitutions in Rac1 greatly diminish modification. Our study establishes a structural paradigm for target selectivity towards Rac subfamily GTPases and provides a highly selective tool for their functional analysis.

Download Full-text

Mapping chromatin accessibility and active regulatory elements reveals pathological mechanisms in human gliomas

Nature Communications ◽

10.1038/s41467-021-23922-2 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Karolina Stępniak ◽

Magdalena A. Machnicka ◽

Jakub Mieczkowski ◽

Anna Macioszek ◽

Bartosz Wojtaś ◽

...

Keyword(s):

Gene Expression ◽

Chromatin Structure ◽

Molecular Mechanisms ◽

Genome Mapping ◽

Malignant Gliomas ◽

Regulatory Elements ◽

Chromatin Accessibility ◽

Multiple Tumor ◽

Genes Encoding ◽

Methylation Patterns

AbstractChromatin structure and accessibility, and combinatorial binding of transcription factors to regulatory elements in genomic DNA control transcription. Genetic variations in genes encoding histones, epigenetics-related enzymes or modifiers affect chromatin structure/dynamics and result in alterations in gene expression contributing to cancer development or progression. Gliomas are brain tumors frequently associated with epigenetics-related gene deregulation. We perform whole-genome mapping of chromatin accessibility, histone modifications, DNA methylation patterns and transcriptome analysis simultaneously in multiple tumor samples to unravel epigenetic dysfunctions driving gliomagenesis. Based on the results of the integrative analysis of the acquired profiles, we create an atlas of active enhancers and promoters in benign and malignant gliomas. We explore these elements and intersect with Hi-C data to uncover molecular mechanisms instructing gene expression in gliomas.

Download Full-text

Epigenomic Analysis of RAD51 ChIP-seq Data Reveals cis-regulatory Elements Associated with Autophagy in Cancer Cell Lines

Cancers ◽

10.3390/cancers13112547 ◽

2021 ◽

Vol 13 (11) ◽

pp. 2547

Author(s):

Keunsoo Kang ◽

Yoonjung Choi ◽

Hyeonjin Moon ◽

Chaelin You ◽

Minjin Seo ◽

...

Keyword(s):

Gene Regulation ◽

Homologous Recombination ◽

Cell Lines ◽

Regulatory Elements ◽

Binding Motif ◽

Genome Wide ◽

E Box ◽

Autophagy Pathway ◽

Mcf 7

RAD51 is a recombinase that plays a pivotal role in homologous recombination. Although the role of RAD51 in homologous recombination has been extensively studied, it is unclear whether RAD51 can be involved in gene regulation as a co-factor. In this study, we found evidence that RAD51 may contribute to the regulation of genes involved in the autophagy pathway with E-box proteins such as USF1, USF2, and/or MITF in GM12878, HepG2, K562, and MCF-7 cell lines. The canonical USF binding motif (CACGTG) was significantly identified at RAD51-bound cis-regulatory elements in all four cell lines. In addition, genome-wide USF1, USF2, and/or MITF-binding regions significantly coincided with the RAD51-associated cis-regulatory elements in the same cell line. Interestingly, the promoters of genes associated with the autophagy pathway, such as ATG3 and ATG5, were significantly occupied by RAD51 and regulated by RAD51 in HepG2 and MCF-7 cell lines. Taken together, these results unveiled a novel role of RAD51 and provided evidence that RAD51-associated cis-regulatory elements could possibly be involved in regulating autophagy-related genes with E-box binding proteins.

Download Full-text

A Genome-Wide Analysis of Pathogenesis-Related Protein-1 (PR-1) Genes from Piper nigrum Reveals Its Critical Role during Phytophthora capsici Infection

Genes ◽

10.3390/genes12071007 ◽

2021 ◽

Vol 12 (7) ◽

pp. 1007

Author(s):

Divya Kattupalli ◽

Asha Sreenivasan ◽

Eppurathu Vasudevan Soniya

Keyword(s):

Phytophthora Capsici ◽

Regulatory Elements ◽

Piper Nigrum ◽

Binding Motif ◽

Altered Expression ◽

Genome Wide ◽

A Genome ◽

Pathogenesis Related Protein ◽

Pathogenesis Related

Black pepper (Piper nigrum L.) is a prominent spice that is an indispensable ingredient in cuisine and traditional medicine. Phytophthora capsici, the causative agent of footrot disease, causes a drastic constraint in P. nigrum cultivation and productivity. To counterattack various biotic and abiotic stresses, plants employ a broad array of mechanisms that includes the accumulation of pathogenesis-related (PR) proteins. Through a genome-wide survey, eleven PR-1 genes that belong to a CAP superfamily protein with a caveolin-binding motif (CBM) and a CAP-derived peptide (CAPE) were identified from P. nigrum. Despite the critical functional domains, PnPR-1 homologs differ in their signal peptide motifs and core amino acid composition in the functional protein domains. The conserved motifs of PnPR-1 proteins were identified using MEME. Most of the PnPR-1 proteins were basic in nature. Secondary and 3D structure analyses of the PnPR-1 proteins were also predicted, which may be linked to a functional role in P. nigrum. The GO and KEGG functional annotations predicted their function in the defense responses of plant-pathogen interactions. Furthermore, a transcriptome-assisted FPKM analysis revealed PnPR-1 genes mapped to the P. nigrum-P. capsici interaction pathway. An altered expression pattern was detected for PnPR-1 transcripts among which a significant upregulation was noted for basic PnPR-1 genes such as CL10113.C1 and Unigene17664. The drastic variation in the transcript levels of CL10113.C1 was further validated through qRT-PCR and it showed a significant upregulation in infected leaf samples compared with the control. A subsequent analysis revealed the structural details, phylogenetic relationships, conserved sequence motifs and critical cis-regulatory elements of PnPR-1 genes. This is the first genome-wide study that identified the role of PR-1 genes during P. nigrum-P. capsici interactions. The detailed in silico experimental analysis revealed the vital role of PnPR-1 genes in regulating the first layer of defense towards a P. capsici infection in Panniyur-1 plants.

Download Full-text

Exploring functionally annotated transcriptional consensus regulatory elements with CONREL

Database ◽

10.1093/database/baaa071 ◽

2020 ◽

Vol 2020 ◽

Author(s):

Davide Dalfovo ◽

Samuel Valentini ◽

Alessandro Romanel

Keyword(s):

Cell Lines ◽

Web Application ◽

Regulatory Networks ◽

Regulatory Elements ◽

Levels Of Abstraction ◽

Binding Motifs ◽

Extensive Collection ◽

Transcription Factor Binding Motifs ◽

Gene Regulatory ◽

Genomic Regions

Abstract Understanding the interaction between human genome regulatory elements and transcription factors is fundamental to elucidate the structure of gene regulatory networks. Here we present CONREL, a web application that allows for the exploration of functionally annotated transcriptional ‘consensus’ regulatory elements at different levels of abstraction. CONREL provides an extensive collection of consensus promoters, enhancers and active enhancers for 198 cell-lines across 38 tissue types, which are also combined to provide global consensuses. In addition, 1000 Genomes Project genotype data and the ‘total binding affinity’ of thousands of transcription factor binding motifs at genomic regulatory elements is fully combined and exploited to characterize and annotate functional properties of our collection. Comparison with other available resources highlights the strengths and advantages of CONREL. CONREL can be used to explore genomic loci, specific genes or genomic regions of interest across different cell lines and tissue types. The resource is freely available at https://bcglab.cibio.unitn.it/conrel.

Download Full-text

Evolution of mouse circadian enhancers from transposable elements

Genome Biology ◽

10.1186/s13059-021-02409-9 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Julius Judd ◽

Hayley Sanderson ◽

Cédric Feschotte

Keyword(s):

Gene Expression ◽

Transposable Elements ◽

Binding Sites ◽

Mouse Liver ◽

Integration Site ◽

Point Mutations ◽

Regulatory Elements ◽

Circadian Gene ◽

Binding Motifs ◽

Reporter Assays

Abstract Background Transposable elements are increasingly recognized as a source of cis-regulatory variation. Previous studies have revealed that transposons are often bound by transcription factors and some have been co-opted into functional enhancers regulating host gene expression. However, the process by which transposons mature into complex regulatory elements, like enhancers, remains poorly understood. To investigate this process, we examined the contribution of transposons to the cis-regulatory network controlling circadian gene expression in the mouse liver, a well-characterized network serving an important physiological function. Results ChIP-seq analyses reveal that transposons and other repeats contribute ~ 14% of the binding sites for core circadian regulators (CRs) including BMAL1, CLOCK, PER1/2, and CRY1/2, in the mouse liver. RSINE1, an abundant murine-specific SINE, is the only transposon family enriched for CR binding sites across all datasets. Sequence analyses and reporter assays reveal that the circadian regulatory activity of RSINE1 stems from the presence of imperfect CR binding motifs in the ancestral RSINE1 sequence. These motifs matured into canonical motifs through point mutations after transposition. Furthermore, maturation occurred preferentially within elements inserted in the proximity of ancestral CR binding sites. RSINE1 also acquired motifs that recruit nuclear receptors known to cooperate with CRs to regulate circadian gene expression specifically in the liver. Conclusions Our results suggest that the birth of enhancers from transposons is predicated both by the sequence of the transposon and by the cis-regulatory landscape surrounding their genomic integration site.

Download Full-text

Receptor and Molecular Mechanism of AGGF1 Signaling in Endothelial Cell Functions and Angiogenesis

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvbaha.121.316867 ◽

2021 ◽

Author(s):

Jingjing Wang ◽

Huixin Peng ◽

Ayse Anil Timur ◽

Vinay Pasupuleti ◽

Yufeng Yao ◽

...

Keyword(s):

Amino Acids ◽

Endothelial Cell ◽

Neutralizing Antibodies ◽

Molecular Mechanisms ◽

Capillary Tube ◽

Therapeutic Angiogenesis ◽

Angiogenic Factor ◽

Single Amino Acid ◽

Binding Motif ◽

Cell Functions

Objective: Angiogenic factor AGGF1 (angiogenic factor and G-patch and FHA [Forkhead-associated] domain 1) promotes angiogenesis as potently as VEGFA (vascular endothelial growth factor A) and regulates endothelial cell (EC) proliferation, migration, specification of multipotent hemangioblasts and venous ECs, hematopoiesis, and vascular development and causes vascular disease Klippel-Trenaunay syndrome when mutated. However, the receptor for AGGF1 and the underlying molecular mechanisms remain to be defined. Approach and Results: Using functional blocking studies with neutralizing antibodies, we identified α5β1 as the receptor for AGGF1 on ECs. AGGF1 interacts with α5β1 and activates FAK (focal adhesion kinase), Src, and AKT. Functional analysis of 12 serial N-terminal deletions and 13 C-terminal deletions by every 50 amino acids mapped the angiogenic domain of AGGF1 to a domain between amino acids 604-613 (FQRDDAPAS). The angiogenic domain is required for EC adhesion and migration, capillary tube formation, and AKT activation. The deletion of the angiogenic domain eliminated the effects of AGGF1 on therapeutic angiogenesis and increased blood flow in a mouse model for peripheral artery disease. A 40-mer or 15-mer peptide containing the angiogenic domain blocks AGGF1 function, however, a 15-mer peptide containing a single amino acid mutation from −RDD- to −RGD- (a classical RGD integrin-binding motif) failed to block AGGF1 function. Conclusions: We have identified integrin α5β1 as an EC receptor for AGGF1 and a novel AGGF1-mediated signaling pathway of α5β1-FAK-Src-AKT for angiogenesis. Our results identify an FQRDDAPAS angiogenic domain of AGGF1 crucial for its interaction with α5β1 and signaling.

Download Full-text

Tyrosine-protein kinase Src regulates Kv1.5 channel activity and membrane expression through interaction with the N-terminus of the channel

Inquiry@Queen's Undergraduate Research Conference Proceedings ◽

10.24908/iqurcp.14024 ◽

2020 ◽

Author(s):

Taylore Dodd ◽

Tingzhong Wang ◽

Shetuan Zhang

Keyword(s):

Protein Kinase ◽

Cell Voltage ◽

Underlying Mechanism ◽

Delayed Rectifier ◽

Binding Motif ◽

Membrane Expression ◽

Binding Motifs ◽

Delayed Rectifier Potassium Current ◽

N Terminus ◽

Tyrosine Protein Kinase

Kv1.5 is a voltage-gated potassium channel that generates the ultra-rapid delayed rectifier potassium current (IKur) important in the repolarization of the atrial action potential. Malfunction of the Kv1.5 channel often results in atrial fibrillation (AFib). A reduction in Kv1.5 current (IKv1.5) occurs upon activation of the endogenous tyrosine-protein kinase Src. The Src SH3 domain binds to proline-rich motifs located within the N-terminus of Kv1.5. Disruption of these binding motifs has been involved in the development of familial AFib. The mechanism underlying the reduction of IKv1.5 upon Src activation has not yet been established and the relationship between Kv1.5 and Src is poorly understood. Therefore, the present study aims to further elucidate the mechanism behind IKv1.5 reduction. The hypothesis that Src regulates Kv1.5 activity by altering the density of mature membrane-localized channels was tested using whole-cell voltage clamp and Western blot analysis. We demonstrate that Src tonically inhibits Kv1.5 activity and decreases the density of mature membrane-localized channels. Kv1.5 channels possessing mutations within the Src binding motifs were also investigated and it was determined that each binding motif contributes to the Kv1.5-Src relationship, however, the binding of Src to an individual motif is sufficiently effective. Our findings indicate that Src regulates Kv1.5 through an interaction with the N-terminal binding motifs and suggests that the inhibition of forward trafficking may be involved in the underlying mechanism. (Supported by the Heart and Stroke foundation of Canada and The Canadian Institutes of Health Research).

Download Full-text

Transcription of the Human 5-Hydroxytryptamine Receptor 2B (HTR2B) Gene Is under the Regulatory Influence of the Transcription Factors NFI and RUNX1 in Human Uveal Melanoma

International Journal of Molecular Sciences ◽

10.3390/ijms19103272 ◽

2018 ◽

Vol 19 (10) ◽

pp. 3272 ◽

Cited By ~ 4

Author(s):

Manel Benhassine ◽

Sylvain Guérin

Keyword(s):

Transcription Factors ◽

Uveal Melanoma ◽

Serotonin Receptor ◽

Molecular Mechanisms ◽

Regulatory Region ◽

Gene Signature ◽

Regulatory Elements ◽

Gene Encoding ◽

Aberrant Expression ◽

Factor I

Because it accounts for 70% of all eye cancers, uveal melanoma (UM) is therefore the most common primary ocular malignancy. In this study, we investigated the molecular mechanisms leading to the aberrant expression of the gene encoding the serotonin receptor 2B (HTR2B), one of the most discriminating among the candidates from the class II gene signature, in metastatic and non-metastatic UM cell lines. Transfection analyses revealed that the upstream regulatory region of the HTR2B gene contains a combination of alternative positive and negative regulatory elements functional in HTR2B− but not in HTR23B+ UM cells. We demonstrated that both the transcription factors nuclear factor I (NFI) and Runt-related transcription factor I (RUNX1) interact with regulatory elements from the HTR2B gene to either activate (NFI) or repress (RUNX1) HTR2B expression in UM cells. The results of this study will help understand better the molecular mechanisms accounting for the abnormal expression of the HTR2B gene in uveal melanoma.

Download Full-text

Non-Coding Genome, Transcription Factors, and Sex Determination

Sexual Development ◽

10.1159/000519725 ◽

2021 ◽

pp. 1-13

Author(s):

Francis Poulat

Keyword(s):

Transcription Factors ◽

Sex Determination ◽

Target Genes ◽

Regulatory Elements ◽

Eukaryotic Cells ◽

Genomic Profiling ◽

Binding Motifs ◽

Male And Female ◽

Genome Transcription ◽

Control Complex

In vertebrates, gonadal sex determination is the process by which transcription factors drive the choice between the testicular and ovarian identity of undifferentiated somatic progenitors through activation of 2 different transcriptional programs. Studies in animal models suggest that sex determination always involves sex-specific transcription factors that activate or repress sex-specific genes. These transcription factors control their target genes by recognizing their regulatory elements in the non-coding genome and their binding motifs within their DNA sequence. In the last 20 years, the development of genomic approaches that allow identifying all the genomic targets of a transcription factor in eukaryotic cells gave the opportunity to globally understand the function of the nuclear proteins that control complex genetic programs. Here, the major transcription factors involved in male and female vertebrate sex determination and the genomic profiling data of mouse gonads that contributed to deciphering their transcriptional regulation role will be reviewed.

Download Full-text