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Article ; Online: NELF focuses sites of initiation and maintains promoter architecture.

Santana, Juan F / Spector, Benjamin M / Suarez, Gustavo A / Luse, Donal S / Price, David H

2024 Volume 52, Issue 6, Page(s) 2977–2994

Abstract: Many factors control the elongation phase of transcription by RNA polymerase II (Pol II), a process that plays an essential role in regulating gene expression. We utilized cells expressing degradation tagged subunits of NELFB, PAF1 and RTF1 to probe the ... ...

Abstract	Many factors control the elongation phase of transcription by RNA polymerase II (Pol II), a process that plays an essential role in regulating gene expression. We utilized cells expressing degradation tagged subunits of NELFB, PAF1 and RTF1 to probe the effects of depletion of the factors on nascent transcripts using PRO-Seq and on chromatin architecture using DFF-ChIP. Although NELF is involved in promoter proximal pausing, depletion of NELFB had only a minimal effect on the level of paused transcripts and almost no effect on control of productive elongation. Instead, NELF depletion increased the utilization of downstream transcription start sites and caused a dramatic, genome-wide loss of H3K4me3 marked nucleosomes. Depletion of PAF1 and RTF1 both had major effects on productive transcript elongation in gene bodies and also caused initiation site changes like those seen with NELFB depletion. Our study confirmed that the first nucleosome encountered during initiation and early elongation is highly positioned with respect to the major TSS. In contrast, the positions of H3K4me3 marked nucleosomes in promoter regions are heterogeneous and are influenced by transcription. We propose a model defining NELF function and a general role of the H3K4me3 modification in blocking transcription initiation.
MeSH term(s)	Transcription, Genetic ; Nucleosomes/genetics ; RNA Polymerase II/genetics ; RNA Polymerase II/metabolism ; Promoter Regions, Genetic
Chemical Substances	Nucleosomes ; RNA Polymerase II (EC 2.7.7.-)
Language	English
Publishing date	2024-01-10
Publishing country	England
Document type	Journal Article
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkad1253
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Article ; Online: Differential dependencies of human RNA polymerase II promoters on TBP, TAF1, TFIIB and XPB.

Santana, Juan F / Collins, Geoffrey S / Parida, Mrutyunjaya / Luse, Donal S / Price, David H

Nucleic acids research

2022 Volume 50, Issue 16, Page(s) 9127–9148

Abstract: The effects of rapid acute depletion of components of RNA polymerase II (Pol II) general transcription factors (GTFs) that are thought to be critical for formation of preinitiation complexes (PICs) and initiation in vitro were quantified in HAP1 cells ... ...

Abstract	The effects of rapid acute depletion of components of RNA polymerase II (Pol II) general transcription factors (GTFs) that are thought to be critical for formation of preinitiation complexes (PICs) and initiation in vitro were quantified in HAP1 cells using precision nuclear run-on sequencing (PRO-Seq). The average dependencies for each factor across >70 000 promoters varied widely even though levels of depletions were similar. Some of the effects could be attributed to the presence or absence of core promoter elements such as the upstream TBP-specificity motif or downstream G-rich sequences, but some dependencies anti-correlated with such sequences. While depletion of TBP had a large effect on most Pol III promoters only a small fraction of Pol II promoters were similarly affected. TFIIB depletion had the largest general effect on Pol II and also correlated with apparent termination defects downstream of genes. Our results demonstrate that promoter activity is combinatorially influenced by recruitment of TFIID and sequence-specific transcription factors. They also suggest that interaction of the preinitiation complex (PIC) with nucleosomes can affect activity and that recruitment of TFIID containing TBP only plays a positive role at a subset of promoters.
MeSH term(s)	Humans ; Transcription Factor TFIIB/genetics ; Transcription Factor TFIIB/metabolism ; RNA Polymerase II/genetics ; RNA Polymerase II/metabolism ; Promoter Regions, Genetic ; Transcription Factors/genetics ; Transcription Factor TFIID/genetics ; Transcription Factor TFIID/metabolism ; TATA-Box Binding Protein/genetics ; TATA-Box Binding Protein/metabolism ; Transcription, Genetic ; TATA Box/genetics ; RNA Polymerase III/genetics
Chemical Substances	Transcription Factor TFIIB ; RNA Polymerase II (EC 2.7.7.-) ; Transcription Factors ; Transcription Factor TFIID ; TATA-Box Binding Protein ; RNA Polymerase III (EC 2.7.7.6)
Language	English
Publishing date	2022-11-10
Publishing country	England
Document type	Journal Article
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkac678
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Article ; Online: Nuclear export restricts Gdown1 to a mitotic function.

Ball, Christopher B / Parida, Mrutyunjaya / Santana, Juan F / Spector, Benjamin M / Suarez, Gustavo A / Price, David H

Nucleic acids research

2022 Volume 50, Issue 4, Page(s) 1908–1926

Abstract: Approximately half of purified mammalian RNA polymerase II (Pol II) is associated with a tightly interacting sub-stoichiometric subunit, Gdown1. Previous studies have established that Gdown1 inhibits transcription initiation through competitive ... ...

Abstract	Approximately half of purified mammalian RNA polymerase II (Pol II) is associated with a tightly interacting sub-stoichiometric subunit, Gdown1. Previous studies have established that Gdown1 inhibits transcription initiation through competitive interactions with general transcription factors and blocks the Pol II termination activity of transcription termination factor 2 (TTF2). However, the biological functions of Gdown1 remain poorly understood. Here, we utilized genetic, microscopic, and multi-omics approaches to functionally characterize Gdown1 in three human cell lines. Acute depletion of Gdown1 caused minimal direct effects on transcription. We show that Gdown1 resides predominantly in the cytoplasm of interphase cells, shuttles between the cytoplasm and nucleus, and is regulated by nuclear export. Gdown1 enters the nucleus at the onset of mitosis. Consistently, genetic ablation of Gdown1 is associated with partial de-repression of mitotic transcription, and Gdown1 KO cells present with evidence of aberrant mitoses coupled to p53 pathway activation. Evidence is presented demonstrating that Gdown1 modulates the combined functions of purified productive elongation factors PAF1C, RTF1, SPT6, DSIF and P-TEFb in vitro. Collectively, our findings support a model wherein the Pol II-regulatory function of Gdown1 occurs during mitosis and is required for genome integrity.
MeSH term(s)	Active Transport, Cell Nucleus ; Adenosine Triphosphatases/genetics ; Cell Line ; DNA-Binding Proteins/genetics ; Humans ; Mitosis ; RNA Polymerase II/metabolism ; Transcription Factors/metabolism ; Transcription, Genetic
Chemical Substances	DNA-Binding Proteins ; POLR2M protein, human ; Transcription Factors ; RNA Polymerase II (EC 2.7.7.-) ; Adenosine Triphosphatases (EC 3.6.1.-) ; TTF2 protein, human (EC 3.6.1.-)
Language	English
Publishing date	2022-02-23
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkac015
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Article: DHODH inhibition enhances the efficacy of immune checkpoint blockade by increasing cancer cell antigen presentation.

Mullen, Nicholas J / Shukla, Surendra K / Thakur, Ravi / Kollala, Sai Sundeep / Wang, Dezhen / Chaika, Nina / Santana, Juan F / Miklavcic, William R / LaBreck, Drew A / Mallareddy, Jayapal Reddy / Price, David H / Natarajan, Amarnath / Mehla, Kamiya / Sykes, David B / Hollingsworth, Michael A / Singh, Pankaj K

bioRxiv : the preprint server for biology

2024

Abstract: Pyrimidine nucleotide biosynthesis is a druggable metabolic dependency of cancer cells, and chemotherapy agents targeting pyrimidine metabolism are the backbone of treatment for many cancers. Dihydroorotate dehydrogenase (DHODH) is an essential enzyme in ...

Abstract	Pyrimidine nucleotide biosynthesis is a druggable metabolic dependency of cancer cells, and chemotherapy agents targeting pyrimidine metabolism are the backbone of treatment for many cancers. Dihydroorotate dehydrogenase (DHODH) is an essential enzyme in the de novo pyrimidine biosynthesis pathway that can be targeted by clinically approved inhibitors. However, despite robust preclinical anticancer efficacy, DHODH inhibitors have shown limited single-agent activity in phase 1 and 2 clinical trials. Therefore, novel combination therapy strategies are necessary to realize the potential of these drugs. To search for therapeutic vulnerabilities induced by DHODH inhibition, we examined gene expression changes in cancer cells treated with the potent and selective DHODH inhibitor brequinar (BQ). This revealed that BQ treatment causes upregulation of antigen presentation pathway genes and cell surface MHC class I expression. Mechanistic studies showed that this effect is 1) strictly dependent on pyrimidine nucleotide depletion, 2) independent of canonical antigen presentation pathway transcriptional regulators, and 3) mediated by RNA polymerase II elongation control by positive transcription elongation factor B (P-TEFb). Furthermore, BQ showed impressive single-agent efficacy in the immunocompetent B16F10 melanoma model, and combination treatment with BQ and dual immune checkpoint blockade (anti-CTLA-4 plus anti-PD-1) significantly prolonged mouse survival compared to either therapy alone. Our results have important implications for the clinical development of DHODH inhibitors and provide a rationale for combination therapy with BQ and immune checkpoint blockade.
Language	English
Publishing date	2024-01-13
Publishing country	United States
Document type	Preprint
DOI	10.1101/2023.04.03.535399
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Article ; Online: IDR-targeting compounds suppress HPV genome replication via disruption of phospho-BRD4 association with DNA damage response factors.

Wu, Shwu-Yuan / Lai, Hsien-Tsung / Sanjib Banerjee, N / Ma, Zonghui / Santana, Juan F / Wei, Shuguang / Liu, Xisheng / Zhang, Meirong / Zhan, Jian / Chen, Haiying / Posner, Bruce / Chen, Yadong / Price, David H / Chow, Louise T / Zhou, Jia / Chiang, Cheng-Ming

Molecular cell

2023 Volume 84, Issue 2, Page(s) 202–220.e15

Abstract: Compounds binding to the bromodomains of bromodomain and extra-terminal (BET) family proteins, particularly BRD4, are promising anticancer agents. Nevertheless, side effects and drug resistance pose significant obstacles in BET-based therapeutics ... ...

Abstract	Compounds binding to the bromodomains of bromodomain and extra-terminal (BET) family proteins, particularly BRD4, are promising anticancer agents. Nevertheless, side effects and drug resistance pose significant obstacles in BET-based therapeutics development. Using high-throughput screening of a 200,000-compound library, we identified small molecules targeting a phosphorylated intrinsically disordered region (IDR) of BRD4 that inhibit phospho-BRD4 (pBRD4)-dependent human papillomavirus (HPV) genome replication in HPV-containing keratinocytes. Proteomic profiling identified two DNA damage response factors-53BP1 and BARD1-crucial for differentiation-associated HPV genome amplification. pBRD4-mediated recruitment of 53BP1 and BARD1 to the HPV origin of replication occurs in a spatiotemporal and BRD4 long (BRD4-L) and short (BRD4-S) isoform-specific manner. This recruitment is disrupted by phospho-IDR-targeting compounds with little perturbation of the global transcriptome and BRD4 chromatin landscape. The discovery of these protein-protein interaction inhibitors (PPIi) not only demonstrates the feasibility of developing PPIi against phospho-IDRs but also uncovers antiviral agents targeting an epigenetic regulator essential for virus-host interaction and cancer development.
MeSH term(s)	Humans ; Transcription Factors/genetics ; Transcription Factors/metabolism ; Nuclear Proteins/genetics ; Nuclear Proteins/metabolism ; Human Papillomavirus Viruses ; Papillomavirus Infections/drug therapy ; Papillomavirus Infections/genetics ; Proteomics ; Cell Cycle Proteins/genetics ; Cell Cycle Proteins/metabolism ; Papillomaviridae/genetics ; Papillomaviridae/metabolism ; Viral Proteins/genetics ; Virus Replication/physiology ; DNA Repair ; Bromodomain Containing Proteins
Chemical Substances	Transcription Factors ; Nuclear Proteins ; Cell Cycle Proteins ; Viral Proteins ; BRD4 protein, human ; Bromodomain Containing Proteins
Language	English
Publishing date	2023-12-15
Publishing country	United States
Document type	Journal Article
ZDB-ID	1415236-8
ISSN	1097-4164 ; 1097-2765
ISSN (online)	1097-4164
ISSN	1097-2765
DOI	10.1016/j.molcel.2023.11.022
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Zs.A 5055: Show issues

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Article ; Online: The Dm-Myb Oncoprotein Contributes to Insulator Function and Stabilizes Repressive H3K27me3 PcG Domains.

Santana, Juan F / Parida, Mrutyunjaya / Long, Abby / Wankum, Joshua / Lilienthal, Anthony J / Nukala, Krishna M / Manak, J Robert

Cell reports

2020 Volume 30, Issue 10, Page(s) 3218–3228.e5

Abstract: Drosophila Myb (Dm-Myb) encodes a protein that plays a key role in regulation of mitotic phase genes. Here, we further refine its role in the context of a developing tissue as a potentiator of gene expression required for proper RNA polymerase II (RNA ... ...

Abstract	Drosophila Myb (Dm-Myb) encodes a protein that plays a key role in regulation of mitotic phase genes. Here, we further refine its role in the context of a developing tissue as a potentiator of gene expression required for proper RNA polymerase II (RNA Pol II) function and efficient H3K4 methylation at promoters. In contrast to its role in gene activation, Myb is also required for repression of many genes, although no specific mechanism for this role has been proposed. We now reveal a critical role for Myb in contributing to insulator function, in part by promoting binding of insulator proteins BEAF-32 and CP190 and stabilizing H3K27me3 Polycomb-group (PcG) domains. In the absence of Myb, H3K27me3 is markedly reduced throughout the genome, leading to H3K4me3 spreading and gene derepression. Finally, Myb is enriched at boundaries that demarcate chromatin environments, including chromatin loop anchors. These results reveal functions of Myb that extend beyond transcriptional regulation.
MeSH term(s)	Animals ; Cell Cycle Proteins/chemistry ; Cell Cycle Proteins/metabolism ; Drosophila Proteins/chemistry ; Drosophila Proteins/metabolism ; Drosophila melanogaster/metabolism ; Histones/metabolism ; Insulator Elements/genetics ; Lysine/metabolism ; Methylation ; Oncogene Proteins/metabolism ; Polycomb-Group Proteins/chemistry ; Protein Binding ; Protein Domains ; Protein Stability ; Proto-Oncogene Proteins c-myb/chemistry ; Proto-Oncogene Proteins c-myb/metabolism ; RNA Polymerase II/metabolism ; Transcription Initiation Site
Chemical Substances	Cell Cycle Proteins ; Drosophila Proteins ; Histones ; Myb protein, Drosophila ; Oncogene Proteins ; Polycomb-Group Proteins ; Proto-Oncogene Proteins c-myb ; RNA Polymerase II (EC 2.7.7.-) ; Lysine (K3Z4F929H6)
Language	English
Publishing date	2020-03-11
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	2649101-1
ISSN	2211-1247 ; 2211-1247
ISSN (online)	2211-1247
ISSN	2211-1247
DOI	10.1016/j.celrep.2020.02.053
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Article ; Online: Connected Elbow Exoskeleton System for Rehabilitation Training Based on Virtual Reality and Context-Aware.

de la Iglesia, Daniel H / Mendes, André Sales / González, Gabriel Villarrubia / Jiménez-Bravo, Diego M / de Paz Santana, Juan F

Sensors (Basel, Switzerland)

2020 Volume 20, Issue 3

Abstract: Traditional physiotherapy rehabilitation systems are evolving into more advanced systems based on exoskeleton systems and Virtual Reality (VR) environments that enhance and improve rehabilitation techniques and physical exercise. In addition, due to ... ...

Abstract	Traditional physiotherapy rehabilitation systems are evolving into more advanced systems based on exoskeleton systems and Virtual Reality (VR) environments that enhance and improve rehabilitation techniques and physical exercise. In addition, due to current connected systems and paradigms such as the Internet of Things (IoT) or Ambient Intelligent (AmI) systems, it is possible to design and develop advanced, effective, and low-cost medical tools that patients may have in their homes. This article presents a low-cost exoskeleton for the elbow that is connected to a Context-Aware architecture and thanks to a VR system the patient can perform rehabilitation exercises in an interactive way. The integration of virtual reality technology in rehabilitation exercises provides an intensive, repetitive and task-oriented capacity to improve patient motivation and reduce work on medical professionals. One of the system highlights is the intelligent ability to generate new exercises, monitor the exercises performed by users in search of progress or possible problems and the dynamic modification of the exercises characteristics. The platform also allows the incorporation of commercial medical sensors capable of collecting valuable information for greater accuracy in the diagnosis and evolution of patients. A case study with real patients with promising results has been carried out.
MeSH term(s)	Biomechanical Phenomena ; Elbow Joint/physiology ; Exercise Therapy ; Exoskeleton Device ; Humans ; Virtual Reality
Language	English
Publishing date	2020-02-06
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2052857-7
ISSN	1424-8220 ; 1424-8220
ISSN (online)	1424-8220
ISSN	1424-8220
DOI	10.3390/s20030858
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Article ; Online: Sirtuin 6 is required for the integrated stress response and resistance to inhibition of transcriptional cyclin-dependent kinases.

Science translational medicine

2023 Volume 15, Issue 694, Page(s) eabn9674

Abstract: Pancreatic ductal adenocarcinoma (PDAC) is classified into two key subtypes, classical and basal, with basal PDAC predicting worse survival. Using in vitro drug assays, genetic manipulation experiments, and in vivo drug studies in human patient-derived ... ...

Abstract	Pancreatic ductal adenocarcinoma (PDAC) is classified into two key subtypes, classical and basal, with basal PDAC predicting worse survival. Using in vitro drug assays, genetic manipulation experiments, and in vivo drug studies in human patient-derived xenografts (PDXs) of PDAC, we found that basal PDACs were uniquely sensitive to transcriptional inhibition by targeting cyclin-dependent kinase 7 (CDK7) and CDK9, and this sensitivity was recapitulated in the basal subtype of breast cancer. We showed in cell lines, PDXs, and publicly available patient datasets that basal PDAC was characterized by inactivation of the integrated stress response (ISR), which leads to a higher rate of global mRNA translation. Moreover, we identified the histone deacetylase sirtuin 6 (SIRT6) as a critical regulator of a constitutively active ISR. Using expression analysis, polysome sequencing, immunofluorescence, and cycloheximide chase experiments, we found that SIRT6 regulated protein stability by binding activating transcription factor 4 (ATF4) in nuclear speckles and protecting it from proteasomal degradation. In human PDAC cell lines and organoids as well as in murine PDAC genetically engineered mouse models where SIRT6 was deleted or down-regulated, we demonstrated that SIRT6 loss both defined the basal PDAC subtype and led to reduced ATF4 protein stability and a nonfunctional ISR, causing a marked vulnerability to CDK7 and CDK9 inhibitors. Thus, we have uncovered an important mechanism regulating a stress-induced transcriptional program that may be exploited with targeted therapies in particularly aggressive PDAC.
MeSH term(s)	Humans ; Mice ; Animals ; Cyclin-Dependent Kinases ; Cell Line, Tumor ; Pancreatic Neoplasms/pathology ; Carcinoma, Pancreatic Ductal/pathology ; Sirtuins/genetics ; Sirtuins/therapeutic use ; Pancreatic Neoplasms
Chemical Substances	Cyclin-Dependent Kinases (EC 2.7.11.22) ; Sirtuins (EC 3.5.1.-) ; SIRT6 protein, human (EC 3.5.1.-) ; Sirt6 protein, mouse (EC 2.4.2.31)
Language	English
Publishing date	2023-05-03
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	2518854-9
ISSN	1946-6242 ; 1946-6234
ISSN (online)	1946-6242
ISSN	1946-6234
DOI	10.1126/scitranslmed.abn9674
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Book ; Conference proceedings: 9th International Conference on Practical Applications of Computational Biology and Bioinformatics

Overbeek, Ross A / Rocha, Miguel P / Fdez.-Riverola, Florentino / Paz Santana, Juan F. de

(Advances in intelligent systems and computing, ; volume 375)

2015

Title variant	Ninth International Conference on Practical Applications of Computational Biology and Bioinformatics
Event/congress	International Conference on Practical Applications of Computational Biology & Bioinformatics (9th, 2015, SalamancaSpain)
Author's details	Ross Overbeek, Miguel P. Rocha, Florentino Fdez-Riverola, Juan F. De Paz, editors
Series title	Advances in intelligent systems and computing, ; volume 375
MeSH term(s)	Computational Biology
Language	English
Size	xiii, 148 pages :, illustrations
Document type	Book ; Conference proceedings
ISBN	9783319197753 ; 9783319197760 ; 3319197754 ; 3319197762
Database	Catalogue of the US National Library of Medicine (NLM)

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Book ; Conference proceedings: 10th International Conference on Practical Applications of Computational Biology & Bioinformatics

Mohd Saberi Mohamad / Rocha, Miguel P / Fdez.-Riverola, Florentino / Domínguez Mayo, Francisco J / Paz Santana, Juan F. de

(Advances in intelligent systems and computing, ; volume 477)

2016

Abstract: Biological and biomedical research are increasingly driven by experimental techniques that challenge our ability to analyse, process and extract meaningful knowledge from the underlying data. The impressive capabilities of next generation sequencing ... ...

Event/congress	International Conference on Practical Applications of Computational Biology & Bioinformatics (10th, 2016, SevillaSpain)
Author's details	Mohd Saberi Mohamad, Miguel P. Rocha, Florentino Fdez-Riverola, Francisco J. Domínguez Mayo, Juan F. De Paz, editors
Series title	Advances in intelligent systems and computing, ; volume 477
Abstract	Biological and biomedical research are increasingly driven by experimental techniques that challenge our ability to analyse, process and extract meaningful knowledge from the underlying data. The impressive capabilities of next generation sequencing technologies, together with novel and ever evolving distinct types of omics data technologies, have put an increasingly complex set of challenges for the growing fields of Bioinformatics and Computational Biology. The analysis of the datasets produced and their integration call for new algorithms and approaches from fields such as Databases, Statistics, Data Mining, Machine Learning, Optimization, Computer Science and Artificial Intelligence. Clearly, Biology is more and more a science of information requiring tools from the computational sciences. In the last few years, we have seen the surge of a new generation of interdisciplinary scientists that have a strong background in the biological and computational sciences. In this context, the interaction of researchers from different scientific fields is, more than ever, of foremost importance boosting the research efforts in the field and contributing to the education of a new generation of Bioinformatics scientists. PACBB'16 hopes to contribute to this effort promoting this fruitful interaction. PACBB'16 technical program included 21 papers spanning many different sub-fields in Bioinformatics and Computational Biology. Therefore, the conference will certainly promote the interaction of scientists from diverse research groups and with a distinct background (computer scientists, mathematicians, biologists). The scientific content will certainly be challenging and will promote the improvement of the work being developed by each of the participants.
MeSH term(s)	Computational Biology ; Data Mining
Language	English
Dates of publication	2016-2016
Size	xv, 224 pages :, illustrations (some color) ;, 24 cm.
Document type	Book ; Conference proceedings
ISBN	9783319401256 ; 9783319401263 ; 3319401254 ; 3319401262
Database	Catalogue of the US National Library of Medicine (NLM)

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