Global landscape of mouse and human cytokine transcriptional regulation

Pro, Sebastian Carrasco; Imedio, Alvaro Dafonte; Santoso, Clarissa Stephanie; Gan, Kok Ann; Sewell, Jared Allan; Martinez, Mélissa; Sereda, Rebecca; Mehta, Suresh L.; Bass, Juan I. Fuxman

doi:10.1101/316612

Cited by 3 publications

(2 citation statements)

References 58 publications

(73 reference statements)

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“…IL32 and PROK2 were two of the most differentially expressed cytokine genes, as well as being present multiple times in the over‐represented categories of “Migration of cells,” “Inflammation of organ,” and “Mobilization of Ca 2+ ” ( Figure , Table ). Prior to initiation of these functional experiments, we performed a strict Bonferroni multiple testing correction ( P < 0.0002) for all cytokine genes in the human genome 30 . Of the 11 cytokine genes defined as significant by FDR ( Table ), 10 remained significant following Bonferroni correction, including IL32 and PROK2 .…”

Section: Resultsmentioning

confidence: 99%

Differences in the Platelet mRNA Landscape Portend Racial Disparities in Platelet Function and Suggest Novel Therapeutic Targets

Garofano

Park

Alarcón

et al. 2021

Clin Pharma and Therapeutics

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The African American (AA) population displays a 1.6 to 3‐fold higher incidence of thrombosis and stroke mortality compared with European Americans (EAs). Current antiplatelet therapies target the ADP‐mediated signaling pathway, which displays significant pharmacogenetic variation for platelet reactivity. The focus of this study was to define underlying population differences in platelet function in an effort to identify novel molecular targets for future antiplatelet therapy. We performed deep coverage RNA‐Seq to compare gene expression levels in platelets derived from a cohort of healthy volunteers defined by ancestry determination. We identified > 13,000 expressed platelet genes of which 480 were significantly differentially expressed genes (DEGs) between AAs and EAs. DEGs encoding proteins known or predicted to modulate platelet aggregation, morphology, or platelet count were upregulated in AA platelets. Numerous G‐protein coupled receptors, ion channels, and pro‐inflammatory cytokines not previously associated with platelet function were likewise differentially expressed. Many of the signaling proteins represent potential pharmacologic targets of intervention. Notably, we confirmed the differential expression of cytokines IL32 and PROK2 in an independent cohort by quantitative real‐time polymerase chain reaction, and provide functional validation of the opposing actions of these two cytokines on collagen‐induced AA platelet aggregation. Using Genotype‐Tissue Expression whole blood data, we identified 516 expression quantitative trait locuses with Fst values > 0.25, suggesting that population‐differentiated alleles may contribute to differences in gene expression. This study identifies gene expression differences at the population level that may affect platelet function and serve as potential biomarkers to identify cardiovascular disease risk. Additionally, our analysis uncovers candidate novel druggable targets for future antiplatelet therapies.

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Section: Resultsmentioning

confidence: 99%

Differences in the Platelet mRNA Landscape Portend Racial Disparities in Platelet Function and Suggest Novel Therapeutic Targets

Garofano

Park

Alarcón

et al. 2021

Clin Pharma and Therapeutics

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show abstract

“…Limitations to eY1H assays include that LTR chromatization may not be recapitulated in yeast, prey transcription factors may not have proper posttranslational modifications, and the screen does not capture heteromultimeric complexes. However, our previous studies examining transcriptional elements by eY1H had 30 to 60% validation rates for identified factors, comparable to, or better than, other methods, including ChIP-seq and motif prediction (26,(45)(46)(47).…”

Section: Discussionmentioning

confidence: 53%

A functional screen identifies transcriptional networks that regulate HIV-1 and HIV-2

Pedro

Agosto

Sewell

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

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The molecular networks involved in the regulation of HIV replication, transcription, and latency remain incompletely defined. To expand our understanding of these networks, we performed an unbiased high-throughput yeast one-hybrid screen, which identified 42 human transcription factors and 85 total protein–DNA interactions with HIV-1 and HIV-2 long terminal repeats. We investigated a subset of these transcription factors for transcriptional activity in cell-based models of infection. KLF2 and KLF3 repressed HIV-1 and HIV-2 transcription in CD4+ T cells, whereas PLAGL1 activated transcription of HIV-2 through direct protein–DNA interactions. Using computational modeling with interacting proteins, we leveraged the results from our screen to identify putative pathways that define intrinsic transcriptional networks. Overall, we used a high-throughput functional screen, computational modeling, and biochemical assays to identify and confirm several candidate transcription factors and biochemical processes that influence HIV-1 and HIV-2 transcription and latency.

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Comprehensive mapping of the human cytokine gene regulatory network

Santoso¹,

Lal

et al. 2020

Preprint

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Proper cytokine gene expression is essential in development, homeostasis, and immune responses. Studies on the transcriptional control of cytokine genes have mostly focused on highly researched transcription factors (TFs) and cytokines, resulting in an incomplete portrait of cytokine gene regulation. Here, we use enhanced yeast one-hybrid (eY1H) assays to derive a comprehensive network comprising 1,380 interactions between 265TFs and 108 cytokine gene promoters, greatly expanding the known repertoire of TFcytokine gene interactions. We found an enrichment of nuclear receptors and confirmed their role in cytokine regulation in primary macrophages. Additionally, we used the eY1Hderived network as a framework to identify pairs of TFs that synergistically modulate cytokine gene expression, and to identify novel TF-cytokine regulatory axes in immune diseases and immune cell lineage development. Overall, the eY1H data provides a rich resource to study cytokine regulation in a variety of physiological and disease contexts. 8 High Fidelity (ThermoFisher) from a pool of human genomic DNA (Clonetech). PCR products were Gateway cloned into the pDONR P1-P4 vector and entry clones were confirmed by Sanger sequencing. Then DNA-baits were Gateway cloned upstream of two reporter genes (HIS3 and LacZ) and both reporter constructs were integrated into the yeast genome to generate chromatinized DNA-bait strains (Deplancke et al., 2006a;Deplancke et al., 2006b). Yeast DNA-bait strains were confirmed by Sanger sequencing.DNA-bait strains were mated with an array of 1,086 TF-prey yeast strains expressing human TFs fused to the yeast Gal4 activation domain (AD) (Fuxman Bass et al., 2015).Matings were performed using a Singer Rotor robotic platform that manipulates yeast strains in a 1,536-colony format. An interaction was detected when a TF-prey binds the DNA-bait and the AD moiety activates reporter gene expression, allowing the mated yeast to grow on media lacking histidine, overcome the addition of 3-amino-triazole (3AT), a competitive inhibitor of the His3p enzyme, and convert the colorless X-gal into a blue compound. Each interaction was tested in quadruplicate and interactions were considered positive if at least two of the four mated colonies tested positively (>90% of detected interactions tested positively for all four colonies).Images of mated colonies on plates lacking histidine and containing 3AT and Xgal were processed using the Mybrid web-tool to detect positive interactions (Reece-Hoyes et al., 2013). Positive interactions detected by Mybrid were then manually checked and curated. False positives detected by Mybrid, which typically occur on plates with uneven background, were removed. False negatives missed by Mybrid, which typically occur when baits exhibit high background reporter gene expression or when interactions occur next to very strong positives, were included. A total of high-quality 1,380 PDIs 9 between 265 TFs and 108 cytokine promoters were included in the final dataset (Table S3). The eY1H interactions...

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Global landscape of mouse and human cytokine transcriptional regulation

Cited by 3 publications

References 58 publications

Differences in the Platelet mRNA Landscape Portend Racial Disparities in Platelet Function and Suggest Novel Therapeutic Targets

Differences in the Platelet mRNA Landscape Portend Racial Disparities in Platelet Function and Suggest Novel Therapeutic Targets

A functional screen identifies transcriptional networks that regulate HIV-1 and HIV-2

Comprehensive mapping of the human cytokine gene regulatory network

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