Publications by authors named "Riccardo Dainese"

7 Publications

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A parallelized, automated platform enabling individual or sequential ChIP of histone marks and transcription factors.

Proc Natl Acad Sci U S A 2020 06 27;117(24):13828-13838. Epub 2020 May 27.

School of Life Sciences, Institute of Bioengineering, École Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland;

Despite its popularity, chromatin immunoprecipitation followed by sequencing (ChIP-seq) remains a tedious (>2 d), manually intensive, low-sensitivity and low-throughput approach. Here, we combine principles of microengineering, surface chemistry, and molecular biology to address the major limitations of standard ChIP-seq. The resulting technology, FloChIP, automates and miniaturizes ChIP in a beadless fashion while facilitating the downstream library preparation process through on-chip chromatin tagmentation. FloChIP is fast (<2 h), has a wide dynamic range (from 10 to 500 cells), is scalable and parallelized, and supports antibody- or sample-multiplexed ChIP on both histone marks and transcription factors. In addition, FloChIP's interconnected design allows for straightforward chromatin reimmunoprecipitation, which allows this technology to also act as a microfluidic sequential ChIP-seq system. Finally, we ran FloChIP for the transcription factor MEF2A in 32 distinct human lymphoblastoid cell lines, providing insights into the main factors driving collaborative DNA binding of MEF2A and into its role in B cell-specific gene regulation. Together, our results validate FloChIP as a flexible and reproducible automated solution for individual or sequential ChIP-seq.
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http://dx.doi.org/10.1073/pnas.1913261117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7306797PMC
June 2020

cis-regulatory variation modulates susceptibility to enteric infection in the Drosophila genetic reference panel.

Genome Biol 2020 01 17;21(1). Epub 2020 Jan 17.

Laboratory of Systems Biology and Genetics, Institute of Bioengineering, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland.

Background: Resistance to enteric pathogens is a complex trait at the crossroads of multiple biological processes. We have previously shown in the Drosophila Genetic Reference Panel (DGRP) that resistance to infection is highly heritable, but our understanding of how the effects of genetic variants affect different molecular mechanisms to determine gut immunocompetence is still limited.

Results: To address this, we perform a systems genetics analysis of the gut transcriptomes from 38 DGRP lines that were orally infected with Pseudomonas entomophila. We identify a large number of condition-specific, expression quantitative trait loci (local-eQTLs) with infection-specific ones located in regions enriched for FOX transcription factor motifs. By assessing the allelic imbalance in the transcriptomes of 19 F1 hybrid lines from a large round robin design, we independently attribute a robust cis-regulatory effect to only 10% of these detected local-eQTLs. However, additional analyses indicate that many local-eQTLs may act in trans instead. Comparison of the transcriptomes of DGRP lines that were either susceptible or resistant to Pseudomonas entomophila infection reveals nutcracker as the only differentially expressed gene. Interestingly, we find that nutcracker is linked to infection-specific eQTLs that correlate with its expression level and to enteric infection susceptibility. Further regulatory analysis reveals one particular eQTL that significantly decreases the binding affinity for the repressor Broad, driving differential allele-specific nutcracker expression.

Conclusions: Our collective findings point to a large number of infection-specific cis- and trans-acting eQTLs in the DGRP, including one common non-coding variant that lowers enteric infection susceptibility.
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http://dx.doi.org/10.1186/s13059-019-1912-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6966807PMC
January 2020

ZFP30 promotes adipogenesis through the KAP1-mediated activation of a retrotransposon-derived Pparg2 enhancer.

Nat Commun 2019 04 18;10(1):1809. Epub 2019 Apr 18.

Laboratory of Systems Biology and Genetics, Institute of Bioengineering, School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), CH-1015, Lausanne, Switzerland.

Krüppel-associated box zinc finger proteins (KZFPs) constitute the largest family of mammalian transcription factors, but most remain completely uncharacterized. While initially proposed to primarily repress transposable elements, recent reports have revealed that KFZPs contribute to a wide variety of other biological processes. Using murine and human in vitro and in vivo models, we demonstrate here that one poorly studied KZFP, ZFP30, promotes adipogenesis by directly targeting and activating a retrotransposon-derived Pparg2 enhancer. Through mechanistic studies, we further show that ZFP30 recruits the co-regulator KRAB-associated protein 1 (KAP1), which, surprisingly, acts as a ZFP30 co-activator in this adipogenic context. Our findings provide an understanding of both adipogenic and KZFP-KAP1 complex-mediated gene regulation, showing that the KZFP-KAP1 axis can also function in a non-repressive manner.
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http://dx.doi.org/10.1038/s41467-019-09803-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6472429PMC
April 2019

A Comprehensive Drosophila melanogaster Transcription Factor Interactome.

Cell Rep 2019 04;27(3):955-970.e7

Department of Medicine, Division of Genetics, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA; Systems Biology Graduate Program, Harvard University, Cambridge, MA 02138, USA; Bioinformatics and Integrative Genomics Ph.D. Program, Harvard University, Cambridge, MA 02138, USA; Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, MA 02115, USA. Electronic address:

Combinatorial interactions among transcription factors (TFs) play essential roles in generating gene expression specificity and diversity in metazoans. Using yeast 2-hybrid (Y2H) assays on nearly all sequence-specific Drosophila TFs, we identified 1,983 protein-protein interactions (PPIs), more than doubling the number of currently known PPIs among Drosophila TFs. For quality assessment, we validated a subset of our interactions using MITOMI and bimolecular fluorescence complementation assays. We combined our interactome with prior PPI data to generate an integrated Drosophila TF-TF binary interaction network. Our analysis of ChIP-seq data, integrating PPI and gene expression information, uncovered different modes by which interacting TFs are recruited to DNA. We further demonstrate the utility of our Drosophila interactome in shedding light on human TF-TF interactions. This study reveals how TFs interact to bind regulatory elements in vivo and serves as a resource of Drosophila TF-TF binary PPIs for understanding tissue-specific gene regulation.
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http://dx.doi.org/10.1016/j.celrep.2019.03.071DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6485956PMC
April 2019

Adaptive damage retention mechanism enables healthier yeast population.

J Theor Biol 2019 07 11;473:52-66. Epub 2019 Apr 11.

Department of Mathematical Sciences, Chalmers University of Technology and University of Gothenburg, Chalmers tvärgata 3, SE-41296 Gothenburg, Sweden. Electronic address:

During cytokinesis in budding yeast (Saccharomyces cerevisiae) damaged proteins are distributed asymmetrically between the daughter and the mother cell. Retention of damaged proteins is a crucial mechanism ensuring a healthy daughter cell with full replicative potential and an ageing mother cell. However, the protein quality control (PQC) system is tuned for optimal reproduction success which suggests optimal health and size of the population, rather than long-term survival of the mother cell. Modelling retention of damage as an adaptable mechanism, we propose two damage retention strategies to find an optimal way of decreasing damage retention efficiency to maximize population size and minimize the damage in the individual yeast cell. A pedigree model is used to investigate the impact of small variations in the strategies over the whole population. These impacts are based on the altruistic effects of damage retention mechanism and are measured by a cost function whose minimum value provides the optimal health and size of the population. We showed that fluctuations in the cost function allow yeast cell to continuously vary its strategy, suggesting that optimal reproduction success is a local minimum of the cost function. Our results suggest that a rapid decrease in the efficiency of damage retention, at the time when the mother cell is almost exhausted, produces fewer daughters with high levels of damaged proteins. In addition, retaining more damage during the early divisions increases the number of healthy daughters in the population.
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http://dx.doi.org/10.1016/j.jtbi.2019.04.005DOI Listing
July 2019

Simplified Drop-seq workflow with minimized bead loss using a bead capture and processing microfluidic chip.

Lab Chip 2019 04;19(9):1610-1620

Laboratory of Systems Biology and Genetics, Institute of Bioengineering, School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland. and Swiss Institute of Bioinformatics, Lausanne, Switzerland.

Single-cell RNA-sequencing (scRNA-seq) has revolutionized biomedical research by enabling the in-depth analysis of cell-to-cell heterogeneity of tissues with unprecedented resolution. One of the catalyzing technologies is single cell droplet microfluidics, which has massively increased the overall cell throughput, routinely allowing the analysis of thousands of cells per experiment at a relatively low cost. Among several existing droplet-based approaches, the Drop-seq platform has emerged as one of the most widely used systems. Yet, this has surprisingly not incentivized major refinements of the method, thus restricting any lab implementation to the original Drop-seq setup, which is known to suffer from up to 80% bead loss during the process. In this study, we present a systematic re-engineering and optimization of Drop-seq: first, we re-designed the original dropleting device to be compatible with both air-pressure systems and syringe pumps, thus increasing the overall flexibility of the platform. Second, we devised an accompanying chip for post-encapsulation bead processing, which simplifies and massively increases Drop-seq's cell processing efficiency. Taken together, the presented optimization efforts result in a more flexible and efficient Drop-seq version.
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http://dx.doi.org/10.1039/c9lc00014cDOI Listing
April 2019

SMiLE-seq identifies binding motifs of single and dimeric transcription factors.

Nat Methods 2017 03 16;14(3):316-322. Epub 2017 Jan 16.

Institute of Bioengineering, École Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland.

Resolving the DNA-binding specificities of transcription factors (TFs) is of critical value for understanding gene regulation. Here, we present a novel, semiautomated protein-DNA interaction characterization technology, selective microfluidics-based ligand enrichment followed by sequencing (SMiLE-seq). SMiLE-seq is neither limited by DNA bait length nor biased toward strong affinity binders; it probes the DNA-binding properties of TFs over a wide affinity range in a fast and cost-effective fashion. We validated SMiLE-seq by analyzing 58 full-length human, mouse, and Drosophila TFs from distinct structural classes. All tested TFs yielded DNA-binding models with predictive power comparable to or greater than that of other in vitro assays. De novo motif discovery on all JUN-FOS heterodimers and several nuclear receptor-TF complexes provided novel insights into partner-specific heterodimer DNA-binding preferences. We also successfully analyzed the DNA-binding properties of uncharacterized human C2H2 zinc-finger proteins and validated several using ChIP-exo.
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http://dx.doi.org/10.1038/nmeth.4143DOI Listing
March 2017