Biology subjects

Duttke, S. H.

Publications and source records attributed to Duttke, S. H..

8 recordsLinked to original sources

bioRxiv2026

EPIC: An open community challenge for sequence-based prediction of transcription initiation in five non-model metazoans

Predicting gene expression from DNA sequence is a central problem with critical biological and clinical implications. Recent sequence-to-function models are reported to achieve improved performance, yet it remains unclear how much of what they learn reflects genuine principles of eukaryotic transcription initiation and to what extent they are able to generalize beyond humans and other primary model species. A fair and blind benchmark has likewise been missing. Here we introduce EPIC, the Eukaryotic Promoter and transcription Initiation prediction Challenge. Teams receive strand-specific, single-nucleotide-resolution initiation profiles for 80-95% of the genome and are asked to predict the remainder from DNA sequence alone. To level the field, the challenge relies on understudied animals spanning three phyla: octopus, oyster, milkweed bug, Indian meal moth, and shark. EPIC is open to everyone and closes on December 31, 2026. All teams clearing the dinucleotide precision baseline are invited to join the consortium authorship of the post-challenge publication, and winners are invited for personal authorship.

bioinformatics↗

bioRxiv2024

Efficient small fragment sequencing of human, cow, and bison miRNA, small RNA or csRNA-seq libraries using AVITI

Next-Generation Sequencing (NGS) catalyzed breakthroughs across various scientific domains. Illuminas sequencing by synthesis method has long been essential for NGS but emerging technologies like Element Biosciences sequencing by avidity (AVITI) represent a novel approach. It has been reported that AVITI offers improved signal-to-noise ratios and cost reductions. However, the method relies on rolling circle amplification which can be impacted by polymer size, raising questions about its efficacy sequencing small RNAs (sRNA) molecules including microRNAs (miRNAs), piwi-interacting RNAs (piRNAs), and others that are crucial regulators of gene expression and involved in various biological processes. In addition, capturing capped small RNAs (csRNA-seq) has emerged as a powerful method to map active or "nascent" RNA polymerase II transcription initiation in tissues and clinical samples. Here, we report a new protocol for seamlessly sequencing short DNA fragments on the AVITI and demonstrate that AVITI and Illumina sequencing technologies equivalently capture human, cattle (Bos taurus) and the bison (Bison bison) sRNA or csRNA sequencing libraries, augmenting the confidence in both approaches. Additionally, analysis of generated nascent transcription start sites (TSSs) data for cattle and bison revealed inaccuracies in their current genome annotations and highlighted the possibility and need to translate small RNA sequencing methodologies to livestock. Our accelerated and optimized protocol therefore bridges the advantages of AVITI sequencing and critical methods that rely on sequencing short DNA fragments.

genomics↗

bioRxiv2023

Enhancers associated with unstable RNAs are rare in plants

Unstable transcripts have emerged as markers of active enhancers in vertebrates and shown to be involved in many cellular processes and medical disorders. However, their prevalence and role in plants is largely unexplored. Here, we comprehensively captured all actively initiating ("nascent") transcripts across diverse crops and other plants using capped small (cs)RNA-seq. We discovered that unstable transcripts are rare, unlike in vertebrates, and often originate from promoters. Additionally, many "distal" elements in plants initiate tissue-specific stable transcripts and are likely bone fide promoters of yet-unannotated genes or non-coding RNAs, cautioning against using genome annotations to infer "enhancers" or transcript stability. To investigate enhancer function, we integrated STARR-seq data. We found that annotated promoters, and other regions that initiate stable transcripts rather than unstable transcripts, function as stronger enhancers in plants. Our findings underscore the blurred line between promoters and enhancers and suggest that cis-regulatory elements encompass diverse structures and mechanisms in eukaryotes.

genomics↗

bioRxiv2023

A single DPE core promoter motif contributes to in vivo transcriptional regulation and affects cardiac function

Transcription is initiated at the core promoter, which confers specific functions depending on the unique combination of core promoter elements. The downstream core promoter element (DPE) is found in many genes related to heart and mesodermal development. However, the function of these core promoter elements has thus far been studied primarily in isolated, in vitro or reporter gene settings. tinman (tin) encodes a key transcription factor that regulates the formation of the dorsal musculature and heart. Pioneering a novel approach utilizing both CRISPR and nascent transcriptomics, we show that a substitution mutation of the functional tin DPE motif within the natural context of the core promoter results in a massive perturbation of Tinmans regulatory network orchestrating dorsal musculature and heart formation. Mutation of endogenous tin DPE reduced the expression of tin and distinct target genes, resulting in significantly reduced viability and an overall decrease in adult heart function. We demonstrate the feasibility and importance of characterizing DNA sequence elements in vivo in their natural context, and accentuate the critical impact a single DPE motif has during Drosophila embryogenesis and functional heart formation.

developmental biology↗

bioRxiv2021

Profiling Transcription Initiation in Peripheral Leukocytes Reveals Severity-Associated Cis-Regulatory Elements in Critical COVID-19

The contribution of transcription factors (TFs) and gene regulatory programs in the immune response to COVID-19 and their relationship to disease outcome is not fully understood. Analysis of genome-wide changes in transcription at both promoter-proximal and distal cis-regulatory DNA elements, collectively termed the active cistrome, offers an unbiased assessment of TF activity identifying key pathways regulated in homeostasis or disease. Here, we profiled the active cistrome from peripheral leukocytes of critically ill COVID-19 patients to identify major regulatory programs and their dynamics during SARS-CoV-2 associated acute respiratory distress syndrome (ARDS). We identified TF motifs that track the severity of COVID- 19 lung injury, disease resolution, and outcome. We used unbiased clustering to reveal distinct cistrome subsets delineating the regulation of pathways, cell types, and the combinatorial activity of TFs. We found critical roles for regulatory networks driven by stimulus and lineage determining TFs, showing that STAT and E2F/MYB regulatory programs targeting myeloid cells are activated in patients with poor disease outcomes and associated with single nucleotide genetic variants implicated in COVID-19 susceptibility. Integration with single-cell RNA-seq found that STAT and E2F/MYB activation converged in specific neutrophils subset found in patients with severe disease. Collectively we demonstrate that cistrome analysis facilitates insight into disease mechanisms and provides an unbiased approach to evaluate global changes in transcription factor activity and stratify patient disease severity.

genomics↗

bioRxiv2021

Transcriptional regulatory features associated with Coccidioides immitis phase transition

Coccidioidomycosis (Valley Fever) is an emerging endemic fungal infection with a rising incidence and an expanding geographic range. It is caused by Coccidiodes, which are thermally dimorphic fungi that grow as mycelia in soil but transition in the lung to form pathogenic spherules. The regulatory mechanisms underlying this transition are not understood. Exploiting capped small (cs)RNA-seq, which identifies actively initiated stable and unstable transcripts and thereby detects acute changes in gene regulation with remarkable sensitivity, here we report the changes in architectural organization and key sequence features underlying phase transition of this highly pathogenic fungus. Spherule transition was accompanied by large-scale transcriptional reprogramming, functional changes in transcript isoforms, and a massive increase in promoter-distal transcription of ncRNAs. Analysis of spherule-activated regulatory elements revealed a motif predicted to recruit a WOPR family transcription factor, which are known regulators of virulence in other fungi. We identify CIMG_02671 as a C. immitis WOPR homologue and show that it activates transcription in a WOPR motif-dependent manner, suggesting it is an important regulator of pathogenic phase transition. Collectively, this also highlights csRNA-seq as a powerful means to identify transcriptional mechanisms that control pathogenesis.

immunology↗

bioRxiv2020

A Chinese hamster transcription start site atlas that enables targeted editing of CHO cells

Chinese hamster ovary (CHO) cells, with their human-compatible glycosylation and high protein titers, are the most widely used cells for producing biopharmaceuticals. Engineering gene expression in CHO is key to improving drug quality and affordability. However, engineering gene expression or activating silent genes requires accurate annotation of the underlying regulatory elements and transcription start sites (TSSs). Unfortunately, most TSSs in the Chinese hamster genome were computationally predicted and are frequently inaccurate. Here, we revised TSS annotations for 15,308 Chinese hamster genes and 4,478 non-coding RNAs based on experimental data from CHO-K1 cells and 10 hamster tissues. The experimental realignment and discovery of TSSs now expose previously hidden motifs, such as the TATA box. We further demonstrate, by targeting the glycosyltransferase gene Mgat3, how accurate annotations readily facilitate activating silent genes by CRISPRa to obtain more human-like glycosylation. Together, we envision our annotation and data will provide a rich resource for the CHO community, improve genome engineering efforts and aid comparative and evolutionary studies.

genomics↗

bioRxiv2018

Diverse motif ensembles specify non-redundant DNA binding activities of AP-1 family members in macrophages

Mechanisms by which members of the AP-1 family of transcription factors play both redundant and non-redundant biological roles despite recognizing the same DNA sequence remain poorly understood. To address this question, we investigated the molecular functions and genome-wide DNA binding patterns of AP-1 family members in macrophages. ChIP-sequencing showed overlapping and distinct binding profiles for each factor that were remodeled following TLR4 ligation. Development of a machine learning approach that jointly weighs hundreds of DNA recognition elements yielded dozens of motifs predicted to drive factor-specific binding profiles. Machine learning-based predictions were confirmed by analysis of the effects of mutations in genetically diverse mice and by loss of function experiments. These findings provide evidence that non-redundant genomic locations of different AP-1 family members in macrophages largely result from collaborative interactions with diverse, locus-specific ensembles of transcription factors and suggest a general mechanism for encoding functional specificities of their common recognition motif.

genomics↗