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Identifying the molecular origins by which new morphological structures evolve is one of the long-standing problems in evolutionary biology. To date, vanishingly few examples provide a compelling account of how new morphologies were initially formed, thereby limiting our understanding of how diverse forms of life derived their complex features. Here, we provide evidence that the large projections on the Drosophila eugracilis phallus that are implicated in sexual conflict have evolved through the partial co-option of the trichome genetic network. These unicellular apical projections on the phallus postgonal sheath are reminiscent of trichomes that cover the Drosophila body but are up to 20-fold larger in size. During their development, they express the transcription factor Shavenbaby, the master regulator of the trichome network. Consistent with the co-option of the Shavenbaby network during the evolution of the D. eugracilis projections, somatic mosaic CRISPR-Cas9 mutagenesis shows that shavenbaby is necessary for their proper length. Moreover, misexpression of Shavenbaby in the sheath of D. melanogaster, a naive species that lacks these projections, is sufficient to induce small trichomes. These induced projections rely on a genetic network that is shared to a large extent with the D. eugracilis projections, indicating its partial co-option but also some genetic rewiring. Thus, by leveraging a genetically tractable evolutionary novelty, our work shows that the trichome-forming network is flexible enough that it can be partially co-opted in a new context and subsequently refined to produce unique apical projections that are barely recognizable compared with their simpler ancestral beginnings., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Congenital heart disease often arises from perturbations of transcription factors (TFs) that guide cardiac development. ISLET1 (ISL1) is a TF that influences early cardiac cell fate, as well as differentiation of other cell types including motor neuron progenitors (MNPs) and pancreatic islet cells. While lineage specificity of ISL1 function is likely achieved through combinatorial interactions, its essential cardiac interacting partners are unknown. By assaying ISL1 genomic occupancy in human induced pluripotent stem cell-derived cardiac progenitors (CPs) or MNPs and leveraging the deep learning approach BPNet, we identified motifs of other TFs that predicted ISL1 occupancy in each lineage, with NKX2.5 and GATA motifs being most closely associated to ISL1 in CPs. Experimentally, nearly two-thirds of ISL1-bound loci were co-occupied by NKX2.5 and/or GATA4. Removal of NKX2.5 from CPs led to widespread ISL1 redistribution, and overexpression of NKX2.5 in MNPs led to ISL1 occupancy of CP-specific loci. These results reveal how ISL1 guides lineage choices through a combinatorial code that dictates genomic occupancy and transcription., Competing Interests: Declaration of interests D.S. is a co-founder and member of the board of directors of Tenaya Therapeutics and has equity in Tenaya Therapeutics. K.N.I. is an employee and shareholder of Tenaya Therapeutics. N.J.K. has received research support from Vir Biotechnology, F. Hoffmann-La Roche, and Rezo Therapeutics. N.J.K. has financially compensated consulting agreements with Maze Therapeutics, Interline Therapeutics, Rezo Therapeutics, and GEn1E Lifesciences, Inc. He is on the Board of Directors of Rezo Therapeutics and is a shareholder in Tenaya Therapeutics, Maze Therapeutics, Rezo Therapeutics, and Interline Therapeutics., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Chromatin accessibility is integral to the process by which transcription factors (TFs) read out cis-regulatory DNA sequences, but it is difficult to differentiate between TFs that drive accessibility and those that do not. Deep learning models that learn complex sequence rules provide an unprecedented opportunity to dissect this problem. Using zygotic genome activation in Drosophila as a model, we analyzed high-resolution TF binding and chromatin accessibility data with interpretable deep learning and performed genetic validation experiments. We identify a hierarchical relationship between the pioneer TF Zelda and the TFs involved in axis patterning. Zelda consistently pioneers chromatin accessibility proportional to motif affinity, whereas patterning TFs augment chromatin accessibility in sequence contexts where they mediate enhancer activation. We conclude that chromatin accessibility occurs in two tiers: one through pioneering, which makes enhancers accessible but not necessarily active, and the second when the correct combination of TFs leads to enhancer activation., Competing Interests: Declaration of interests J.Z. owns a patent on ChIP-nexus (no. 10287628). A.K. is on the scientific advisory board of PatchBio, SerImmune, AINovo, TensorBio and OpenTargets, was a consultant with Illumina, and owns shares in Illumina, Deep Genomics, Immunai, and Freenome Inc. All other authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

What new questions can we ask about transcriptional regulation given recent developments in large-scale approaches?, Competing Interests: Declaration of interests J.Z. is an investigator at the Stowers Institute for Medical Research and also a professor (affiliate track) at the Department of Pathology and Laboratory Medicine at the University of Kansas Medical Center. H.J.B. is a co-founder and shareholder of Metric Biotechnologies, Inc. Columbia University has filed a patent on a technology tangentially related, on which H.J.B. is one of the inventors, to the topic of the Voices piece that H.J.B. contributed., (Copyright © 2023. Published by Elsevier Inc.)

Mouse embryonic stem cells (ESCs) cultured in defined medium resemble the pre-implantation epiblast in the ground state, with full developmental capacity including the germline. β-Catenin is required to maintain ground state pluripotency in mouse ESCs, but its exact role is controversial. Here, we reveal a Tcf3-independent role of β-catenin in restraining germline and somatic lineage differentiation genes. We show that β-catenin binds target genes with E2F6 and forms a complex with E2F6 and HMGA2 or E2F6 and HP1γ. Our data indicate that these complexes help β-catenin restrain and fine-tune germ cell and neural developmental potential. Overall, our data reveal a previously unappreciated role of β-catenin in preserving lineage differentiation integrity in ground state ESCs., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

FACT (facilitates chromatin transcription) is an evolutionarily conserved histone chaperone that was initially identified as an activity capable of promoting RNA polymerase II (Pol II) transcription through nucleosomes in vitro. In this report, we describe a global analysis of FACT function in Pol II transcription in Drosophila. We present evidence that loss of FACT has a dramatic impact on Pol II elongation-coupled processes including histone H3 lysine 4 (H3K4) and H3K36 methylation, consistent with a role for FACT in coordinating histone modification and chromatin architecture during Pol II transcription. Importantly, we identify a role for FACT in the maintenance of promoter-proximal Pol II pausing, a key step in transcription activation in higher eukaryotes. These findings bring to light a broader role for FACT in the regulation of Pol II transcription., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

Hyperactivating mutations in Ras signaling are hallmarks of carcinomas. Ras signaling mediates cell fate decisions as well as proliferation during development. It is not known what dictates whether Ras signaling drives differentiation versus proliferation. Here we show that the Hippo pathway is critical for this decision. Loss of Hippo switches Ras activation from promoting cellular differentiation to aggressive cellular proliferation. Transcriptome analysis combined with genetic tests show that this excessive proliferation depends on the synergistic induction of Ras target genes. Using ChIP-nexus, we find that Hippo signaling keeps Ras targets in check by directly regulating the expression of two key downstream transcription factors of Ras signaling: the ETS-domain transcription factor Pointed and the repressor Capicua. Our results highlight how independent signaling pathways can impinge on each other at the level of transcription factors, thereby providing a safety mechanism to keep proliferation in check under normal developmental conditions., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Paused RNA polymerase (Pol II) is a pervasive feature of Drosophila embryos and mammalian stem cells, but its role in development is uncertain. Here, we demonstrate that a spectrum of paused Pol II determines the "time to synchrony"-the time required to achieve coordinated gene expression across the cells of a tissue. To determine whether synchronous patterns of gene activation are significant in development, we manipulated the timing of snail expression, which controls the coordinated invagination of ∼1,000 mesoderm cells during gastrulation. Replacement of the strongly paused snail promoter with moderately paused or nonpaused promoters causes stochastic activation of snail expression and increased variability of mesoderm invagination. Computational modeling of the dorsal-ventral patterning network recapitulates these variable and bistable gastrulation profiles and emphasizes the importance of timing of gene activation in development. We conclude that paused Pol II and transcriptional synchrony are essential for coordinating cell behavior during morphogenesis., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Poised RNA polymerase II (Pol II) is predominantly found at developmental control genes and is thought to allow their rapid and synchronous induction in response to extracellular signals. How the recruitment of poised RNA Pol II is regulated during development is not known. By isolating muscle tissue from Drosophila embryos at five stages of differentiation, we show that the recruitment of poised Pol II occurs at many genes de novo and this makes them permissive for future gene expression. A comparison with other tissues shows that these changes are stage specific and not tissue specific. In contrast, Polycomb group repression is tissue specific, and in combination with Pol II (the balanced state) marks genes with highly dynamic expression. This suggests that poised Pol II is temporally regulated and is held in check in a tissue-specific fashion. We compare our data with findings in mammalian embryonic stem cells and discuss a framework for predicting developmental programs on the basis of the chromatin state., (Copyright © 2012 The Authors. Published by Elsevier Inc. All rights reserved.)

Eukaryotic genomes are packaged into nucleosomes whose position and chemical modification state can profoundly influence regulation of gene expression. We profiled nucleosome modifications across the yeast genome using chromatin immunoprecipitation coupled with DNA microarrays to produce high-resolution genome-wide maps of histone acetylation and methylation. These maps take into account changes in nucleosome occupancy at actively transcribed genes and, in doing so, revise previous assessments of the modifications associated with gene expression. Both acetylation and methylation of histones are associated with transcriptional activity, but the former occurs predominantly at the beginning of genes, whereas the latter can occur throughout transcribed regions. Most notably, specific methylation events are associated with the beginning, middle, and end of actively transcribed genes. These maps provide the foundation for further understanding the roles of chromatin in gene expression and genome maintenance.

Specialized gene expression programs are induced by signaling pathways that act on transcription factors. Whether these transcription factors can function in multiple developmental programs through a global switch in promoter selection is not known. We have used genome-wide location analysis to show that the yeast Ste12 transcription factor, which regulates mating and filamentous growth, is bound to distinct program-specific target genes dependent on the developmental condition. This condition-dependent distribution of Ste12 requires concurrent binding of the transcription factor Tec1 during filamentation and is differentially regulated by the MAP kinases Fus3 and Kss1. Program-specific distribution across the genome may be a general mechanism by which transcription factors regulate distinct gene expression programs in response to signaling.

Genome-wide location analysis was used to determine how the yeast cell cycle gene expression program is regulated by each of the nine known cell cycle transcriptional activators. We found that cell cycle transcriptional activators that function during one stage of the cell cycle regulate transcriptional activators that function during the next stage. This serial regulation of transcriptional activators forms a connected regulatory network that is itself a cycle. Our results also reveal how the nine transcriptional regulators coordinately regulate global gene expression and diverse stage-specific functions to produce a continuous cycle of cellular events. This information forms the foundation for a complete map of the transcriptional regulatory network that controls the cell cycle.