Long-range promoter-enhancer contacts are conserved during evolution and contribute to gene expression robustness

Gene expression is regulated through complex molecular interactions, involving cis-acting elements that can be situated far away from their target genes. Data on long-range contacts between promoters and regulatory elements is rapidly accumulating. However, it remains unclear how these regulatory relationships evolve and how they contribute to the establishment of robust gene expression profiles. Here, we address these questions by comparing genome-wide maps of promoter-centered chromatin contacts in mouse and human. We show that there is significant evolutionary conservation of cis-regulatory landscapes, indicating that selective pressures act to preserve not only regulatory element sequences but also their chromatin contacts with target genes. The extent of evolutionary conservation is remarkable for long-range promoter-enhancer contacts, illustrating how the structure of regulatory landscapes constrains large-scale genome evolution. We show that the evolution of cis-regulatory landscapes, measured in terms of distal element sequences, synteny or contacts with target genes, is significantly associated with gene expression evolution.

Sex, Gender, and Diversity Analysis in Research Policies of Major Public Granting Agencies: A Global Review

National research agencies are funded by taxpayer monies and, as such, are responsible for promoting excellent research that benefits all of society. Integrating sex, gender and diversity analysis (SG&DA) into the design of research, where relevant, can improve research methodology and provide new insights. To realize this potential, funding agencies have developed policies for integrating this type of analysis into the grant proposal process. This study reviews those policies for 23 agencies across six continents. Overall, one agency achieved superior performance, six agencies scored excellent performance, five showed average performance, two need some improvement and nine require improvement. Our study developed a five-part SG&DA policy roadmap for agencies and collected best practices across that guide. Standard practices, tailored as appropriate to country-specific cultures and regulatory landscapes, will enhance collaboration potential, global equity, research excellence and reproducibility.

Cell-specific alterations in Pitx1 regulatory landscape activation caused by the loss of a single enhancer

Developmental genes are frequently controlled by multiple enhancers sharing similar specificities. As a result, deletions of such regulatory elements have often failed to reveal their full function. Here, we use the Pitx1 testbed locus to characterize in detail the regulatory and cellular identity alterations following the deletion of one of its enhancers (Pen). By combining single cell transcriptomics and an in-embryo cell tracing approach, we observe an increased fraction of Pitx1 non/low-expressing cells and a decreased fraction of Pitx1 high-expressing cells. We find that the over-representation of Pitx1 non/low-expressing cells originates from a failure of the Pitx1 locus to coordinate enhancer activities and 3D chromatin changes. This locus mis-activation induces a localized heterochrony and a concurrent loss of irregular connective tissue, eventually leading to a clubfoot phenotype. This data suggests that, in some cases, redundant enhancers may be used to locally enforce a robust activation of their host regulatory landscapes.

THEORISING THE LOCAL ADAPTATION OF TRANSNATIONAL CERTIFICATION STANDARDS

Over the last three decades, transnational certification standards have proliferated to fill perceived ‘governance gaps’ in developing countries. Transnational non-governmental organisations and private standards-setting agencies have developed standards that cover a vast range of areas such as labour rights, social justice and environmental protection. As a form of private transnational regulation, certification standards travel through transnational production networks that link lead firms in developed countries with supplier firms in developing countries. This article draws on a case study about coffee certification to challenge the conventional understanding of transnational certification as a contractual conduit that transfers encoded certification standards from senders to receivers. It shows how transnational certification standards interact with, and remake local regulatory landscapes as they pass through. This interaction between global and local knowledge compels us to see transnational standards as a protean, highly localised regulatory process rather than stable universal norms. The article concludes that transnational certification does not function like an integrated ‘joined-up’ process and it is better understood as a mode of polycentric regulation that decentres and fragments transnational norms and standards.

Artificial Intelligence, Heuristic Biases, and the Optimization of Health Outcomes: Cautionary Optimism

The use of artificial intelligence (AI) and machine learning (ML) in clinical care offers great promise to improve patient health outcomes and reduce health inequity across patient populations. However, inherent biases in these applications, and the subsequent potential risk of harm can limit current use. Multi-modal workflows designed to minimize these limitations in the development, implementation, and evaluation of ML systems in real-world settings are needed to improve efficacy while reducing bias and the risk of potential harms. Comprehensive consideration of rapidly evolving AI technologies and the inherent risks of bias, the expanding volume and nature of data sources, and the evolving regulatory landscapes, can contribute meaningfully to the development of AI-enhanced clinical decision making and the reduction in health inequity.

Sequential in cis mutagenesis in vivo reveals various functions for CTCF sites at the mouse HoxD cluster

Mammalian Hox gene clusters contain a range of CTCF binding sites. In addition to their importance in organizing a TAD border, which isolates the most posterior genes from the rest of the cluster, the positions and orientations of these sites suggest that CTCF may be instrumental in the selection of various subsets of contiguous genes, which are targets of distinct remote enhancers located in the flanking regulatory landscapes. We examined this possibility by producing an allelic series of cumulative in cis mutations in these sites, up to the abrogation of CTCF binding in the five sites located on one side of the TAD border. In the most impactful alleles, the global chromatin architecture of the locus was modified, yet not drastically, illustrating that CTCF sites located on one side of a strong TAD border are sufficient to organize at least part of this insulation. Spatial colinearity in the expression of these genes along the major body axis was nevertheless maintained, despite abnormal expression boundaries. In contrast, strong effects were scored in the selection of target genes responding to particular enhancers, leading to the misregulation of Hoxd genes in specific structures. Altogether, while most enhancer–promoter interactions can occur in the absence of this series of CTCF sites, the binding of CTCF in the Hox cluster is required to properly transform a rather unprecise process into a highly discriminative mechanism of interactions, which is translated into various patterns of transcription accompanied by the distinctive chromatin topology found at this locus. Our allelic series also allowed us to reveal the distinct functional contributions for CTCF sites within this Hox cluster, some acting as insulator elements, others being necessary to anchor or stabilize enhancer–promoter interactions, and some doing both, whereas they all together contribute to the formation of a TAD border. This variety of tasks may explain the amazing evolutionary conservation in the distribution of these sites among paralogous Hox clusters or between various vertebrates.

Promoter repression and 3D-restructuring resolves divergent developmental gene expression in TADs

Cohesin loop extrusion facilitates precise gene expression by continuously driving promoters to sample all enhancers located within the same topologically-associated domain (TAD). However, many TADs contain multiple genes with divergent expression patterns, thereby indicating additional forces further refine how enhancer activities are utilised. Here, we unravel the mechanisms enabling a new gene, Rex1, to emerge with divergent expression within the ancient Fat1 TAD in placental mammals. We show that such divergent expression is not determined by a strict enhancer-promoter compatibility code, intra-TAD position or nuclear envelope-attachment. Instead, TAD-restructuring in embryonic stem cells (ESCs) separates Rex1 and Fat1 with distinct proximal enhancers that independently drive their expression. By contrast, in later embryonic tissues, DNA methylation renders the inactive Rex1 promoter profoundly unresponsive to Fat1 enhancers within the intact TAD. Combined, these features adapted an ancient regulatory landscape during evolution to support two entirely independent Rex1 and Fat1 expression programs. Thus, rather than operating only as rigid blocks of co-regulated genes, TAD-regulatory landscapes can orchestrate complex divergent expression patterns in evolution.

Building regulatory landscapes: enhancer recruits cohesin to create contact domains, engage CTCF sites and activate distant genes

ABSTRACTDevelopmental gene expression is often controlled by distal tissue-specific enhancers. Enhancer action is restricted to topological chromatin domains, typically formed by cohesin-mediated loop extrusion between CTCF-associated boundaries. To better understand how individual regulatory DNA elements form topological domains and control expression, we used a bottom-up approach, building active regulatory landscapes of different sizes in inactive chromatin. We demonstrate that transcriptional output and protection against gene silencing reduces with increased enhancer distance, but that enhancer contact frequencies alone do not dictate transcription activity. The enhancer recruits cohesin to stimulate the formation of local chromatin contact domains and activate flanking CTCF sites for engagement in chromatin looping. Small contact domains can support strong and stable expression of distant genes. The enhancer requires transcription factors and mediator to activate genes over all distance ranges, but relies on cohesin exclusively for the activation of distant genes. Our work supports a model that assigns two functions to enhancers: its classic role to stimulate transcription initiation and elongation from target gene promoters and a role to recruit cohesin for the creation of contact domains, the engagement of flanking CTCF sites in chromatin looping, and the activation of distal target genes.

Street markets, urban development and immigrant entrepreneurship: Unpacking precarity in Moore Street, Dublin

Traditional markets represent vital spaces of opportunity for livelihood-building, intercultural contact and for developing familiarity with the city. Yet, worldwide, markets are under pressure due to redevelopment agendas driven by neoliberalised forms of urban governance. Although precarious sites of occupation and employment, markets still maintain an attractiveness for immigrant micro-entrepreneurs as a foothold into the labour market and urban economy. Through a case study of the historic Moore Street market in central Dublin (Ireland), we explore the experiences of immigrant entrepreneurs. While these may be different in terms of their familiarity with the urban, institutional and regulatory landscapes, they are not entirely dissimilar from the experiences of longer-term traders in Moore Street. However what is evident is how precarity is tactically exploited by newcomer entrepreneurs for particular reasons. These traders prize the autonomy brought by market trading and use it as a meso-scale between low-paid waged employment and higher-level employment that may be out of reach for a variety of reasons. We argue that in examining urban precarity, increased attention should be paid to exploring the context-specific nature of the processes that give rise to it as well as the agentic capacity exercised to exploit it even within structural constraints.

CTCF knockout in zebrafish induces alterations in regulatory landscapes and developmental gene expression

Coordinated chromatin interactions between enhancers and promoters are critical for gene regulation. The architectural protein CTCF mediates chromatin looping and is enriched at the boundaries of topologically associating domains (TADs), which are sub-megabase chromatin structures. In vitro CTCF depletion leads to a loss of TADs but has only limited effects over gene expression, challenging the concept that CTCF-mediated chromatin structures are a fundamental requirement for gene regulation. However, how CTCF and a perturbed chromatin structure impacts gene expression during development remains poorly understood. Here we link the loss of CTCF and gene regulation during patterning and organogenesis in a ctcf knockout zebrafish model. CTCF absence leads to loss of chromatin structure and affects the expression of thousands of genes, including many developmental regulators. Our results demonstrate the essential role of CTCF in providing the structural context for enhancer-promoter interactions, thus regulating developmental genes.