Biological Questions Research Articles

Probabilistic association of differentially expressed genes with cis-regulatory elements.

Differential gene expression in response to perturbations is mediated at least in part by changes in binding of transcription factors (TFs) and other proteins at specific genomic regions. Association of these cis-regulatory elements (CREs) with their target genes is a challenging task that is essential to address many biological and mechanistic questions. Many current approaches rely on chromatin conformation capture techniques or single-cell correlational methods to establish CRE-to-gene associations. These methods can be effective but have limitations, including resolution, gaps in detectable association distances, and cost. As an alternative, we have developed DegCre, a nonparametric method that evaluates correlations between measurements of perturbation-induced differential gene expression and differential regulatory signal at CREs to score possible CRE-to-gene associations. It has several unique features, including the ability to use any type of CRE activity measurement, yield probabilistic scores for CRE-to-gene pairs, and assess CRE-to-gene pairings across a wide range of sequence distances. We apply DegCre to six data sets, each using different perturbations and containing a variety of regulatory signal measurements, including chromatin openness, histone modifications, and TF occupancy. To test their efficacy, we compare DegCre associations to Hi-C loop calls and CRISPR-validated CRE-to-gene associations, establishing good performance by DegCre that is comparable or superior to competing methods. DegCre is a novel approach to the association of CREs to genes from a perturbation-differential perspective, with strengths that are complementary to existing approaches and allow for new insights into gene regulation.

A translational and multidisciplinary approach to studying the Garcia effect, a higher form of learning with deep evolutionary roots.

Animals, including humans, learn and remember to avoid a novel food when its ingestion is followed, hours later, by sickness - a phenomenon initially identified during World War II as a potential means of pest control. In the 1960s, John Garcia (for whom the effect is now named) demonstrated that this form of conditioned taste aversion had broader implications, showing that it is a rapid but long-lasting taste-specific food aversion with a fundamental role in the evolution of behaviour. From the mid-1970s onward, the principles of the Garcia effect were translated to humans, showing its role in different clinical conditions (e.g. side-effects linked to chemotherapy). However, in the last two decades, the number of studies on the Garcia effect has undergone a considerable decline. Since its discovery in rodents, this form of learning was thought to be exclusive to mammals; however, we recently provided the first demonstration that a Garcia effect can be formed in an invertebrate model organism, the pond snail Lymnaea stagnalis. Thus, in this Commentary, after reviewing the experiments that led to the first characterization of the Garcia effect in rodents, we describe the recent evidence for the Garcia effect in L. stagnalis, which may pave the way for future studies in other invertebrates and mammals. This article aims to inspire future translational and ecological studies that characterize the conserved mechanisms underlying this form of learning with deep evolutionary roots, which can be used to address a range of different biological questions.

Exploring stem cell frontiers: definitions, challenges, and perspectives for regenerative medicine.

Each year, the European Summer School on Stem Cell Biology and Regenerative Medicine (SCSS) attracts early-career researchers and actively practicing clinicians who specialise in stem cell and regenerative biology. The 16th edition of this influential course took place from 12th to 19th September 2023 on the charming Greek island of Spetses. Focusing on important concepts and recent advances in stem cells, the distinguished faculty included experts spanning the spectrum from fundamental research to clinical trials to market-approved therapies. Alongside an academically intensive programme that bridges the various contexts of stem cell research, delegates were encouraged to critically address relevant questions in stem cell biology and medicine, including broader societal implications. Here, we present a comprehensive overview and key highlights from the SCSS 2023.

Micro- and Macroevolution: A Continuum or Two Distinct Types of Change?

How microevolution and macroevolution are related is one of the major unanswered questions in evolutionary biology. The most prevalent view is that microevolution and macroevolution are part of a continuum of one type of change and that macroevolution is the cumulative result of microevolution. Mathematics, however, distinguishes two fundamentally different, singular types of change: change of a vector in its parameters versus its dimensions. This mathematical distinction may help to articulate the concept of evolution by distinction of two fundamentally different types of evolution: the change of the state vector of an organism in 1) its parameters (= ‘first-order evolution’) and 2) its dimensions (= ‘second-order evolution’). This distinction can be operationalized by identifying genes and regulatory elements in the nucleotide code of an organism as dimensions and the level of expression as parameters of its state vector. This operationalization allows us to substitute the phenotype-based analysis of evolution with a genotype-based analysis and draws attention to the molecular mechanisms that change the parameters or the dimensions of the state vector, respectively. We illustrate the distinction between first- and second-order evolution with a simulation of the adaptive dynamics of a population of digital amoebae. Our genotype-based systems approach reveals that micro- and macroevolution are largely similar to first- and second-order evolution respectively, and are not a continuum of change.

Extreme genome scrambling in marine planktonic Oikopleura dioica cryptic species.

Genome structural variations within species are rare. How selective constraints preserve gene order and chromosome structure is a central question in evolutionary biology that remains unsolved. Our sequencing of several genomes of the appendicularian tunicate Oikopleura dioica around the globe reveals extreme genome scrambling caused by thousands of chromosomal rearrangements, although showing no obvious morphological differences between these animals. The breakpoint accumulation rate is an order of magnitude higher than in ascidian tunicates, nematodes, Drosophila, or mammals. Chromosome arms and sex-specific regions appear to be the primary unit of macrosynteny conservation. At the microsyntenic level, scrambling did not preserve operon structures, suggesting an absence of selective pressure to maintain them. The uncoupling of the genome scrambling with morphological conservation in O. dioica suggests the presence of previously unnoticed cryptic species and provides a new biological system that challenges our previous vision of speciation in which similar animals always share similar genome structures.

Refinement of a technique for collecting and evaluating the osmolality of haemolymph from Drosophila larvae.

Ex vivo physiological experiments using small insect models such as Drosophila larvae have become increasingly useful to address fundamental biological questions. To perform such experiments, various artificial saline solutions have been developed, but their osmolality varies significantly from one to the next. Such a variation of osmolality stems, in part, from the difficulty of determining the true value of haemolymph osmolality in Drosophila larvae. Thus, there is a pressing need to refine protocols for collecting and measuring the osmolality of the larval haemolymph. Two major obstacles are thought to impede the accurate analysis of haemolymph collected from small insects: melanin formation and gut-derived contamination. Here, we greatly refined existing haemolymph collection methods, evaluated the purity of the collected haemolymph under melanin-free conditions, and concluded that the true value of haemolymph osmolality is close to 306.0 mOsm kg-1 in Drosophila larvae.

Demuxafy: improvement in droplet assignment by integrating multiple single-cell demultiplexing and doublet detection methods

Recent innovations in single-cell RNA-sequencing (scRNA-seq) provide the technology to investigate biological questions at cellular resolution. Pooling cells from multiple individuals has become a common strategy, and droplets can subsequently be assigned to a specific individual by leveraging their inherent genetic differences. An implicit challenge with scRNA-seq is the occurrence of doublets—droplets containing two or more cells. We develop Demuxafy, a framework to enhance donor assignment and doublet removal through the consensus intersection of multiple demultiplexing and doublet detecting methods. Demuxafy significantly improves droplet assignment by separating singlets from doublets and classifying the correct individual.

Asymmetric segregation of maternal mRNAs and germline-related determinants in Cephalochordate embryos: implications for the evolution of early patterning events in chordates.

How animal embryos determine their early cell fates is an important question in developmental biology. In various model animals asymmetrically localized maternal transcripts play important roles in axial patterning and cell fate specification. Cephalochordates (amphioxus), which have three living genera (Asymmetron, Epigonichthys, Branchiostoma), are an early branching chordate lineage and thus occupy a key phylogenetic position for understanding the evolution of chordate developmental mechanisms. It has been shown that in the zygote of Brachiostoma amphioxus, which possess bilateral gonads flanking both sides of their trunk region, maternal transcripts of germline determinants form a compact granule. During early embryogenesis this granule is inherited by a single blastomere that subsequently gives rise to a cluster of cells displaying typical characteristics of primordial germ cells (PGC). These PGCs then come to lie in the tailbud region and proliferate during posterior elongation of the larva to join in the gonad anlagen at the ventral tip of the developing myomeres in amphioxus larvae. However, in Asymmetron and Epigonichthys amphioxus, whose gonads are present only on the right side of their body, nothing is known about their PGC development or the cellular/morphogenetic processes resulting in the asymmetric distribution of gonads. Using conserved germline determinants as markers, we show that similarly to Brachiostoma amphioxus, Asymmetron also employ a preformation mechanism to specify their PGCs, suggesting that this mechanism represents an ancient trait dating back to the common ancestor of Cephalochordates. Surprisingly, we found that Asymmetron PGCs are initially deposited on both sides of the body during early larval development; however, the left side PGCs cease to exist in young juveniles, suggesting that PGCs are eliminated from the left body side during larval development or following metamorphosis. This is reminiscent of the PGC development in the sea urchin embryo, and we discuss the implications of this observation for the evolution of developmental mechanisms.

Mathematical Modelling of Parasite Dynamics: A Stochastic Simulation-Based Approach and Parameter Estimation via Modified Sequential-Type Approximate Bayesian Computation

The development of mathematical models for studying newly emerging and re-emerging infectious diseases has gained momentum due to global events. The gyrodactylid-fish system, like many host-parasite systems, serves as a valuable resource for ecological, evolutionary, and epidemiological investigations owing to its ease of experimental manipulation and long-term monitoring. Although this system has an existing individual-based model, it falls short in capturing information about species-specific microhabitat preferences and other biological details for different Gyrodactylus strains across diverse fish populations. This current study introduces a new individual-based stochastic simulation model that uses a hybrid τ\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$\ au $$\\end{document}-leaping algorithm to incorporate this essential data, enhancing our understanding of the complexity of the gyrodactylid-fish system. We compare the infection dynamics of three gyrodactylid strains across three host populations. A modified sequential-type approximate Bayesian computation (ABC) method, based on sequential Monte Carlo and sequential importance sampling, is developed. Additionally, we establish two penalised local-linear regression methods (based on L1 and L2 regularisations) for ABC post-processing analysis to fit our model using existing empirical data. With the support of experimental data and the fitted mathematical model, we address open biological questions for the first time and propose directions for future studies on the gyrodactylid-fish system. The adaptability of the mathematical model extends beyond the gyrodactylid-fish system to other host-parasite systems. Furthermore, the modified ABC methodologies provide efficient calibration for other multi-parameter models characterised by a large set of correlated or independent summary statistics.

Halophytes and heavy metals: A multi-omics approach to understand the role of gene and genome duplication in the abiotic stress tolerance of Cakile maritima.

The origin of diversity is a fundamental biological question. Gene duplications are one mechanism that provides raw material for the emergence of novel traits, but evolutionary outcomes depend on which genes are retained and how they become functionalized. Yet, following different duplication types (polyploidy and tandem duplication), the events driving gene retention and functionalization remain poorly understood. Here we used Cakile maritima, a species that is tolerant to salt and heavy metals and shares an ancient whole-genome triplication with closely related salt-sensitive mustard crops (Brassica), as a model to explore the evolution of abiotic stress tolerance following polyploidy. Using a combination of ionomics, free amino acid profiling, and comparative genomics, we characterize aspects of salt stress response in C. maritima and identify retained duplicate genes that have likely enabled adaptation to salt and mild levels of cadmium. Cakile maritima is tolerant to both cadmium and salt treatments through uptake of cadmium in the roots. Proline constitutes greater than 30% of the free amino acid pool in C. maritima and likely contributes to abiotic stress tolerance. We find duplicated gene families are enriched in metabolic and transport processes and identify key transport genes that may be involved in C. maritima abiotic stress tolerance. These findings identify pathways and genes that could be used to enhance plant resilience and provide a putative understanding of the roles of duplication types and retention on the evolution of abiotic stress response.

Compensatory growth and recovery of cartilage cytoarchitecture after transient cell death in fetal mouse limbs

A major question in developmental and regenerative biology is how organ size and architecture are controlled by progenitor cells. While limb bones exhibit catch-up growth (recovery of a normal growth trajectory after transient developmental perturbation), it is unclear how this emerges from the behaviour of chondroprogenitors, the cells sustaining the cartilage anlagen that are progressively replaced by bone. Here we show that transient sparse cell death in the mouse fetal cartilage is repaired postnatally, via a two-step process. During injury, progression of chondroprogenitors towards more differentiated states is delayed, leading to altered cartilage cytoarchitecture and impaired bone growth. Then, once cell death is over, chondroprogenitor differentiation is accelerated and cartilage structure recovered, including partial rescue of bone growth. At the molecular level, ectopic activation of mTORC1 correlates with, and is necessary for, part of the recovery, revealing a specific candidate to be explored during normal growth and in future therapies.

Abstract LB187: Investigating the roles of ROR1/2 in non-small cell lung carcinoma NSCLC and potential therapeutic applications

Abstract Lung cancer is the leading cause of cancer mortality worldwide, representing an unmet medical need. Receptor tyrosine kinase-like orphan receptors (RORs) are oncofetal antigens that act as receptors/co-receptors of WNT5A and other related noncanonical WNT proteins, in transducing β-catenin independent WNT signaling. Dysregulated ROR/WNT have been linked to cancer progression and therapy resistance. Recently, ROR receptors have drawn increasing attention as anti-cancer targets of antibody-drug conjugates (ADCs) due to their selective expression on the tumor cell surface. The ROR family contains ROR1 and ROR2, which share 58% sequence homology. However, their functional interactions between ROR/WNT5A and whether RORs have distinct or redundant roles in tumor phenotypes remain unclear. In this study, we sought to address these biological questions in NSCLC, and investigate the potential application of anti-ROR therapies. To this end, we used CRISPR/Cas9 to generate clones of ROR1 or ROR2 single knockouts (KOs) in the NSCLC LCLC-103H cell line and implemented an array of assays to characterize the effect of single ROR KOs. ROR KOs were confirmed by both western blot and flow cytometry. We found reduced ROR2 mean fluorescence intensity (MFI) on ROR1 KOs and increased ROR1 MFI on ROR2 KOs in flow cytometrical analysis. Phenotypically, ROR1 KOs did not affect tumor doubling time, but ROR2 KOs promoted cell proliferation in 2D cultures. Unlike 2D cultures, tumor 3D spheroid proliferation was significantly inhibited in both ROR1 and ROR2 KOs, implying that each ROR is indispensable in maintaining tumor growth featured by cell-cell/extracellular matrix interactions. Mechanistically, ROR1 KOs tended to decrease ROR2 and WNT5A/B protein expression, while ROR2 KOs caused compensatory upregulation of ROR1 and WNT5A/B. Both ROR1 and ROR2 KOs decreased SNAIL expression, which is a key driver of epithelial-to-mesenchymal plasticity (EMP). Furthermore, alterations in ROR1/2 signaling pathways were observed. We have identified pathways downstream of ROR1/2 including WNT, EMP, and survival, which may shed light on the biological roles of RORs in cancer and potential therapeutic intervention. Finally, the therapeutic application of ROR1/2 was evaluated by monomethyl auristatin F (MMAF) containing anti-ROR ADCs. ROR1 KOs demonstrated the on-target cytotoxicity of anti-ROR1 ADC, while ROR2 KOs showed increased sensitivity to anti-ROR1 ADC due to the increased ROR1 expression in ROR2 KOs. In vivo analysis is currently underway. Collectively, our results demonstrate that RORs have both distinct and compensatory functions in regulating tumor proliferation. We believe a greater understanding in ROR1/2 biology may lead to better targeted therapeutic applications. Citation Format: Wen-An Pan, Xin Guo, Christina Galang, Mira Ko, Jeff Kucharski, Nicole Miller, Haim Moskowitz, Jeff Watkins, Brian Lannutti, Katti Jessen, Hao Liu. Investigating the roles of ROR1/2 in non-small cell lung carcinoma NSCLC and potential therapeutic applications [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB187.

Abstract SY38-02: Clinical investigations of obesity in cancer: BMI and other confounders

Abstract SY38-02: Clinical investigations of obesity in cancer: BMI and other confounders

Investigation of the Allelopathic Effect of Two Invasive Plant Species in Rhizotron System.

A key question in plant invasion biology is why invasive plants are more competitive in their introduced habitat than in their native habitat. Studies show that invasive species exhibit allelopathy, influencing other plants by releasing chemicals. Research on allelopathy uses in vitro tests, investigating effects on seed germination and seedling development. Although soil plays a role in modifying allelopathic effects, observations with soil are rare and almost nothing is known about the root development of test plants developing in soil and the effects of allelopathic compounds on root architecture. Our study evaluates the allelopathic effects of false indigo-bush (Amorpha fruticosa L.) and common milkweed (Asclepias syriaca L.) on oilseed rape growth as a model plant. The rhizotron system was used to study the effect of morphology and root architecture. Leaf-soil mixtures at 0.5%, 1%, and 5% concentrations were used. Shoot and root development was strongly inhibited at 5%. But there was no difference between the allelopathy of the two species, and the application of lower concentrations did not show any effect, demonstrating that soil has a significant modifying effect on their allelopathy. Our results highlight that the development of roots growing in the soil is also worth investigating in connection with allelopathy, which can strengthen the ecological importance of allelochemicals during successful invasions.

The effect of pseudoknot base pairing on cotranscriptional structural switching of the fluoride riboswitch.

A central question in biology is how RNA sequence changes influence dynamic conformational changes during cotranscriptional folding. Here we investigated this question through the study of transcriptional fluoride riboswitches, non-coding RNAs that sense the fluoride anion through the coordinated folding and rearrangement of a pseudoknotted aptamer domain and a downstream intrinsic terminator expression platform. Using a combination of Escherichia coli RNA polymerase in vitro transcription and cellular gene expression assays, we characterized the function of mesophilic and thermophilic fluoride riboswitch variants. We showed that only variants containing the mesophilic pseudoknot function at 37°C. We next systematically varied the pseudoknot sequence and found that a single wobble base pair is critical for function. Characterizing thermophilic variants at 65°C through Thermus aquaticus RNA polymerase in vitro transcription showed the importance of this wobble pair for function even at elevated temperatures. Finally, we performed all-atom molecular dynamics simulations which supported the experimental findings, visualized the RNA structure switching process, and provided insight into the important role of magnesium ions. Together these studies provide deeper insights into the role of riboswitch sequence in influencing folding and function that will be important for understanding of RNA-based gene regulation and for synthetic biology applications.

Nuclear deformation and cell division of single cell on elongated micropatterned substrates fabricated by DMD lithography

Cells sense mechanical signals from the surrounding environment and transmit them to the nucleus through mechanotransduction to regulate cellular behavior. Microcontact printing, which utilizes elastomer stamps, is an effective method for simulating the cellular microenvironment and manipulating cell morphology. However, the conventional fabrication process of silicon masters and elastomer stamps requires complex procedures and specialized equipment, which restricts the widespread application of micropatterning in cell biology and hinders the investigation of the role of cell geometry in regulating cell behavior. In this study, we present an innovative method for convenient resin stamp microfabrication based on digital micromirror device planar lithography. Using this method, we generated a series of patterns ranging from millimeter to micrometer scales and validated their effectiveness in controlling adhesion at both collective and individual cell levels. Additionally, we investigated mechanotransduction and cell behavior on elongated micropatterned substrates. We then examined the effects of cell elongation on cytoskeleton organization, nuclear deformation, focal adhesion formation, traction force generation, nuclear mechanics, and the growth of HeLa cells. Our findings reveal a positive correlation between cell length and mechanotransduction. Interestingly, HeLa cells with moderate length exhibit the highest cell division and proliferation rates. These results highlight the regulatory role of cell elongation in mechanotransduction and its significant impact on cancer cell growth. Furthermore, our methodology for controlling cell adhesion holds the potential for addressing fundamental questions in both cell biology and biomedical engineering.

ARGONAUTE10 controls cell fate specification and formative cell divisions in the Arabidopsis root

A key question in plant biology is how oriented cell divisions are integrated with patterning mechanisms to generate organs with adequate cell type allocation. In the root vasculature, a gradient of miRNA165/6 controls the abundance of HD-ZIP III transcription factors, which in turn control cell fate and spatially restrict vascular cell proliferation to specific cells. Here, we show that vascular development requires the presence of ARGONAUTE10, which is thought to sequester miRNA165/6 and protect HD-ZIP III transcripts from degradation. Our results suggest that the miR165/6-AGO10-HDZIP III module acts by buffering cytokinin responses and restricting xylem differentiation. Mutants of AGO10 show faster growth rates and strongly enhanced survival under severe drought conditions. However, this superior performance is offset by markedly increased variation and phenotypic plasticity in sub-optimal carbon supply conditions. Thus, AGO10 is required for the control of formative cell division and coordination of robust cell fate specification of the vasculature, while altering its expression provides a means to adjust phenotypic plasticity.

General Principles, Designs, and Statistical Analyses in Experimental Animal Studies

Research using animals contributes significantly to many research and development studies, especially in the biomedical field. Within the scope of the study, conducting animal experiments in accordance with scientific principles and ensuring the ethical use and welfare of animals are issues that should be taken into consideration. In this context, the scientific contribution to be achieved by conducting these studies in line with scientific and ethical principles will be directly proportional. In many studies, while investigating the biological significance, it is seen that the effects of many factors are ignored, the answer to the biological question is investigated with simple experimental designs, or the accurate statistical analyses are not chosen. Therefore, in this study, the principles that a researcher planning an animal study should follow within the scope of the research (animal ethics, 3R, and other R rules, determination of sample size, randomization, and blinding) are briefly mentioned. Then, completely randomized design, regression design, split-unit design, hierarchical (nested) design, mixed effects design, and appropriate statistical analyses for these designs, which are thought to be useful in these studies, are discussed. It is thought that this review will be useful as it contains important summative information that will guide all researchers in planning animal studies accurately and quickly.

ICDP workshop on the Lake Victoria Drilling Project (LVDP): scientific drilling of the world's largest tropical lake

Abstract. Lake Victoria, which is bordered by Uganda, Tanzania, Kenya, and has a catchment that extends to Rwanda and Burundi, is home to the largest human population surrounding any lake in the world and provides critical resources across eastern Africa. Lake Victoria is also the world's largest tropical lake by surface area, but it is relatively shallow and without a major inlet, making it very sensitive to changes in climate, and especially hydroclimate. Furthermore, its size creates abundant habitats for aquatic fauna, including the iconic hyper-diverse cichlids, and serves as a major geographic barrier to terrestrial fauna across equatorial Africa. Given Lake Victoria's importance to the eastern African region, its sensitivity to climate, and its influences on terrestrial and aquatic faunal evolution and dispersal, it is vital to understand the connection between the lake and regional climate and how the lake size, shape, and depth have changed through its depositional history. This information can only be ascertained by collecting a complete archive of Lake Victoria's sedimentary record. To evaluate the Lake Victoria basin as a potential drilling target, ∼ 50 scientists from 10 countries met in Dar es Salaam, Tanzania, in July 2022 for the International Continental Scientific Drilling Program (ICDP)-sponsored Lake Victoria Drilling Project (LVDP) workshop. Discussions of the main scientific objectives for a future drilling project included (1) recovering the Pleistocene and Holocene sedimentary records of Lake Victoria that document the dynamic nature of the lake, including multiple lacustrine and paleosol sequences; (2) establishing the chronology of recovered sediments, including using extensive tephra fingerprinting and other techniques from deposits in the region; (3) reconstructing past climate, environment, lacustrine conditions, and aquatic fauna, using an integrated multi-proxy approach, combined with climate and hydrologic modeling; and (4) connecting new records with existing sedimentary snapshots and fossils exposed in deposits around the lake, tying archaeological, paleontological, sedimentological, tectonic, and volcanic findings to new drilling results. The LVDP provides an innovative way to address critical geological, paleontological, climatological, and evolutionary biological questions about Quaternary to modern landscapes and ecosystems in eastern Africa. Importantly, this project affords an excellent opportunity to help develop conservation and management strategies for regional responses to current and future changes in climate, land use, fisheries, and resiliency of at-risk communities in equatorial Africa.

Botany and geogenomics: Constraining geological hypotheses in the neotropics with large-scale genetic data derived from plants.

Decades of empirical research have revealed how the geological history of our planet shaped plant evolution by establishing well-known patterns (e.g., how mountain uplift resulted in high rates of diversification and replicate radiations in montane plant taxa). This follows a traditional approach where botanical data are interpreted in light of geological events. In this synthesis, I instead describe how by integrating natural history, phylogenetics, and population genetics, botanical research can be applied alongside geology and paleontology to inform our understanding of past geological and climatic processes. This conceptual shift aligns with the goals of the emerging field of geogenomics. In the neotropics, plant geogenomics is a powerful tool for the reciprocal exploration of two long standing questions in biology and geology: how the dynamic landscape of the region came to be and how it shaped the evolution of the richest flora. Current challenges that are specific to analytical approaches for plant geogenomics are discussed. I describe the scale at which various geological questions can be addressed from biological data and what makes some groups of plants excellent model systems for geogenomics research. Although plant geogenomics is discussed with reference to the neotropics, the recommendations given here for approaches to plant geogenomics can and should be expanded to exploring long-standing questions on how the earth evolved with the use of plant DNA.