Biology subjects

Mukherjee, A.

Publications and source records attributed to Mukherjee, A..

At least 73 records · Page 4Linked to original sources

bioRxiv2024

Checkpoint-independent association of Mad3BUBR1 with Stu1CLASP directs homolog alignment in meiosis I

Gametes are produced via meiosis, a specialized cell division associated with frequent errors which cause birth defects and infertility. Uniquely in meiosis I, homologous chromosomes segregate to opposite poles, usually requiring their linkage by chiasmata, the products of crossover recombination1. The spindle checkpoint delays cell cycle progression until all chromosomes are properly attached to microtubules2 but the steps leading to the capture and alignment of chromosomes on the meiosis I spindle remain poorly understood. In budding yeast meiosis I, Mad2 and Mad3BUBR1 are equally important for spindle checkpoint delay, but biorientation of homologs on the meiosis I spindle requires Mad2, but not Mad3BUBR1 3,4. Here we show that Mad3BUBR1 promotes accurate meiosis I homolog segregation outside its canonical checkpoint role, independently of Mad2. We find that Mad3BUBR1 associates with the TOGL1 domain of Stu1CLASP, a conserved plus-end microtubule protein which is important for chromosome capture onto the spindle. Homologous chromosome pairs that are proficient in crossover formation, but which fail to biorient, rely on Mad3BUBR1-Stu1CLASP to ensure their efficient attachment to microtubules and segregation during meiosis I. Furthermore, we show that Mad3BUBR1-Stu1CLASP are essential to rescue the segregation of mini-chromosomes lacking crossovers. Our findings define a new pathway ensuring microtubule-dependent chromosome capture and demonstrate that spindle checkpoint proteins safeguard the fidelity of chromosome segregation both by actively promoting chromosome alignment and delaying cell cycle progression until this has occurred.

cell biology↗

bioRxiv2023

Analysing the distribution of SARS-CoV-2 infections in schools: integrating model predictions with real world observations

School closures were used as strategies to mitigate transmission in the COVID-19 pandemic. Understanding the nature of SARS-CoV-2 outbreaks and the distribution of infections in classrooms could help inform targeted or precision preventive measures and outbreak management in schools, in response to future pandemics. In this work, we derive an analytical model of Probability Density Function (PDF) of SARS-CoV-2 secondary infections and compare the model with infection data from all public schools in Ontario, Canada between September-December, 2021. The model accounts for major sources of variability in airborne transmission like viral load and dose-response (i.e., the human bodys response to pathogen exposure), air change rate, room dimension, and classroom occupancy. Comparisons between reported cases and the modeled PDF demonstrated the intrinsic overdispersed nature of the real-world and modeled distributions, but uncovered deviations stemming from an assumption of homogeneous spread within a classroom. The inclusion of near-field transmission effects resolved the discrepancy with improved quantitative agreement between the data and modeled distributions. This study provides a practical tool for predicting the size of outbreaks from one index infection, in closed spaces such as schools, and could be applied to inform more focused mitigation measures. Author summaryAt the start of the COVID-19 pandemic, there was huge uncertainty around the risks of SARS-CoV-2 spread in classrooms. In the absence of early predictions surrounding classroom risks, many jurisdictions across countries closed in-person education. There is great interest in adopting a more precision approach to better inform future interventions in the context of airborne virus risks. For this purpose, we need tools that can predict the probability of the size of outbreaks within classrooms along with the impact of interventions including masks, better ventilation, and physical distancing by limiting the number of students per classroom. To this end, we have developed a robust but practical model that yields the probability of secondary infections stemming from index cases occurring within schools on a given day. During model development, the major underlying physical and biological factors that dictate the disease transmission process, both at long-range and close-range, have been accounted for. This enables our model to modify its predictions for different scenarios - and possibly allows its use beyond schools. Finally, the models predictive capability has been verified by comparing its outputs with publicly available data on SARS-CoV-2 diagnoses in Ontario public schools. To our knowledge, this is the first time an analytical model derived from mostly first principles describes real-world infection distributions, satisfactorily. The quantitative match between the theoretical prediction and real-world data offers the proposed model as a possible powerful tool for better-informed precision pandemic mitigation strategies in indoor environments like schools.

biophysics↗

bioRxiv2023

Engineering water exchange is a safe and effective method for magnetic resonance imaging in diverse cell types

Aquaporin-1 (Aqp1), a water channel, has garnered significant interest for cell-based medicine and in vivo synthetic biology due to its ability to be genetically encoded to produce magnetic resonance signals by increasing the rate of water diffusion in cells. However, concerns regarding the effects of Aqp1 overexpression and increased membrane diffusivity on cell physiology have limited its widespread use as a deep-tissue reporter. In this study, we present evidence that Aqp1 generates strong diffusion-based magnetic resonance signals without adversely affecting cell viability or morphology in diverse cell lines derived from mice and humans. Our findings indicate that Aqp1 overexpression does not induce ER stress, which is frequently associated with heterologous expression of membrane proteins. Furthermore, we observed that Aqp1 expression had no detrimental effects on native biological activities, such as phagocytosis, immune response, insulin secretion, and tumor cell migration in the analyzed cell lines. These findings should serve to alleviate any lingering safety concerns regarding the utilization of Aqp1 as a genetic reporter and should foster its broader application as a noninvasive reporter for in vivo studies.

bioengineering↗

bioRxiv2023

Membrane potential mediates an ancient mechano-transduction mechanism for multi-cellular homeostasis

Mechanical forces have been shown to influence cellular decisions to grow, die, or differentiate, through largely mysterious mechanisms. Separately, changes in resting membrane potential have been observed in development, differentiation, regeneration, and cancer. We now demonstrate that membrane potential is the central mediator of cellular response to mechanical pressure. We show that mechanical forces acting on the cell change cellular biomass density, which in turn alters membrane potential. Membrane potential then regulates cell number density in epithelia by controlling cell growth, proliferation, and cell elimination. Mechanistically, we show that changes in membrane potential control signaling through the Hippo and MAPK pathways, and potentially other signaling pathways that originate at the cell membrane. While many molecular interactions are known to affect Hippo signaling, the upstream signal that activates the canonical Hippo pathway at the membrane has previously been elusive. Our results establish membrane potential as a central regulator of growth and tissue homeostasis.

cell biology↗

bioRxiv2023

Scale-independent topological interactions drive the first fate decision in the Drosophila embryo

During embryogenesis, the earliest cell fate decision is often linked to nuclear positioning, whose control arises from the integration of the cell cycle oscillator and associated cytoskeletal dynamics. Yet, the mechanisms that ensure that the correct number of nuclei move to the appropriate place remain poorly understood. Here, using light sheet microscopy, we show that in Drosophila embryos spindle orientation controls which nuclei migrate towards the cortex and which remains inside the embryo, thereby determining nuclear fate and the number of cells undergoing development. Combining computational methods inspired by integral geometry and manipulations of cell cycle genes, we show that spindle orientation is controlled by topological spindle-spindle interactions and not by internuclear distance. Using arguments describing the behavior of space-filling systems, we develop a theory for topological dependency in microtubule structures. Our work shows how topological interplay of microtubule mechanics can ensure robust control of density and cell fate determination.

developmental biology↗

bioRxiv2023

Lysine 36 of Drosophila histone H3.3 supports adult longevity

Aging is a multifactorial process that disturbs homeostasis, increases disease susceptibility, and ultimately results in death. Although the definitive set of molecular mechanisms responsible for aging remain to be discovered, epigenetic change over time is proving to be a promising piece of the puzzle. Several posttranslational histone modifications (PTMs) have been linked to the maintenance of longevity. Here, we focus on lysine-36 of the replication-independent histone protein, H3.3 (H3.3K36). To interrogate the role of this residue in Drosophila developmental gene regulation, we generated a lysine to arginine mutant that blocks the activity of its cognate modifying enzymes. We found that an H3.3BK36R mutation causes a significant reduction in adult lifespan, accompanied by dysregulation of the genomic and transcriptomic architecture. Transgenic co-expression of wild-type H3.3B completely rescues the longevity defect. Because H3.3 is known to accumulate in non-dividing tissues, we carried out transcriptome profiling of young vs aged adult fly heads. The data show that loss of H3.3K36 results in age-dependent misexpression of NF-{kappa}B and other innate immune target genes, as well as defects in silencing of heterochromatin. We propose H3.3K36 maintains the postmitotic epigenomic landscape, supporting longevity by regulating both pericentric and telomeric retrotransposons and by suppressing aberrant immune signaling.

genetics↗

bioRxiv2023

Structure of the portal complex from Staphylococcus aureus Pathogenicity Island 1 transducing particles in situ and in solution

Staphylococcus aureus is an important human pathogen, and the prevalence of antibiotic resistance is a major public health concern. The evolution of pathogenicity and resistance in S. aureus often involves acquisition of mobile genetic elements (MGEs). Bacteriophages play an especially important role, since transduction represents the main mechanism for horizontal gene transfer. S. aureus pathogenicity islands (SaPIs), including SaPI1, are MGEs that carry genes encoding virulence factors, and are mobilized at high frequency through interactions with specific "helper" bacteriophages, such as 80, leading to packaging of the SaPI genomes into virions made from structural proteins supplied by the helper. Among these structural proteins is the portal protein, which forms a ring-like portal at a fivefold vertex of the capsid, through which the DNA is packaged during virion assembly and ejected upon infection of the host. We have used high- resolution cryo-electron microscopy to determine structures of the S. aureus bacteriophage 80 portal in solution and in situ in the empty and full SaPI1 virions, and show how the portal interacts with the capsid. These structures provide a basis for understanding portal and capsid assembly and the conformational changes that occur upon DNA packaging and ejection.

microbiology↗

bioRxiv2023

Cell wall fluidization by mechano-endopeptidases sets up a turgor-mediated volumetric pacemaker

The cell wall is an essential cellular component of bacteria and the target of many antibiotics. However, how bacteria regulate the rate of cell wall biosynthesis as growth rates change remains unresolved. In E. coli, cell wall growth was thought to proceed independently from turgor pressure1, the osmotic pressure that the cytoplasm exerts on the cell wall. Here, we uncover a striking increase of turgor pressure with growth rate. Modulating turgor pressure and measuring cell wall biosynthesis, we find that turgor pressure is directly controls the rate of cell wall biosynthesis. The picture that emerges is that turgor pressure is largely generated by counterions of negatively charged cellular biomass. The increase in turgor pressure with growth rates results from more ribosomes and therefore higher concentrations of negatively charged ribosomal RNA. Elegantly, the coupling between biomass composition, turgor pressure and cell wall biosynthesis simultaneously explains how bacteria achieve homeostasis of cytoplasmic crowding and how they regulate the rate of cell wall biosynthesis across growth rates.

microbiology↗

bioRxiv2023

Plasticity of growth laws tunes resource allocation strategies in bacteria

Bacteria like E. coli grow at vastly different rates on different substrates, however, the precise reason for this variability is poorly understood. Different growth rates have been attributed to nutrient quality, a key parameter in bacterial growth laws. However, it remains unclear to what extent nutrient quality is rooted in fundamental biochemical constraints like the energy content of nutrients, the protein cost required for their uptake and catabolism, or the capacity of the plasma membrane for nutrient transporters. Here, we show that while nutrient quality is indeed reflected in protein investment in substrate-specific transporters and enzymes, this is not a fundamental limitation on growth rate. We show that it is possible to turn mannose, one of the poorest substrates of E. coli, into one of the best substrates by reengineering chromosomal promoters of the mannose transporter and metabolic enzymes required for mannose degradation. However, we show that this faster growth rate comes at the cost of diverse cellular capabilities, reflected in longer lag phases, worse starvation survival and lower motility. We show that addition of cAMP to the medium can rescue these phenotypes but imposes a corresponding growth cost. Based on these data, we propose that nutrient quality is largely a self-determined, plastic property that can be modulated by the fraction of proteomic resources devoted to a specific substrate in the much larger proteome sector of catabolically activated genes. Rather than a fundamental biochemical limitation, nutrient quality reflects resource allocation decisions that are shaped by evolution in specific ecological niches and can be quickly adapted if necessary. Author summaryBacteria grow at very different rates on different substrates. Therefore, the substrates themselves are often denoted as rich substrates versus poor substrates, referring to their nutrient quality. Nutrient quality is also a key parameter in bacterial growth laws that determines substrate-specific growth rate. However, it remains unclear which properties make a specific carbon source a good substrate or a bad substrate and a host of different explanations have been suggested, such as the energy content of the nutrient, the protein investment required for efficient catabolism of the substrate, or limitations in membrane capacity for transporters of the substrate, all of which are actively debated. Here, we show instead that nutrient quality can be a plastic property that can be dialed by evolution or by genetic perturbations. While we report a correlation between nutrient quality and protein cost of substrate utilization, this correlation is not causal. Instead, nutrient quality is encoded by the proteome fraction of a core catabolic sector within a much larger co-regulated catabolic sector. Different nutrients thus enable bacteria to grow, but they also serve as a major signal that allows microbes to infer information about their environment. We propose that nutrient quality encoded in a combination regulatory architecture and enzymatic properties, serves as both as a map of the safety and reliability of the environment and as a regulatory mechanism implementing proteome allocation decisions.

microbiology↗

bioRxiv2023

Long-term history dependence of growth rates of E. coli after nutrient shifts

According to a widely accepted paradigm of microbiology, steady-state growth rates are determined solely by current growth conditions1-3 and adaptations between growth states are rapid, as recently recapitulated by simple resource allocation models4. However, even in microbes overlapping regulatory networks can yield multi-stability or long-term cellular memory. Species like Listeria monocytogenes5 and Bacillus subtilis "distinguish" distinct histories for the commitment to sporulation6, but it is unclear if these states can persist over many generations. Remarkably, studying carbon co-utilization of Escherichia coli, we found that growth rates on combinations of carbon sources can depend critically on the previous growth condition. Growing in identical conditions, we observed differences in growth rates of up to 25% and we did not observe convergence of growth rates over 15 generations. We observed this phenomenon occurs across combinations of different phosphotransferase (PTS) substrates with various gluconeogenic carbon sources and found it to depend on the transcription factor Mlc.

microbiology↗

bioRxiv2023

Olfactomedin-4+ neutrophils exacerbate intestinal epithelial damage and worsen host survival after Clostridioides difficile infection.

Neutrophils are key first responders to Clostridioides difficile infection (CDI). Excessive tissue and blood neutrophils are associated with worse histopathology and adverse outcomes, however their functional role during CDI remains poorly defined. Utilizing intestinal epithelial cell (IEC)-neutrophil co-cultures and a pre-clinical animal model of CDI, we show that neutrophils exacerbate C. difficile-induced IEC injury. We utilized cutting-edge single-cell transcriptomics to illuminate neutrophil subtypes and biological pathways that could exacerbate CDI-associated IEC damage. As such, we have established the first transcriptomics atlas of bone marrow (BM), blood, and colonic neutrophils after CDI. We found that CDI altered the developmental trajectory of BM and blood neutrophils towards populations that exhibit gene signatures associated with pro-inflammatory responses and neutrophil-mediated tissue damage. Similarly, the transcriptomic signature of colonic neutrophils was consistent with hyper-inflammatory and highly differentiated cells that had amplified expression of cytokine-mediated signaling and degranulation priming genes. One of the top 10 variable features in colonic neutrophils was the gene for neutrophil glycoprotein, Olfactomedin 4 (OLFM4). CDI enhanced OLFM4 mRNA and protein expression in neutrophils, and OLFM4+ cells aggregated to areas of severe IEC damage. Compared to uninfected controls, both humans and mice with CDI had higher concentrations of circulating OLFM4; and in mice, OLFM4 deficiency resulted in faster recovery and better survival after infection. Collectively, these studies provide novel insights into neutrophil-mediated pathology after CDI and highlight the pathogenic role of OLFM4+ neutrophils in regulating CDI-induced IEC damage. One Sentence SummaryUtilizing single-cell transcriptomics, IEC-epithelial co-cultures, and pre-clinical models of CDI, we have identified a subset of neutrophils that are marked by OLFM4 expression as pathogenic determinants of IEC barrier damage after CDI.

immunology↗

bioRxiv2023

Combined effects of P25 TiO2 nanoparticles and disposable face mask leachate on microalgae Scenedesmus obliquus: Analysing the effects of heavy metals

Disposable surgical masks have been extensively employed as protective medical equipment due to the widespread breakout and transmission of the COVID-19 virus across the globe. These masks were made up of plastic polymer materials that would emit microplastics after entering the environment. Therefore, their careless disposal might lead to new and bigger microplastic contamination. The impacts of plastics that seep into waterways and their subsequent interactions with aquatic life are yet largely unexplored. In this study, we determined the quantity and kind of microplastics that were discharged from disposable surgical face masks. Furthermore, we also quantified heavy metals leached from the face masks (HML). In contrast, the increasing usage of nTiO2 in consumer items has led to its ubiquitous presence in freshwater systems. Four different concentrations of nTiO2, 0.5, 1, 2, and 4 mg L-1 were mixed with face mask leachates (FML) to perform the mixture toxicity test on freshwater algae, Scenedesmus obliquus. Reduced cell viability and photosynthetic activity were noticed in the treatment groups containing nTiO2 and FML. This was accompanied by increased oxidative stress and antioxidant activities. Furthermore, the heavy metals leached from the face masks were also tested for toxicity. In addition to that, changes in the cellular morphology were also studied with the help of FE-SEM and FTIR analysis. Our study reveals that leachates from disposable surgical face masks along with nTiO2 possess a serious threat to the environment. Environmental significanceDuring COVID-19, surgical face masks were widely used and discarded. These discarded face masks end up in lakes, rivers, and ponds. The facemasks were composed of polypropylene and other polymers. These masks release microplastics and heavy metals when discarded into water bodies. The current research focuses on assessing the environmental toxicity of the microplastics and heavy metals leached from the masks using algae as a model system. Our work further demonstrates the combined toxic effects of nTiO2 in the presence of the face mask leachate. Algae plays a crucial role as the primary producer in the freshwater ecosystem. These emerging contaminants may act as environmental stressors to the microalgae, and this may impair the ecosystems structure and function.

pharmacology and toxicology↗

bioRxiv2023

Molecular imaging with aquaporin-based reporter genes: quantitative considerations from Monte Carlo diffusion simulations

Aquaporins provide a new class of genetic tools for imaging molecular activity in deep tissues by increasing the rate of cellular water diffusion, which generates magnetic resonance contrast. However, distinguishing aquaporin contrast from the tissue background is challenging because water diffusion is also influenced by structural factors such as cell size and packing density. Here, we developed and experimentally validated a Monte Carlo model to analyze how cell radius and intracellular volume fraction quantitatively affect aquaporin signals. We demonstrated that a differential imaging approach based on time-dependent changes in diffusivity can improve specificity by unambiguously isolating aquaporin-driven contrast from the tissue background. Finally, we used Monte Carlo simulations to analyze the connection between diffusivity and the percentage of cells engineered to express aquaporin, and established a simple mapping that accurately determined the volume fraction of aquaporin-expressing cells in mixed populations. This study creates a framework for broad applications of aquaporins, particularly in biomedicine and in vivo synthetic biology, where quantitative methods to measure the location and performance of genetic devices in whole vertebrates are necessary.

bioengineering↗

bioRxiv2023

E3 ubiquitin ligase Deltex facilitates the expansion of Wingless gradient and antagonizes Wingless signaling through a conserved mechanism of transcriptional effector Armadillo/β-catenin degradation

The Wnt/Wg pathway controls myriads of biological phenomena throughout the development and adult life of all organisms across the phyla. Thus, an aberrant Wnt signaling is associated with a wide range of pathologies in humans. Tight regulation of Wnt/Wg signaling is required to maintain proper cellular homeostasis. Here we report a novel role of E3 ubiquitin ligase Deltex in Wg signaling regulation. Drosophila dx genetically interacts with wg and its pathway components. Further, Dx LOF results in a reduced spreading of Wg while its over-expression expands the diffusion gradient of the morphogen. We attribute this change in Wg gradient to the endocytosis of Wg through Dx which directly affects the short and long-range Wg targets. We also demonstrate the role of Dx in regulating Wg effector Armadillo where Dx down-regulates Arm through proteasomal degradation. We also showed the conservation of Dx function in the mammalian system where DTX1 is shown to bind with {beta}-catenin and facilitates its proteolytic degradation, spotlighting a novel step that potentially modulates Wnt/Wg signaling cascade.

developmental biology↗

bioRxiv2023

A semiconductor 96-microplate platform for real-time impedance-based high-throughput screening

Profiling compounds and genetic perturbations via high-content imaging has become increasingly popular for drug discovery, but the technique is limited to endpoint images of fixed cells. In contrast, electronic-based devices offer label-free, functional information of live cells, yet current approaches suffer from low-spatial resolution or single-well throughput. Here, we report a semiconductor 96-microplate platform designed for high-resolution real-time impedance "imaging" at scale. Each well features 4,096 electrodes at 25 {micro}m spatial resolution while a miniaturized data interface allows 8x parallel plate operation (768 total wells) within each incubator for enhanced throughputs. New electric field-based, multi-frequency measurement techniques capture >20 parameter images including tissue barrier, cell-surface attachment, cell flatness, and motility every 15 min throughout experiments. Using these real-time readouts, we characterized 16 cell types, ranging from primary epithelial to suspension, and quantified heterogeneity in mixed epithelial and mesenchymal co-cultures. A proof-of-concept screen of 904 diverse compounds using 13 semiconductor microplates demonstrates the platforms capability for mechanism of action (MOA) profiling with 25 distinct responses identified. The scalability of the semiconductor platform combined with the translatability of the high dimensional live-cell functional parameters expands high-throughput MOA profiling and phenotypic drug discovery applications.

bioengineering↗

bioRxiv2023

Engineering ligand stabilized aquaporin reporters for magnetic resonance imaging

Imaging transgene expression in live tissues requires reporters that are detectable with deeply penetrant modalities, such as magnetic resonance imaging (MRI). Here, we show that LSAqp1, a water channel engineered from aquaporin-1, can be used to create background-free, drug-gated, and multiplex images of gene expression using MRI. LSAqp1 is a fusion protein composed of aquaporin-1 and a degradation tag that is sensitive to a cell-permeable ligand, which allows for dynamic small molecule modulation of MRI signals. LSAqp1 improves specificity for imaging gene expression by allowing reporter signals to be conditionally activated and distinguished from the tissue background by difference imaging. In addition, by engineering destabilized aquaporin-1 variants with different ligand requirements, it is possible to image distinct cell types simultaneously. Finally, we expressed LSAqp1 in a tumor model and showed successful in vivo imaging of gene expression without background activity. LSAqp1 provides a conceptually unique approach to accurately measure gene expression in living organisms by combining the physics of water diffusion and biotechnology tools to control protein stability.

bioengineering↗

bioRxiv2023

Inverse blebs operate as hydraulic pumps during mouse blastocyst formation

During preimplantation development, mouse embryos form a fluid-filled lumen, which sets their first axis of symmetry1,2. Pressurized fluid breaks open cell-cell contacts and accumulates into pockets, which gradually coarsen into a single lumen3-5. During coarsening, the adhesive and contractile properties of cells are thought to guide intercellular fluid (IF) but what cell behavior may control fluid movements is unknown. Here, we report large fluid-filled spherical membrane intrusions called inverse blebs6,7 growing into cells at adhesive contacts. At the onset of lumen coarsening, we observed hundreds of inverse blebs throughout the embryo, each dynamically filling with IF and retracting within a minute. We find that inverse blebs grow due to pressure build-up resulting from luminal fluid accumulation and cell-cell adhesion, which locally confines fluid. Inverse blebs then retract due to actomyosin contraction, which effectively redistributes fluid within the intercellular space. Importantly, inverse blebs show topological specificity and only occur at contacts between two cells, not at contacts formed by multiple cells, which essentially serve as fluid sinks. Manipulating the topology of the embryo reveals that, in the absence of sinks, inverse blebs pump fluid into one another in a futile cycle. We propose that inverse blebs operate as hydraulic pumps to promote luminal coarsening, thereby constituting an instrument used by cells to control fluid movement.

developmental biology↗

bioRxiv2023

Predictable genetic recruitment for luciferin sulfation in the convergent evolution of bioluminescence

Genes from ancient families are sometimes involved in the convergent evolutionary origins of similar traits, even across vast phylogenetic distances. Sulfotransferases are an ancient family of enzymes that transfer sulfate from a donor to a wide variety of substrates, including probable roles in some bioluminescence systems. Here we demonstrate multiple sulfotransferases, highly expressed in light organs of the bioluminescent ostracod Vargula tsujii, transfer sulfate in vivo to the luciferin substrate, vargulin. We find luciferin sulfotransferases of ostracods are not orthologous to known luciferin sulfotransferases of fireflies or sea pansies; animals with distinct and convergently evolved bioluminescence systems compared to ostracods. Therefore, distantly related sulfotransferases were independently recruited at least three times, leading to parallel evolution of luciferin metabolism in three highly diverged organisms. Re-use of homologous genes is surprising in these bioluminescence systems because the other components, including luciferins and luciferases, are completely distinct. Whether convergently evolved traits incorporate ancient genes with similar functions or instead use distinct, often newer, genes may be constrained by how many genetic solutions exist for a particular function. When fewer solutions exist, as in genetic sulfation of small molecules, evolution may be more constrained to use the same genes time and again.

evolutionary biology↗