Ediacaran marine animal forests and the ventilation of the oceans.

The rise of animals across the Ediacaran-Cambrian transition marked a step-change in the history of life, from a microbially dominated world to the complex macroscopic biosphere we see today.1,2,3 While the importance of bioturbation and swimming in altering the structure and function of Earth systems is well established,4,5,6 the influence of epifaunal animals on the hydrodynamics of marine environments is not well understood. Of particular interest are the oldest "marine animal forests,"7 which comprise a diversity of sessile soft-bodied organisms dominated by the fractally branching rangeomorphs.8,9 Typified by fossil assemblages from the Ediacaran of Mistaken Point, Newfoundland,8,10,11 these ancient communities might have played a pivotal role in structuring marine environments, similar to modern ecosystems,7,12,13 but our understanding of how they impacted fluid flow in the water column is limited. Here, we use ecological modeling and computational flow simulations to explore how Ediacaran marine animal forests influenced their surrounding environment. Our results reveal how organism morphology and community structure and composition combined to impact vertical mixing of the surrounding water. We find that Mistaken Point communities were capable of generating high-mixing conditions, thereby likely promoting gas and nutrient transport within the "canopy." This mixing could have served to enhance local-scale oxygen concentrations and redistribute resources like dissolved organic carbon. Our work suggests that Ediacaran marine animal forests may have contributed to the ventilation of the oceans over 560 million years ago, well before the Cambrian explosion of animals.

Raptorial appendages of the Cambrian apex predator Anomalocaris canadensis are built for soft prey and speed.

The stem-group euarthropod Anomalocaris canadensis is one of the largest Cambrian animals and is often considered the quintessential apex predator of its time. This radiodont is commonly interpreted as a demersal hunter, responsible for inflicting injuries seen in benthic trilobites. However, controversy surrounds the ability of A. canadensis to use its spinose frontal appendages to masticate or even manipulate biomineralized prey. Here, we apply a new integrative computational approach, combining three-dimensional digital modelling, kinematics, finite-element analysis (FEA) and computational fluid dynamics (CFD) to rigorously analyse an A. canadensis feeding appendage and test its morphofunctional limits. These models corroborate a raptorial function, but expose inconsistencies with a capacity for durophagy. In particular, FEA results show that certain parts of the appendage would have experienced high degrees of plastic deformation, especially at the endites, the points of impact with prey. The CFD results demonstrate that outstretched appendages produced low drag and hence represented the optimal orientation for speed, permitting acceleration bursts to capture prey. These data, when combined with evidence regarding the functional morphology of its oral cone, eyes, body flaps and tail fan, suggest that A. canadensis was an agile nektonic predator that fed on soft-bodied animals swimming in a well-lit water column above the benthos. The lifestyle of A. canadensis and that of other radiodonts, including plausible durophages, suggests that niche partitioning across this clade influenced the dynamics of Cambrian food webs, impacting on a diverse array of organisms at different sizes, tiers and trophic levels.

The rangeomorph Pectinifrons abyssalis: Hydrodynamic function at the dawn of animal life.

Rangeomorphs are among the oldest putative eumetazoans known from the fossil record. Establishing how they fed is thus key to understanding the structure and function of the earliest animal ecosystems. Here, we use computational fluid dynamics to test hypothesized feeding modes for the fence-like rangeomorph Pectinifrons abyssalis, comparing this to the morphologically similar extant carnivorous sponge Chondrocladia lyra. Our results reveal complex patterns of flow around P. abyssalis unlike those previously reconstructed for any other Ediacaran taxon. Comparisons with C. lyra reveal substantial differences between the two organisms, suggesting they converged on a similar fence-like morphology for different functions. We argue that the flow patterns recovered for P. abyssalis do not support either a suspension feeding or osmotrophic feeding habit. Instead, our results indicate that rangeomorph fronds may represent organs adapted for gas exchange. If correct, this interpretation could require a dramatic reinterpretation of the oldest macroscopic animals.

Large size in aquatic tetrapods compensates for high drag caused by extreme body proportions.

Various Mesozoic marine reptile lineages evolved streamlined bodies and efficient lift-based swimming, as seen in modern aquatic mammals. Ichthyosaurs had low-drag bodies, akin to modern dolphins, but plesiosaurs were strikingly different, with long hydrofoil-like limbs and greatly variable neck and trunk proportions. Using computational fluid dynamics, we explore the effect of this extreme morphological variation. We find that, independently of their body fineness ratio, plesiosaurs produced more drag than ichthyosaurs and modern cetaceans of equal mass due to their large limbs, but these differences were not significant when body size was accounted for. Additionally, necks longer than twice the trunk length can substantially increase the cost of forward swimming, but this effect was cancelled out by the evolution of big trunks. Moreover, fast rates in the evolution of neck proportions in the long-necked elasmosaurs suggest that large trunks might have released the hydrodynamic constraints on necks thus allowing their extreme enlargement.

The locomotion of extinct secondarily aquatic tetrapods.

The colonisation of freshwater and marine ecosystems by land vertebrates has repeatedly occurred in amphibians, reptiles, birds and mammals over the course of 300 million years. Functional interpretations of the fossil record are crucial to understanding the forces shaping these evolutionary transitions. Secondarily aquatic tetrapods have acquired a suite of anatomical, physiological and behavioural adaptations to locomotion in water. However, much of this information is lost for extinct clades, with fossil evidence often restricted to osteological data and a few extraordinary specimens with soft tissue preservation. Traditionally, functional morphology in fossil secondarily aquatic tetrapods was investigated through comparative anatomy and correlation with living functional analogues. However, in the last two decades, biomechanics in palaeobiology has experienced a remarkable methodological shift. Anatomy-based approaches are increasingly rigorous, informed by quantitative techniques for analysing shape. Moreover, the incorporation of physics-based methods has enabled objective tests of functional hypotheses, revealing the importance of hydrodynamic forces as drivers of evolutionary innovation and adaptation. Here, we present an overview of the latest research on the locomotion of extinct secondarily aquatic tetrapods, with a focus on amniotes, highlighting the state-of-the-art experimental approaches used in this field. We discuss the suitability of these techniques for exploring different aspects of locomotory adaptation, analysing their advantages and limitations and laying out recommendations for their application, with the aim to inform future experimental strategies. Furthermore, we outline some unexplored research avenues that have been successfully deployed in other areas of palaeobiomechanical research, such as the use of dynamic models in feeding mechanics and terrestrial locomotion, thus providing a new methodological synthesis for the field of locomotory biomechanics in extinct secondarily aquatic vertebrates. Advances in imaging technology and three-dimensional modelling software, new developments in robotics, and increased availability and awareness of numerical methods like computational fluid dynamics make this an exciting time for analysing form and function in ancient vertebrates.

Effects of body plan evolution on the hydrodynamic drag and energy requirements of swimming in ichthyosaurs.

Ichthyosaurs are an extinct group of fully marine tetrapods that were well adapted to aquatic locomotion. During their approximately 160 Myr existence, they evolved from elongate and serpentine forms into stockier, fish-like animals, convergent with sharks and dolphins. Here, we use computational fluid dynamics (CFD) to quantify the impact of this transition on the energy demands of ichthyosaur swimming for the first time. We run computational simulations of water flow using three-dimensional digital models of nine ichthyosaurs and an extant functional analogue, a bottlenose dolphin, providing the first quantitative evaluation of ichthyosaur hydrodynamics across phylogeny. Our results show that morphology did not have a major effect on the drag coefficient or the energy cost of steady swimming through geological time. We show that even the early ichthyosaurs produced low levels of drag for a given volume, comparable to those of a modern dolphin, and that deep 'torpedo-shaped' bodies did not reduce the cost of locomotion. Our analysis also provides important insight into the choice of scaling parameters for CFD applied to swimming mechanics, and underlines the great influence of body size evolution on ichthyosaur locomotion. A combination of large bodies and efficient swimming modes lowered the cost of steady swimming as ichthyosaurs became increasingly adapted to a pelagic existence.

Autophagy maintains stemness by preventing senescence.

During ageing, muscle stem-cell regenerative function declines. At advanced geriatric age, this decline is maximal owing to transition from a normal quiescence into an irreversible senescence state. How satellite cells maintain quiescence and avoid senescence until advanced age remains unknown. Here we report that basal autophagy is essential to maintain the stem-cell quiescent state in mice. Failure of autophagy in physiologically aged satellite cells or genetic impairment of autophagy in young cells causes entry into senescence by loss of proteostasis, increased mitochondrial dysfunction and oxidative stress, resulting in a decline in the function and number of satellite cells. Re-establishment of autophagy reverses senescence and restores regenerative functions in geriatric satellite cells. As autophagy also declines in human geriatric satellite cells, our findings reveal autophagy to be a decisive stem-cell-fate regulator, with implications for fostering muscle regeneration in sarcopenia.

Bmi1 regulates murine intestinal stem cell proliferation and self-renewal downstream of Notch.

Genetic data indicate that abrogation of Notch-Rbpj or Wnt-ß-catenin pathways results in the loss of the intestinal stem cells (ISCs). However, whether the effect of Notch is direct or due to the aberrant differentiation of the transit-amplifying cells into post-mitotic goblet cells is unknown. To address this issue, we have generated composite tamoxifen-inducible intestine-specific genetic mouse models and analyzed the expression of intestinal differentiation markers. Importantly, we found that activation of ß-catenin partially rescues the differentiation phenotype of Rbpj deletion mutants, but not the loss of the ISC compartment. Moreover, we identified Bmi1, which is expressed in the ISC and progenitor compartments, as a gene that is co-regulated by Notch and ß-catenin. Loss of Bmi1 resulted in reduced proliferation in the ISC compartment accompanied by p16(INK4a) and p19(ARF) (splice variants of Cdkn2a) accumulation, and increased differentiation to the post-mitotic goblet cell lineage that partially mimics Notch loss-of-function defects. Finally, we provide evidence that Bmi1 contributes to ISC self-renewal.

Geriatric muscle stem cells switch reversible quiescence into senescence.

Regeneration of skeletal muscle depends on a population of adult stem cells (satellite cells) that remain quiescent throughout life. Satellite cell regenerative functions decline with ageing. Here we report that geriatric satellite cells are incapable of maintaining their normal quiescent state in muscle homeostatic conditions, and that this irreversibly affects their intrinsic regenerative and self-renewal capacities. In geriatric mice, resting satellite cells lose reversible quiescence by switching to an irreversible pre-senescence state, caused by derepression of p16(INK4a) (also called Cdkn2a). On injury, these cells fail to activate and expand, undergoing accelerated entry into a full senescence state (geroconversion), even in a youthful environment. p16(INK4a) silencing in geriatric satellite cells restores quiescence and muscle regenerative functions. Our results demonstrate that maintenance of quiescence in adult life depends on the active repression of senescence pathways. As p16(INK4a) is dysregulated in human geriatric satellite cells, these findings provide the basis for stem-cell rejuvenation in sarcopenic muscles.

PAI-1-regulated miR-21 defines a novel age-associated fibrogenic pathway in muscular dystrophy.

Disruption of skeletal muscle homeostasis by substitution with fibrotic tissue constitutes the principal cause of death in Duchenne muscular dystrophy (DMD) patients, yet the implicated fibrogenic mechanisms remain poorly understood. This study identifies the extracellular PAI-1/urokinase-type plasminogen activator (uPA) balance as an important regulator of microribonucleic acid (miR)-21 biogenesis, controlling age-associated muscle fibrosis and dystrophy progression. Genetic loss of PAI-1 in mdx dystrophic mice anticipated muscle fibrosis through these sequential mechanisms: the alteration of collagen metabolism by uPA-mediated proteolytic processing of transforming growth factor (TGF)-ß in muscle fibroblasts and the activation of miR-21 expression, which inhibited phosphatase and tensin homologue and enhanced AKT signaling, thus endowing TGF-ß with a remarkable cell proliferation-promoting potential. Age-associated fibrogenesis and muscle deterioration in mdx mice, as well as exacerbated dystrophy in young PAI-1(-/-) mdx mice, could be reversed by miR-21 or uPA-selective interference, whereas forced miR-21 overexpression aggravated disease severity. The PAI-1-miR-21 fibrogenic axis also appeared dysregulated in muscle of DMD patients, providing a basis for effectively targeting fibrosis and muscular dystrophies in currently untreatable individuals.

Functional cooperation between Snail1 and twist in the regulation of ZEB1 expression during epithelial to mesenchymal transition.

Snail1 and Zeb1 are E-cadherin-transcriptional repressors induced during epithelial mesenchymal transition (EMT). In this article we have analyzed the factors controlling Zeb1 expression during EMT. In NMuMG cells treated with TGF-ß, Snail1 RNA and protein are induced 1 h after addition of the cytokine preceding Zeb1 up-regulation that requires 6-8 h. Zeb1 gene expression is caused by increased RNA levels but also by enhanced protein stability and is markedly dependent on Snail1 because depletion of this protein prevents Zeb1 protein and RNA up-regulation. In addition to Snail1, depletion of the Twist transcriptional factor retards Zeb1 stimulation by TGF-ß or decreases Zeb1 expression in other cellular models indicating that this factor is also required for Zeb1 expression. Accordingly, Snail1 and Twist cooperate in the induction of Zeb1: co-transfection of both cDNAs is required for the maximal expression of ZEB1 mRNA. Unexpectedly, the expression of Snail1 and Twist shows a mutual dependence although to a different extent; whereas Twist depletion retards Snail1 up-regulation by TGF-ß, Snail1 is necessary for the rapid increase in Twist protein and later up-regulation of Twist1 mRNA induced by the cytokine. Besides this effect on Twist, Snail1 also induces the nuclear translocation of Ets1, another factor required for Zeb1 expression. Both Twist and Ets1 bind to the ZEB1 promoter although to different elements: whereas Ets1 interacts with the proximal promoter, Twist does it with a 700-bp sequence upstream of the transcription start site. These results indicate that Snail1 controls Zeb1 expression at multiple levels and acts cooperatively with Twist in the ZEB1 gene transcription induction.