Form, function, mind: What doesn't compute (and what might).

The applicability of computational and dynamical systems models to organisms is scrutinized, using examples from developmental biology and cognition. Developmental morphogenesis is dependent on the inherent material properties of developing animal (metazoan) tissues, a non-computational modality, but cell differentiation, which utilizes chromatin-based revisable memory banks and program-like function-calling, via the developmental gene co-expression system unique to the metazoans, has a quasi-computational basis. Multi-attractor dynamical models are argued to be misapplied to global properties of development, and it is suggested that along with computationalism, classic forms of dynamicism are similarly unsuitable to accounting for cognitive phenomena. Proposals are made for treating brains and other nervous tissues as novel forms of excitable matter with inherent properties which enable the intensification of cell-based basal cognition capabilities present throughout the tree of life. Finally, some connections are drawn between the viewpoint described here and active inference models of cognition, such as the Free Energy Principle.

The many roads to and from multicellularity.

The multiple origins of multicellularity had far-reaching consequences ranging from the appearance of phenotypically complex life-forms to their effects on Earth's aquatic and terrestrial ecosystems. Yet, many important questions remain. For example, do all lineages and clades share an ancestral developmental predisposition for multicellularity emerging from genomic and biophysical motifs shared from a last common ancestor, or are the multiple origins of multicellularity truly independent evolutionary events? In this review, we highlight recent developments and pitfalls in understanding the evolution of multicellularity with an emphasis on plants (here defined broadly to include the polyphyletic algae), but also draw upon insights from animals and their holozoan relatives, fungi and amoebozoans. Based on our review, we conclude that the evolution of multicellular organisms requires three phases (origination by disparate cell-cell attachment modalities, followed by integration by lineage-specific physiological mechanisms, and autonomization by natural selection) that have been achieved differently in different lineages.

Cell differentiation: What have we learned in 50 years?

I revisit two theories of cell differentiation in multicellular organisms published a half-century ago, Stuart Kauffman's global genome regulatory dynamics (GGRD) model and Roy Britten's and Eric Davidson's modular gene regulatory network (MGRN) model, in light of newer knowledge of mechanisms of gene regulation in the metazoans (animals). The two models continue to inform hypotheses and computational studies of differentiation of lineage-adjacent cell types. However, their shared notion (based on bacterial regulatory systems) of gene switches and networks built from them have constrained progress in understanding the dynamics and evolution of differentiation. Recent work has described unique write-read-rewrite chromatin-based expression encoding in eukaryotes, as well metazoan-specific processes of gene activation and silencing in condensed-phase, enhancer-recruiting regulatory hubs, employing disordered proteins, including transcription factors, with context-dependent identities. These findings suggest an evolutionary scenario in which the origination of differentiation in animals, rather than depending exclusively on adaptive natural selection, emerged as a consequence of a type of multicellularity in which the novel metazoan gene regulatory apparatus was readily mobilized to amplify and exaggerate inherent cell functions of unicellular ancestors. The plausibility of this hypothesis is illustrated by the evolution of the developmental role of Grainyhead-like in the formation of epithelium.

Does resource availability help determine the evolutionary route to multicellularity?

Genetic heterogeneity and homogeneity are associated with distinct sets of adaptive advantages and bottlenecks, both in developmental biology and population genetics. Whereas populations of individuals are usually genetically heterogeneous, most multicellular metazoans are genetically homogeneous. Observing that resource scarcity fuels genetic heterogeneity in populations, we propose that monoclonal development is compatible with the resource-rich and stable internal environments that complex multicellular bodies offer. In turn, polyclonal development persists in tumors and in certain metazoans, both exhibiting a closer dependence on external resources. This eco-evo-devo approach also suggests that multicellularity may originally have emerged through polyclonal development in early metazoans, because of their reduced shielding from environmental fluctuations.

Inherent forms and the evolution of evolution.

John Bonner presented a provocative conjecture that the means by which organisms evolve has itself evolved. The elements of his postulated nonuniformitarianism in the essay under discussion-the emergence of sex, the enhanced selection pressures on larger multicellular forms-center on a presumed close mapping of genotypic to phenotypic change. A different view emerges from delving into earlier work of Bonner's in which he proposed the concept of "neutral phenotypes" and "neutral morphologies" allied to D'Arcy Thompson's analysis of physical determinants of form and studied the conditional elicitation of intrinsic organizational properties of cell aggregates in social amoebae. By comparing the shared and disparate mechanistic bases of morphogenesis and developmental outcomes in the embryos of metazoans (animals), closely related nonmetazoan holozoans, more distantly related dictyostelids, and very distantly related volvocine algae, I conclude, in agreement with Bonner's earlier proposals, that understanding the evolution of multicellular evolution requires knowledge of the inherent forms of diversifying lineages, and that the relevant causative factors extend beyond genes and adaptation to the physics of materials.

John Tyler Bonner: Remembering a scientific pioneer.

Throughout his life, John Tyler Bonner contributed to major transformations in the fields of developmental and evolutionary biology. He pondered the evolution of complexity and the significance of randomness in evolution, and was instrumental in the formation of evolutionary developmental biology. His contributions were vast, ranging from highly technical scientific articles to numerous books written for a broad audience. This historical vignette gathers reflections by several prominent researchers on the greatness of John Bonner and the implications of his work.

Sleeper cells: the stringent response and persistence in the Borreliella (Borrelia) burgdorferi enzootic cycle.

Infections with tick-transmitted Borreliella (Borrelia) burgdorferi, the cause of Lyme disease, represent an increasingly large public health problem in North America and Europe. The ability of these spirochetes to maintain themselves for extended periods of time in their tick vectors and vertebrate reservoirs is crucial for continuance of the enzootic cycle as well as for the increasing exposure of humans to them. The stringent response mediated by the alarmone (p)ppGpp has been determined to be a master regulator in B. burgdorferi. It modulates the expression of identified and unidentified open reading frames needed to deal with and overcome the many nutritional stresses and other challenges faced by the spirochete in ticks and animal reservoirs. The metabolic and morphologic changes resulting from activation of the stringent response in B. burgdorferi may also be involved in the recently described non-genetic phenotypic phenomenon of tolerance to otherwise lethal doses of antimicrobials and to other antimicrobial activities. It may thus constitute a linchpin in multiple aspects of infections with Lyme disease borrelia, providing a link between the micro-ecological challenges of its enzootic life-cycle and long-term residence in the tissues of its animal reservoirs, with the evolutionary side effect of potential persistence in incidental human hosts.

On the application of principal component analysis to the calculation of the bulk integral optical properties for radiation parameterizations in climate models.

Rigorous electromagnetic computations required for the calculation of high-resolution monochromatic bulk integral optical properties of irregular atmospheric particles are onerous in memory and in time requirements. Here, it is shown that from a set of 145 monochromatic bulk integral ice optical properties, it is possible to reduce the set to eight hinge wavelengths by using the method of principal component analysis (PCA) regression. From the eight hinge wavelengths, the full set can be reconstructed to within root mean square errors of ≪1%. To obtain optimal reconstruction, the training set must cover as wide a range of parameter space as possible. Rigorous electromagnetic methods can now be routinely applied to represent accurately the integral optical properties of atmospheric particles in climate models.

Highly polygenic architecture of antidepressant treatment response: Comparative analysis of SSRI and NRI treatment in an animal model of depression.

Response to antidepressant (AD) treatment may be a more polygenic trait than previously hypothesized, with many genetic variants interacting in yet unclear ways. In this study we used methods that can automatically learn to detect patterns of statistical regularity from a sparsely distributed signal across hippocampal transcriptome measurements in a large-scale animal pharmacogenomic study to uncover genomic variations associated with AD. The study used four inbred mouse strains of both sexes, two drug treatments, and a control group (escitalopram, nortriptyline, and saline). Multi-class and binary classification using Machine Learning (ML) and regularization algorithms using iterative and univariate feature selection methods, including InfoGain, mRMR, ANOVA, and Chi Square, were used to uncover genomic markers associated with AD response. Relevant genes were selected based on Jaccard distance and carried forward for gene-network analysis. Linear association methods uncovered only one gene associated with drug treatment response. The implementation of ML algorithms, together with feature reduction methods, revealed a set of 204 genes associated with SSRI and 241 genes associated with NRI response. Although only 10% of genes overlapped across the two drugs, network analysis shows that both drugs modulated the CREB pathway, through different molecular mechanisms. Through careful implementation and optimisations, the algorithms detected a weak signal used to predict whether an animal was treated with nortriptyline (77%) or escitalopram (67%) on an independent testing set. The results from this study indicate that the molecular signature of AD treatment may include a much broader range of genomic markers than previously hypothesized, suggesting that response to medication may be as complex as the pathology. The search for biomarkers of antidepressant treatment response could therefore consider a higher number of genetic markers and their interactions. Through predominately different molecular targets and mechanisms of action, the two drugs modulate the same Creb1 pathway which plays a key role in neurotrophic responses and in inflammatory processes. © 2016 The Authors. American Journal of Medical Genetics Part B: Neuropsychiatric Genetics Published by Wiley Periodicals, Inc.

Deep phylogenomics of a tandem-repeat galectin regulating appendicular skeletal pattern formation.

BACKGROUND: A multiscale network of two galectins Galectin-1 (Gal-1) and Galectin-8 (Gal-8) patterns the avian limb skeleton. Among vertebrates with paired appendages, chondrichthyan fins typically have one or more cartilage plates and many repeating parallel endoskeletal elements, actinopterygian fins have more varied patterns of nodules, bars and plates, while tetrapod limbs exhibit tandem arrays of few, proximodistally increasing numbers of elements. We applied a comparative genomic and protein evolution approach to understand the origin of the galectin patterning network. Having previously observed a phylogenetic constraint on Gal-1 structure across vertebrates, we asked whether evolutionary changes of Gal-8 could have critically contributed to the origin of the tetrapod pattern. RESULTS: Translocations, duplications, and losses of Gal-8 genes in Actinopterygii established them in different genomic locations from those that the Sarcopterygii (including the tetrapods) share with chondrichthyans. The sarcopterygian Gal-8 genes acquired a potentially regulatory non-coding motif and underwent purifying selection. The actinopterygian Gal-8 genes, in contrast, did not acquire the non-coding motif and underwent positive selection. CONCLUSION: These observations interpreted through the lens of a reaction-diffusion-adhesion model based on avian experimental findings can account for the distinct endoskeletal patterns of cartilaginous, ray-finned, and lobe-finned fishes, and the stereotypical limb skeletons of tetrapods.

Blastocyst genotyping for quality control of mouse mutant archives: an ethical and economical approach.

With the advent of modern developmental biology and molecular genetics, the scientific community has generated thousands of newly genetically altered strains of laboratory mice with the aim of elucidating gene function. To this end, a large group of Institutions which form the International Mouse Phenotyping Consortium is generating and phenotyping a knockout mouse strain for each of the ~20,000 protein-coding genes using the mutant ES cell resource produced by the International Knockout Mouse Consortium. These strains are made available to the research community via public repositories, mostly as cryopreserved sperm or embryos. To ensure the quality of this frozen resource there is a requirement that for each strain the frozen sperm/embryos are proven able to produce viable mutant progeny, before the live animal resource is removed from cages. Given the current requirement to generate live pups to demonstrate their mutant genotype, this quality control check necessitates the use and generation of many animals and requires considerable time, cage space, technical and economic resources. Here, we describe a simple and efficient method of genotyping pre-implantation stage blastocysts with significant ethical and economic advantages especially beneficial for current and future large-scale mouse mutagenesis projects.

Form and function remixed: developmental physiology in the evolution of vertebrate body plans.

The most widely accepted model of evolutionary change, the Modern Evolutionary Synthesis, is based on the gradualism of Darwin and Wallace. They, in turn, developed their ideas in the context of 19th century concepts of how matter, including the tissues of animals and plants, could be reshaped and repatterned. A new physics of condensed, chemically, electrically and mechanically excitable materials formulated in the 20th century was, however, readily taken up by physiologists, who applied it to the understanding of dynamical, external condition-dependent and homeostatic properties of individual organisms. Nerve conduction, vascular and airway dynamics, and propagation of electrical excitations in heart and brain tissue all benefited from theories of biochemical oscillation, fluid dynamics, reaction-diffusion-based pattern instability and allied dissipative processes. When, in the late 20th century, the development of body and organ form was increasingly seen to involve dynamical, frequently non-linear processes similar to those that had become standard in physiology, a strong challenge to the evolutionary synthesis emerged. In particular, large-scale changes in organismal form now had a scientific basis other than gradualistic natural selection based on adaptive advantage. Moreover, heritable morphological changes were seen to be capable of occurring abruptly with little or no genetic change, with involvement of the external environment, and in preferred directions. This paper discusses three examples of morphological motifs of vertebrate bodies and organs, the somites, the skeletons of the paired limbs, and musculoskeletal novelties distinctive to birds, for which evolutionary origination and transformation can be understood on the basis of the physiological and biophysical determinants of their development.

Why are there eggs?

A description and update of the "egg-as-novelty" hypothesis is presented. It is proposed that the major animal phylum-characteristic suites of morphological motifs first emerged more than a half-billion years ago in multicellular aggregates and clusters that did not exhibit an egg-soma divergence. These pre-metazoan bodies were organized by "dynamical patterning modules" (DPMs), physical processes and effects mobilized on the new multicellular scale by ancient conserved genes that came to mediate cell-cell interactions in these clusters. "Proto-eggs" were enlarged cells that through cleavage, or physical confinement by a secreted matrix, served to enforce genomic and genetic homogeneity in the cell clusters arising from them. Enlargement of the founder cell was the occasion for spontaneous intra-egg spatiotemporal organization based on single-cell physiological functions - calcium transients and oscillations, cytoplasmic flows - operating on the larger scale. Ooplasmic segregation by egg-patterning processes, while therefore not due to adaptive responses to external challenges, served as evolutionarily fertile "pre-adaptations" by making the implementation of the later-acting (at the multicellular "morphogenetic stage" of embryogenesis) DPMs more reliable, robust, and defining of sub-phylum morphotypes. This perspective is seen to account for a number of otherwise difficult to understand features of the evolution of development, such as the rapid diversification of biological forms with a conserved genetic toolkit at the dawn of animal evolution, the capability of even obligatory sexual reproducers to propagate vegetatively, and the "embryonic hourglass" of comparative developmental biology.

Ernest Everett Just: Egg and embryo as excitable systems.

Ernest Everett Just (1883-1941) was an African American embryologist of international standing whose research interests lay in the area of fertilization and early development in marine invertebrates. Perhaps best known for his discovery of the dynamical and structural blocks to polyspermy that sweep over the egg upon fertilization, E. E. Just also was the first to associate cell surface changes with stages of embryonic development. He was deeply familiar with the natural history of the animals whose eggs he studied, and his knowledge of natural settings led him to emphasize the importance of using laboratory conditions that closely match those in nature. Based on more than 30 years of work, he came to believe that it was the cell surface that played the most critical role in development, heredity, and evolution. He promoted a holistic view of cells and organisms in opposition to the gene-centric view that was becoming more prevalent with the rise of genetics, but rejected the vitalism espoused by some biologists of his era, calling instead for "a physics and chemistry in a new dimension …superimposed upon the now known physics and chemistry" to account for biological phenomena. Just's incisive critique of genetic reductionism finds echoes in contemporary multiscale, systems approaches in biology. His speculations on the relationship between developmental and evolutionary mechanisms resonate with today's evolutionary developmental biology. After a brief biographical sketch, this paper outlines and discusses some of Just's scientific contributions, and shows how his ideas remain relevant today.

Rapid conversion of EUCOMM/KOMP-CSD alleles in mouse embryos using a cell-permeable Cre recombinase.

We describe here use of a cell-permeable Cre to efficiently convert the EUCOMM/KOMP-CSD tm1a allele to the tm1b form in preimplantation mouse embryos in a high-throughput manner, consistent with the requirements of the International Mouse Phenotyping Consortium-affiliated NIH KOMP2 project. This method results in rapid allele conversion and minimizes the use of experimental animals when compared to conventional Cre transgenic mouse breeding, resulting in a significant reduction in costs and time with increased welfare benefits.

The origins of multicellular organisms.

Multicellularity has evolved in several eukaryotic lineages leading to plants, fungi, and animals. Theoretically, in each case, this involved (1) cell-to-cell adhesion with an alignment-of-fitness among cells, (2) cell-to-cell communication, cooperation, and specialization with an export-of-fitness to a multicellular organism, and (3) in some cases, a transition from "simple" to "complex" multicellularity. When mapped onto a matrix of morphologies based on developmental and physical rules for plants, these three phases help to identify a "unicellular ⇒ colonial ⇒ filamentous (unbranched ⇒ branched) ⇒ pseudoparenchymatous ⇒ parenchymatous" morphological transformation series that is consistent with trends observed within each of the three major plant clades. In contrast, a more direct "unicellular ⇒ colonial or siphonous ⇒ parenchymatous" series is observed in fungal and animal lineages. In these contexts, we discuss the roles played by the cooptation, expansion, and subsequent diversification of ancestral genomic toolkits and patterning modules during the evolution of multicellularity. We conclude that the extent to which multicellularity is achieved using the same toolkits and modules (and thus the extent to which multicellularity is homologous among different organisms) differs among clades and even among some closely related lineages.

Reaction-diffusion systems and external morphogen gradients: the two-dimensional case, with an application to skeletal pattern formation.

We investigate a reaction-diffusion system consisting of an activator and an inhibitor in a two-dimensional domain. There is a morphogen gradient in the domain. The production of the activator depends on the concentration of the morphogen. Mathematically, this leads to reaction-diffusion equations with explicitly space-dependent terms. It is well known that in the absence of an external morphogen, the system can produce either spots or stripes via the Turing bifurcation. We derive first-order expansions for the possible patterns in the presence of an external morphogen and show how both stripes and spots are affected. This work generalizes previous one-dimensional results to two dimensions. Specifically, we consider the quasi-one-dimensional case of a thin rectangular domain and the case of a square domain. We apply the results to a model of skeletal pattern formation in vertebrate limbs. In the framework of reaction-diffusion models, our results suggest a simple explanation for some recent experimental findings in the mouse limb which are much harder to explain in positional-information-type models.

Borreliella burgdorferi Antimicrobial-Tolerant Persistence in Lyme Disease and Posttreatment Lyme Disease Syndromes.

The annual incidence of Lyme disease, caused by tick-transmitted Borreliella burgdorferi, is estimated to be at least 476,000 cases in the United States and many more worldwide. Ten to 20% of antimicrobial-treated Lyme disease patients display posttreatment Lyme disease syndrome (PTLDS), a clinical complication whose etiology and pathogenesis remain uncertain. Autoimmunity, cross-reactivity, molecular mimicry, coinfections, and borrelial tolerance to antimicrobials/persistence have been hypothesized and studied as potential causes of PTLDS. Studies of borrelial tolerance/persistence in vitro in response to antimicrobials and experimental studies in mice and nonhuman primates, taken together with clinical reports, have revealed that B. burgdorferi becomes tolerant to antimicrobials and may sometimes persist in animals and humans after the currently recommended antimicrobial treatment. Moreover, B. burgdorferi is pleomorphic and can generate viable-but-nonculturable bacteria, states also involved in antimicrobial tolerance. The multiple regulatory pathways and structural genes involved in mediating this tolerance to antimicrobials and environmental stressors by persistence might include the stringent (rel and dksA) and host adaptation (rpoS) responses, sugar metabolism (glpD), and polypeptide transporters (opp). Application of this recently reported knowledge to clinical studies can be expected to clarify the potential role of bacterial antibacterial tolerance/persistence in Lyme disease and PTLDS.

A regulatory network of two galectins mediates the earliest steps of avian limb skeletal morphogenesis.

BACKGROUND: The skeletal elements of vertebrate embryonic limbs are prefigured by rod- and spot-like condensations of precartilage mesenchymal cells. The formation of these condensations depends on cell-matrix and cell-cell interactions, but how they are initiated and patterned is as yet unresolved. RESULTS: Here we provide evidence that galectins, ß-galactoside-binding lectins with ß-sandwich folding, play fundamental roles in these processes. We show that among the five chicken galectin (CG) genes, two, CG-1A, and CG-8, are markedly elevated in expression at prospective sites of condensation in vitro and in vivo, with their protein products appearing earlier in development than any previously described marker. The two molecules enhance one another's gene expression but have opposite effects on condensation formation and cartilage development in vivo and in vitro: CG-1A, a non-covalent homodimer, promotes this process, while the tandem-repeat-type CG-8 antagonizes it. Correspondingly, knockdown of CG-1A inhibits the formation of skeletal elements while knockdown of CG-8 enhances it. The apparent paradox of mutual activation at the gene expression level coupled with antagonistic roles in skeletogenesis is resolved by analysis of the direct effect of the proteins on precartilage cells. Specifically, CG-1A causes their aggregation, whereas CG-8, which has no adhesive function of its own, blocks this effect. The developmental appearance and regulation of the unknown cell surface moieties ("ligands") to which CG-1A and CG-8 bind were indicative of specific cognate- and cross-regulatory interactions. CONCLUSION: Our findings indicate that CG-1A and CG-8 constitute a multiscale network that is a major mediator, earlier-acting than any previously described, of the formation and patterning of precartilage mesenchymal condensations in the developing limb. This network functions autonomously of limb bud signaling centers or other limb bud positional cues.