Creatine kinase (CK) and arginine kinase (AK) are members of a highly conserved family of phosphoryl transferase enzymes known as phosphagen kinases. CK and AK play a central role in cells that display high and variable rates of ATP turnover such as neurons, muscle fibers, spermatozoa, and transport epithelia. All CK isoforms exhibit targeting to specific tissues. In vertebrates CK exists as two dimeric, cytoplasmic isoforms, B (brain) and M (muscle) and two octameric, mitochondrial isoforms termed sarcomeric (SarMtCK) and ubiquitous (UbiMtCK). In some groups cytoplasmic dimers and mitochondrial octamers are found as well as a contiguous trimeric form, flagellar CK (flgCK). This latter isoform is found in the primitive-type spermatozoa of protochordates and certain protostome and deuterostome invertebrates. FlgCKs are myristolated and are found anchored to the flagellar membrane. AK isoforms are found as monomers, dimers, and contiguous dimers. AK is present in protozoans, protostome and deuterostome invertebrates, and protochordates, but not in craniates. Recently, a dimeric mitochondrial creatine kinase (MtCK) and a protoflagellar creatine kinase (protoflgCK) have been cloned and sequenced from the demosponge Tethya aurantia (Sona et al. 2004) and AK has been cloned and sequenced from the demosponge Suberites domuncula (Perovic-Ottstadt et al. 2005). Based on this earlier work we have surveyed all three classes of the poriferans, arguably the most primitive, basal metazoans, for the presence of CK and AK using spectrophotometric assays of tissue extracts as well as molecular biology protocols. Total RNA was isolated from three demosponges and three hexactinellids. RT-PCR was performed and cDNAs were amplified consisting of both mitochondrial and protoflagellar CK isoforms for two demosponges and two out of three hexactinellids tested. A gene for protoflgCK was amplified from a third hexactinellid. AKs were amplified from two demosponges and one hexactinellid. Our results show that AK is widely distributed in all three sponge classes. CK is present in two of the sponge classes and the genes for the mitochondrial and protoflagellar isoforms evolved prior to the divergence of the hexactinellids. It is highly probable that CK evolved in the sponges or in their immediate ancestors, the urmetazoans, or possibly even earlier. The deduced amino acid sequences from the amplified cDNAs were aligned and used in phylogenetic analyses along with CKs from other invertebrates and vertebrates and AKs from protozoans, invertebrates, and protochordates to elucidate the early evolution of the CK and AK gene families. Maximum Likelihood analysis using PHYLIP produced a CK tree with two distinct clades: mitochondrial and cytoplasmic-protoflagellar-flagellar. Using an outgroup composed of AKs, GKs, LKs, and TKs for evolutionary direction, our tree suggests that the ancestral CK dimer was MtCK-like and strongly suggests that the MtCK isoform was the first to evolve. Cyt, protoflg, and flgCK are closely related as well and phylogenetic evidence supports the idea that protoflgCKs are ancestral to flgCKs. The phylogenetic analysis completed in this effort provides the strongest evidence yet for a gene duplication event producing the true CytCKs and protoflgCKs. Maximum Likelihood analysis produced an AK tree with a base of protozoan, poriferan, and cnidarian AKs, and a clade of higher ecdysozoan and lophotrochozoan invertebrate AKs. To supplement this phylogenetic data and to further understand the early evolution of the CK isoforms, genomic DNA was isolated from Suberites ficus and the mitochondrial and protoflagellar genes were amplified by PCR. The gene organization data provides additional support that protoflgCKs are ancestral to flgCKs and that MtCKs are highly conserved in terms of their intron:exon organization. The present effort focuses on the distribution and early evolution of AK and CK in the poriferans, the pattern of gene duplication and divergence responsible for CK isoform diversity, and the development of intracellular targeting. The location and physiological roles of AK and CK will also be hypothesized and well as examining the utility of CK and AK as characters for sponge systematics. We will also explore structure-function relationships of these proteins and propose an evolutionary pathway for phosphagen kinases. / A Thesis Submitted to the Department of Biological Science in Partial Fulfillment of the Requirements for the Degree of Master of Science. / Spring Semester, 2006. / March 31, 2006. / Molecular Evolution / Includes bibliographical references. / W. Ross Ellington, Professor Directing Thesis; Gavin J. P. Naylor, Committee Member; Janie Wulff, Committee Member.

published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

In the mid-twentieth century, French philosophers looked to contemporary biological research as they attempted to come to grips with the philosophical and historical crises of the previous decades. My dissertation provides a genealogy of the relationship between developments in the fields of evolutionary theory, genetics, and embryology, and the emergence of French structuralism and posthumanist history. The story centers around two generations of French philosophers, including Raymond Aron, Georges Canguilhem, Maurice Merleau-Ponty, and Raymond Ruyer, and the biologists they turned to as resources for their philosophy, including Maurice Caullery, George E. Coghill, Étienne Rabaud, and Étienne Wolff. As I show, because these philosophers did not look to “life” as a metaphor or to “science” understood as either mere ideology or pure rationality, but instead grappled directly with the specific content of evolutionary theory, embryology, and genetics, biology profoundly reshaped the philosophical concepts of human and history.

This dissertation is an exploration of phenomena on varying scales, built on the backbone of Cenozoic mammalian biochronologic units (Land Mammal ‘Ages’): the integration of fossil and geological data to constrain spatiotemporal patterns in evolution. I develop and test hypotheses about the origins and ordinal-level relationships of the extinct South American endemic placental order Notoungulata, as well as about some more specific macroevolutionary patterns at a familial level within notoungulates. Major novel outcomes include a new biochronologic timescale for the terrestrial Cenozoic of South America, numerically calibrated through synthesis of new and existing high-precision geochronological data (particularly U-Pb and 40Ar/39Ar dating), along with an explicit logical framework for Land Mammal “Age” calibration; description of two new interatheriid notoungulate taxa from the central Chilean Andes; a new phylogenetic hypothesis for the position of Notoungulata within Placentalia; and the first ever DNA sequences obtained from the extinct musk ox relative Bootherium bombifrons. The first study, chapter two, is a review of geochronologic (radiometric and magnetostratigraphic) constraints for the South American Land Mammal “Age” timescale. I present a revised, updated timescale, and a descriptive, logical framework for the synthesis of geochronologic and biochronologic data from a variety of sources and analytical methods. Significant changes to the calibration of individual SALMAs in this update are concentrated in the Paleogene, while Neogene calibrations have remained fairly stable, with small refinements to the core age ranges of the Huayquerian, Chasicoan, Colloncuran, and Friasian. This section also investigates the influence of latitudinal biotic provinciality on correlations and chronologic calibrations, particularly as far as provinciality reflects the climatic evolution of the continent. Marked provinciality is evident at least as early as the early Eocene, with faunas like Itaboraí differing from potential correlative faunas at higher latitudes, potentially representing unique periods in mammalian evolution, both faunally and chronologically. Instead of trying to recognize and correlate the classical high latitude SALMAs to highly distinctive tropical assemblages, the SALMA timescale should allow for the development of separate mammalian biochronologic zonations for low and high latitudes. Chapter three presents and describes two new notoungulate taxa, representing the first species formally described from the Los Queñes Fauna, a late Eocene mammal assemblage from the Andean Main Range of central Chile. These two taxa, Anabalcarcel ignimbritae and Jackconrad carreterensis, represent the earliest hypsodont interatheres known. Based on ancestral state reconstructions using parsimony, hypsodonty appeared no later than the latest Eocene (34.6 ± 0.8 Ma; likely correlative with the Mustersan South American Land Mammal Age) in interatheres, a time when this dental specialization was not yet pervasive among other mammalian herbivores. Tree-based comparative analyses revealed two significant taxonomic radiations of interatheres, the early radiation of basal interatheriids and a later radiation of hypselodont taxa after the interval of dramatic global climatic change associated with the Eocene/Oligocene boundary. In chapter four, I attempted to apply some of the geochronologic methods incorporated in the first two chapters, to date a newly discovered site in Abanico Formation, central Chilean Andes. This formation yields fossil mammals in numerous areas, including at several localities in the Río Las Leñas and Río Cachapoal drainages. In the Cachapoal Valley, steeply-dipping beds have yielded fossils of Tinguirirican age, including a polydolopine marsupial and an interatheriid notoungulate. The results of this study, the first 40Ar/39Ar analysis from the Cachapoal Valley, are a date of 11.1 ± 1.8 Ma, from stratigraphically higher levels loosely constraining the minimum age for fossiliferous deposits. Chapters five and six change tack a little, and investigate analytical methods in ancient-DNA phylogenomics, time-calibrated with radiocarbon dates, using as a study system Bootherium bombifrons, the endemic North American helmeted musk ox that went extinct, along with most of its megafaunal colleagues, at the end of the Pleistocene. This switch to a North American, rather than South American, endemic mammal was driven by availability of molecular comparative material and the presence of hundreds of helmeted musk ox specimens in the American Museum paleontology collections, which I was able to sample destructively. I present the first complete mitochondrial genome of the extinct musk ox Bootherium bombifrons, and new mitochondrial sequence data from seven individuals referred to Bootherium bombifrons Additionally, I obtained new radiocarbon dates from these same specimens, of 38580 ± 720, 30240 ± 260, 44570 ± 190, 41040 ± 910, 44240 ± 1500, 39080 ± 770, 28370 ± 210, and 47190 ± 2100 calibrated 14C years BP. Phylogenetic analysis supports placement of Bootherium as sister to the living musk ox, Ovibos moschatus, in agreement with morphological taxonomy. I also tested hypotheses regarding the impact that locus choice has on divergence date estimates using tip calibrations in this taxon. Estimates of the root age of three different gene trees for Bootherium specimens ranged from 19 ka, for cytochrome B, to over 50 ka for the mitochondrial control region. The final study in this dissertation (chapter seven) is a superordinal scale approach to fitting Notoungulata into the picture of all placental mammals through synthesis and expansion of existing total-evidence matrices. Results presented here corroborate previous findings that notoungulates are most closely related to Afrotheria, but do not support afrothere and notoungulate reciprocal monophyly, suggesting a complex biogeographic relationship between South America and Africa persisting from Late Cretaceous through mid-Paleocene.

The complexity of ecological and evolutionary processes that govern species distributions has long presented a challenge to understanding community assembly history. The work presented here develops a conceptual framework for integrating phylogenetics and biogeography to reconstruct the assembly of communities, provides empirical support for the broad applicability of this framework, tests whether morphology can serve as a proxy for behavioral ecology, and develops a novel metric of assemblage vulnerability and shows how vulnerability is related to biogeographic history. This dissertation demonstrates the need to merge evolution and ecology to reconstruct community assembly, and provides a framework for doing so. Further, the findings presented here suggest that such an interdisciplinary approach has the potential to both reveal fundamental processes shaping the assembly of natural systems, and to illuminate the functions and properties of ecosystems based on the evolutionary histories of their constituent species.

Evolutionary processes in any population depend upon patterns of phenotypic variation available to selection and their underlying heritability. In this thesis, I used the filamentous red seaweed Asparagopsis armata, with particular focus on its modularity, to test several key questions underlying its growth-form evolution in heterogeneous environments. I established that experimental manipulations of light quantity and quality mimicking variation in underwater light due to shading or depth induce growthform plasticity in A. armata that may be evolutionarily significant given its variability among clones. Current patterns of plasticity displayed by A. armata appear adaptive, moreover, given that a reciprocal transplant of phenotypes between light environments found densely-branched (phalanx-like) phenotypes to have higher relative growth rates than sparsely-branched (guerrilla-like) phenotypes in well-lit patches, but lower relative growth rates than the latter in shaded patches. Using the capacity for rapid growth as a proxy for fitness, multivariate selection analyses identified environment-dependent patterns of directional selection on single traits coupled with linear and nonlinear selection on multi-trait combinations that shape growth-form variation within patches of differing light intensity, thereby reinforcing plasticity across light environments. Quantitative genetic analyses, however, suggest that the modular iteration of genes in morphogenesis may limit further growth-form evolution in A. armata populations exposed to spatial heterogeneity in light by constraining thallus responses to environment-dependent selection. Last, heritable responses to artificial selection on growth-form variation among clonal cell-lineages revealed the surprising capacity for A.armata to circumvent genetic constraints inherent to its development by adapting to environmental change in the absence of sexually-generated variance among clones.

The release of cells from a biofilm to the surrounding environment is poorly understood and the importance of this stage of biofilm development has only recently been realized. A key part of this process is the generation of phenotypic variants in the biofilm dispersal population. This thesis reports on the characterization of biofilm development of Serratia marcescens MG1, the analysis of the biofilm dispersal population, and the identification of the conditions that trigger phenotypic diversification. Furthermore, it provides an insight into the molecular understanding of how phenotypic variation is being generated, and demonstrates the clinical and environmental implications of phenotypic diversification during bacterial pathogenesis and bacterial persistence. Characterization of the microcolony biofilm development of S. marcescens revealed that the S. marcescens biofilm develops through a process involving microcolony formation, hollowing of mature microcolonies, and a sudden biofilm expansion within a very short period (&lt 24h) resulting in an increase in biofilm biomass with a radiation of biofilm structures at days 3 to 4. The biofilm expansion phase consistently correlated to an increase in the number of dispersal variant morphotypes. Studies of variant induction in planktonic cultures and biofilm flow cells demonstrated that phenotypic diversification in S. marcescens is not only a biofilm-specific phenomenon, but also involves biofilm-specific morphotypes. These morphological variants can only be isolated from the microcolony biofilm morphotype and not from the filamentous biofilms, leading to the hypothesis that there is a strong diversifying selection that is specific to the microcolony biofilms. To further explore how these variants were generated, molecular analyses revealed that exopolysaccharides and lipopolysaccharides are important moieties that are involved in phenotypic variation in S. marcescens biofilms. The etk gene, encoding a tyrosine protein kinase within the exopolysaccharide biosynthesis operon, was found to contain single nucleotide polymorphisms (SNPs) that were present in the 'sticky' variants but not in the 'non-sticky' wild-type or the 'non sticky' small colony variants. Furthermore, infrequent-restriction-site PCR (IRS-PCR), BIOLOG metabolic profiling, and gene sequence analyses, suggest that phenotypic diversification in S. marcescens is likely to involve mutational hotspots in specific genes. The biofilm-derived morphotypic variants differed extensively in cell ultrastructure properties, and exhibited specialized colonization and virulence traits, such as attachment, biofilm formation, swimming and swarming motilities, protease production, and hemolysin production. It was also demonstrated that phenotypic diversification contributed to a varying degree of resistance to protozoan predation, and bacterial pathogenecity in Caenorhabditis elegans, highlighting the complexity of the dispersal populations from S. marcescens biofilms. Furthermore, mixed-culture experiments involving multiple variant isolates (with or without the parental wild-type) showed that the persistence and virulence potential of S. marcescens can be synergistically enhanced in the Acanthamoeba castellanii grazing model and in the C. elegans infection model, respectively. This indicates that the different bacterial morphotypes work in concert to provide S. marcescens with enhanced protection against environmental perturbations and a competitive edge during the infection process. It was proposed that phenotypic diversification is not only an integral part of S. marcescens biofilm life-cycle, but also represents an important strategy for bacteria to greatly enhance its survival and persistence in different environments, ranging from aquatic and soil ecosystems, to those of the infected hosts.

Caenorhabditis elegans is arguably the best understood animal on the planet. Used for over 50 years to study development, we have a vast amount of knowledge of the inner workings of this worm. Our knowledge is incomplete, however, without placing this organism in its evolutionary and ecological context. In this body of work, I focused on examining the evolutionary forces shaping Caenorhabditis nematodes, with a particular emphasis on C. briggsae. In the first part, I examined the evolution of mitochondrial genomes throughout the genus. I tested for signatures of selection and examined the evolution of mitochondrial genome architecture. Through this, I have shown that the mitochondrial genomes of Caenorhabditis nematodes appear to be primarily influenced by purifying selection and that molecular evolutionary inference is greatly limited by mutational saturation. The evolutionary forces acting on mitochondrial genomes have been examined before, however, this study, extensively examining this within a single genus, provides a much better characterization than any of the studies to date. In the second part, I characterized the evolutionary dynamics of mitochondrial pseudogenes in C. briggsae and its closest relatives. I showed that these elements, while they might not evolve under strictly neutral terms, are still quite useful in uncovering cryptic diversity and population structure. I also observed that they appear/disappear in a manner that appears inconsistent with one commonly held model for mitochondrial pseudogene evolution. In the final part, I examined the evolution of C. briggsae in response to a biotic environment. I showed that fitness in a parasite-containing environment incurs a trade-off with fitness in the absence of parasites. Together, the chapters of this dissertation demonstrate the strength of Caenorhabditis, and in particular C. briggsae, for examining evolutionary questions and advances this system as a tool for evolutionary biology research. / Graduation date: 2013

In social and biological evolution, infelicity can operate as a driving motor to force change. In this essay, for life other than human, infelicity is equated with physical unfitness to compete for the resources of a specific niche. For humanity it is defined as the result of an incongruity between a nation's culture and its government. The purpose of this study is to investigate how, for irrational life, unfitness can stimulate the creation of a new species and, for men, how the unhappiness of a nation may enhance its opportunity to enter a new socio-economic order. An evolutionary account about a possible way in which life could have evolved is offered, concentrating mainly on the transition from ape to a less remote ancestor of man, but also taking into consideration other life forms. Then, a parallel to social evolution is established. A study of the rise of capitalism in England, as well as the recent attempts to institute socialism in Latin America, are also explained as consequences of infelicity. / Graduation date: 1994

Thesis (Ph. D.)--University of Chicago, Dept. of Philosophy, 2002. / Includes bibliographical references. Also available on the Internet.