Synovial joints are among the most important structures that give us complex motor abilities as humans. Degenerative joint diseases, such as arthritis, cause loss of normal joint functioning and affect over 40 million people in the USA and approximately 350 million people worldwide. Therapies based on regenerative medicine hold the promise of effectively repairing or replacing damaged joints permanently. Here, we introduce a model for synovial joint regeneration utilizing the chick embryo. In this model, a block of tissue that contains the prospective elbow is excised, leaving a window with strips of anterior and posterior tissue intact (window excision, WE). In addition, we also slice out the same area containing the elbow and the distal piece of the limb is pinned back onto the stump (Slice Excision, SE). For making excisions precisely we first carried out a cell fate analysis for elbow forming tissue in the developing limb and carefully determined the tissue to be removed. Interestingly, when the elbow is removed via WE, regeneration of the joint takes place, whereas the elbow joint does not regenerate following SE. In order to investigate whether the regeneration response recapitulates the developmental program of forming joints, first we made a gene expression analysis for the elbow joint because a specific gene expression analysis for the elbow joint was not available in the literature. Among the genes analyzed, we used GDF-5 and Autotaxin (Atx) as joint tissue specific markers and Sox-9 and Col-9 as cartilage markers for in situ hybridization on sections at different time points after WE and SE surgeries. Re-expression of GDF-5 and Atx is observed in the WE samples by 60 hours after the surgery. In contrast, the majority of the samples that underwent SE surgery did not express GDF-5 and Atx. Also, in SE fusion of cartilage elements takes place and the joint interzone does not form. This is indicated by continuous Col-9 expression in SE, whereas Col-9 is down-regulated at the joint interzone in the regenerating WE samples. This order and pattern of gene expression observed in regenerates is similar to the development of a joint suggesting that regeneration recapitulates development at the molecular level. Various growth factors have been shown to trigger or enhance the regenerative response in different models and organs. The regeneration response we observe in WE is present in 50% of the embryos. In order to test the effect of growth factors on this response, we implanted window excised limbs with BMP-2, Noggin, or BSA beads as control. BMP-2 inhibits the joint regeneration, while Noggin does not improve regeneration of the joint tissue. On the other hand, Noggin treatment resulted in elongation of the cartilage elements from the amputated surface This model defines some of the conditions required for inducing joint regeneration in an otherwise nonregenerating environment. This knowledge can be useful for designing new therapeutic approaches for joint loss or for conditions affecting joint integrity in humans / acase@tulane.edu

<p> Microbial biofilms on leaf litter can assimilate nutrients, thus altering the elemental composition (i.e. stoichiometric composition) of detritus. Aquatic macroinvertebrates consuming stoichiometrically-altered detritus may increase, decrease, or not change growth rates depending upon their body stoichiometry. Invertebrates with high body phosphorus (P) can allocate more P to rRNA, increasing protein production and growth. Thus, if diet nutrient limitation is alleviated, high P invertebrates are expected to increase growth rates. Conversely, low P invertebrates might not respond or decrease growth rates, due to the metabolic costs of eliminating excess nutrients. Diptera from two families were used in growth studies and fed either maple or oak leaves with different levels of carbon:phosphorus (C:P) to determine if diet P influenced growth rates and body stoichiometry of these high P organisms. </p><p> <i>Tipula abdominalis</i> from the family Tipulidae, were hypothesized to increase growth and consumption rates, and decrease body C:P as diet C:P decreased. It was also hypothesized that the proportion of consumed material composed of a particular element used in growth would increase (GGE), and that the diet ratio at which neither C or P is limiting, and at which maximum growth would occur, would decrease as diet C:P decreased (TERC:P). It was observed that <i>T. abdominalis</i> growth and consumption varied based on leaf type and elemental composition. <i>Tipula abdominalis</i> fed oak had high growth (5.4% day-1) and consumption (1.98mg mg tipulid-1 day-1) compared to maple diets, while individuals fed maple increased growth (2.6 to 5.0% day-1) and consumption (1.1 to 2.1mg mg tipulid-1 day-1) across treatments as maple C:P decreased. <i>Tipula abdominalis</i> were overall homeostatic in body stoichiometry. Contrary to expectations, GGEs decreased as diet C:P decreased, maintaining stoichiometric homeostasis. Also, current TER calculations were not representative of observed growth rates. Thus, results indicate that leaf stoichiometry and leaf type interact to influence growth rates. </p><p> Three genera of Chironomidae (<i>Chironomus, Polypedilum, Micropsectra </i>) were fed a gradient of diet C:P to determine if there are differences in taxon-specific growth responses to changes in diet stoichiometry. It was hypothesized that genera with higher body P would increase growth more than genera with lower body P, and that body P in high P genera would increase as they consumed more dietary P. The three genera had different growth responses to decreased diet C:P that did not appear to be influenced by organism body stoichiometry or diet stoichiometry. <i>Chironomus</i> fed maple and oak and <i>Micropsectra</i> fed maple had similar growth rates (17.5% day^-1), while having significantly different body C:P, 141 and 249 for <i>Chironomus</i> fed oak and maple, respectively, and 359 for <i>Micropsectra</i> fed maple. Despite similarities in diet C:P ranges (1000 to 8000), <i>Chironomus</i> and <i> Polypedilum</i> had different growth responses: <i>Chironomus</i> had high growth, while <i>Polypedilum</i> fed maple did not grow and lost mass (- 4.8% day^-1), and Polypedilum fed oak exhibited relatively low growth rates (5.9% day^-1). Leaf type and genus identity appear to be important factors in determining growth response. However, most Dipterans in this study either increased growth or had no response to dietary P enrichment. Understanding how organism genus identity interacts with leaf type and leaf stoichiometry to influence growth rates will allow more accurate predictions of community changes in nutrient enriched systems. </p>

<p> Many organisms live in temporally or spatially heterogeneous environments. One adaptive response to environmental heterogeneity is phenotypic plasticity, or the ability of an organism to change its phenotype in response to environmental variation. Plasticity is particularly important for plants, which cannot easily escape environmental stresses (e.g., competition, herbivory, or drought), and therefore is common in many morphological and physiological traits that allow plants to cope with stresses. Despite numerous examples of the adaptive value of plasticity in plant traits, the reaction norms of many plant traits do not evolve to their predicted optima: there is often considerable genetic variation in trait plasticities, and some populations display sub-optimal responses to environmental stresses. This mismatch may reflect a balance between the fitness benefits (adaptive value) of plasticity and evolutionary constraints, such as strong genetic correlations with other traits, a lack of genetic variation in the reaction norm, and fitness costs of plasticity. </p><p> This dissertation focuses on the evolution of plasticity in plant defense responses to insect herbivores. Plant induced defenses are well-studied ecologically, but little is known about potential constraints on their evolution or the effects of these constraints on insect herbivores. Using the andromonecious herb <i>Solanum carolinense</i>, I examined the adaptive value of plasticity and tested for several evolutionary constraints on plant induced defenses and tolerance. I found damage-induced plasticity in several defense traits, and herbivores significantly reduced plant fitness in the field. However, neither tolerance nor plasticity in induced defenses was adaptive, and most traits were selected against (i.e., costly) in both environments with and without insect herbivores. </p><p> I uncovered several genetic constraints on plasticity in the plant defense response, including a) a lack of genetic variation in some resistance traits, b) strong genetic correlations between constitutive resistance and inducibility, and c) genetic correlations among the plasticities of different defense traits. Combined, these genetic constraints structured variation in the plant-mediated interaction between the specialist folivores <i>Manduca sexta</i> and <i>Leptinotarsa juncta</i>; this is the first study to demonstrate that genetic trade-offs within a plant species can affect interactions at higher trophic levels. </p><p> Although there were no fitness costs of plasticity in induced chemical/mechanical defenses, there were fitness costs of tolerance, and I was able to detect selection acting directly on trait plasticities. Selection on plasticities was generally positive or stabilizing, indicating fitness benefits of canalization of two defense traits I measured. Fitness costs of tolerance were present both in environments with and without herbivores, meaning that tolerance of herbivory was never adaptive. My dissertation is the first to simultaneously test for both the adaptive value and a breadth of evolutionary constraints (including costs of plasticity) on the plant defense response. </p><p> Combined, my results indicate that selection should reduce both the plasticity and mean expression of <i>S. carolinense</i> defenses; however, correlated selection on plastic traits or a lack of genetic variation in plasticity may prevent this from occurring. Most importantly, these results suggest that two basic predictions of evolutionary theory - that phenotypic plasticity should be both adaptive and costly - do not always hold true for plant defense traits. This underscores the importance of measuring selection on any plastic defense trait assumed to be adaptive, and investigating other potential evolutionary constraints on defensive plasticity besides fitness costs. My study shows that genetic correlations may also be important, but correlations among defense trait plasticities have been rarely investigated and most studies do not account for the presence of these correlations when examining selection on individual defense traits.</p>

<p> The formation of mineralized tissue was critical to the evolution and diversification of metazoans and remains functionally significant in most animal lineages. Of special importance is the protein found occluded within the mineral matrix, which facilitates the process of biomineralization and modulates the final mineral structure. These skeletal matrix proteins have well been described in several species, including the sea urchin <i> Stronglyocentrotus purpuratus,</i> an important model organism. Biomineralization research is limited in other echinoderm classes. This research encompasses the first description of mineral matrix proteins in a member of the echinoderm class Ophiuroidea. This work describes the skeletal matrix proteins of the brittle star <i>Ophiocoma wendtii</i> using bioinformatic and proteomic techniques. General characteristics of matrix protein are described and a number of candidate biomineralization related genes have been identified, cloned, and sequenced. The unique evolutionary and biochemical properties of brittle star skeletal matrix proteins are also described.</p>

<p>This project explores species boundaries, coevolution, biodiversity, parasite life cycles, and ecology using the cestode (tapeworm) fauna parasitizing the spiny dogfish, <i>Squalus acanthias</i>, and several of its close relatives. One of the aims was to verify the species identity of all the shark specimens from which cestodes were collected. Chapter 1 details use of the elasmobranch "barcoding" gene, NADH2, to verify host identifications, as well as to raise doubt about the wisdom of recognizing the Black Sea population of <i>S. acanthias</i> as a distinct subspecies. Chapter 2 examines diversity in the monotypic cestode genus, <i>Trilocularia</i>, throughout the widespread anti-tropical distribution of <i>S. acanthias</i> (i.e., the North and South Pacific, the North Atlantic, and the Black Sea), and in its congeners. An integrative approach, including morphological (i.e., light microscopy, histology, and scanning electron microscopy) and molecular methods (i.e., 28S, ITS1, 16S genes), was employed. Results reveal a large amount of undiscovered diversity in this genus and suggest that species of <i> Trilocularia</i> may be undergoing speciation more rapidly than their hosts. Chapter 3 describes one of the new species discovered, <i>Trilocularia eberti</i> n. sp. from <i>S.</i> cf. <i>mitsukurii</i>, and provides a prototype for future descriptions of species in this genus. Chapter 4 investigates microthrix variation in <i>Trilocularia</i> from the stomach and the spiral intestine of <i>S. acanthias</i> off Rhode Island. Results suggest that variation seen within a host individual is likely due to developmental changes rather than species differences. Chapter 5 aims to further the understanding of cestode infections in a marine environment through space and time by examining seasonal infection parameters in the cestode community of <i>S. acanthias</i> from Rhode Island across three years. While some general trends may be maintained across disparate localities, spatial variation is likely due to differences in accessibility to intermediate hosts and host diet across sites. The knowledge gained from understanding cestode infections in the vast ocean environment allows us to speculate about the factors driving fluctuations in parasite infections in elasmobranchs. </p>

<p>Centromeres are the chromosomal loci where microtubule spindles bind, via the kinetochore, during mitosis and meiosis. Paradoxically the centromere, as a functional unit, is essential to guarantee faithful chromosome segregation, whereas its underlying DNA sequences and associated kinetochore proteins are fast evolving. In most animals and plants that have been studied, centromeres contain megabase-scale arrays of tandem repeats. In spite of their importance, very little is known about the degree to which centromeric tandem repeats share common properties between different species across different phyla. We used bioinformatic methods to identify high-copy tandem repeats from species using publicly available genomic sequence and our own data. We found that despite an overall lack of sequence conservation, centromeric tandem repeats from diverse species showed similar modes of evolution. Furthermore, phylogenetic analysis of sequence homology showed little evidence of sequence conservation beyond approximately 50 million years of divergence. In addition, we performed a survey of fungi genomes for the presence of high-copy tandem repeats, but found little evidence to suggest that high-copy centromeric repeats are a common feature feature in fungi, with the possible exception of the <i>Zygomycota</i>. phylum. Finally, in most species the kinetochore assembles at a single locus, but in some cases the kinetochore forms along the entire length of the chromosomes forming holocentric chromosomes. Following a literature review we estimate that holocentricity is very common and has evolved at least thirteen times.

<p> In this work, I describe my doctoral work studying the regulation of transcription with both computational and experimental methods on the natural genetic variation in a population. This works integrates an investigation of the consequences of polymorphisms at three stages of gene regulation in the developing fly embryo: the diversity at <i>cis</i>-regulatory modules, the integration of transcription factor binding into changes in chromatin state and the effects of these inputs on the final phenotype of embryonic gene expression.</p>

<p> This study used fine-scale phylogeography of populations of side-blotched lizard, <i>Uta stansburiana</i> to determine concordance with previously identified phylogeographic patterns in the Transverse Mountain Range of Southern California. Mitochondrial DNA analysis revealed clade distributions that both agreed and conflicted with previously identified clade breaks. This study revealed two new distinct clade breaks that transect the San Gabriel Mountains and detected haplotype mixtures in populations sampled between the San Gabriel and San Bernardino Mountains. Genetic landscape GIS analyses identified areas of genetic divergence and diversity for this species. This combined analysis enabled the discovery of a suture zone between the San Gabriel and San Bernardino Mountains that represents a phylogenetic crossroads with high levels of diversity and divergence and complex phylogeographic structure. My results suggest the importance of the use of fine-scale phylogeographic analysis in the discovery of new clade boundaries within a geologically complex and hyper-diverse region.</p>

<p> Signaling theory originated in evolutionary biology and explains the mechanisms behind the honest communication of information between organisms. Communication scholars are increasingly turning to signaling theory as a way to test evolutionary explanations for human behavior. The present study tests if receiver-dependent costly signals can be used to predict the moment of aggressive behavior in video game environments. High status (but not high trait aggression) male subjects were fastest to engage in combat against a low voice pitch male opponent - but only when subject skill was high. This result underscores the importance of video game skill as a variable of interest as well as the need for video games researchers to tease out when real-world behaviors map to video game contexts.</p>

<p>Much of plant growth is directly or indirectly regulated by the plant hormone auxin. Although some genes involved in the auxin pathway have been characterized, there are still gaps in our knowledge of this genetic pathway. Recently, the orthologous maize genes <i>BARREN STALK2</i> (<i> BA2</i>) and rice gene <i>LAX PANICLE2</i> (<i>LAX2</i>) have been cloned and characterized. Maize <i>ba2</i> and rice <i> lax2</i> mutant plants both show significant flaws in axillary meristem (AM) initiation, suggesting a role in auxin regulation. In support of this hypothesis, <i>LAX2</i> interacts with the auxin regulating <i> LAX PANICLE1</i> (<i>LAX1</i>) protein. My research reconstructs the evolutionary history of the <i>BA2</i>/<i>LAX2</i> lineage and tests for conservation of <i>BA2</i>/<i>LAX2</i> mRNA expression in diverse grasses. My results indicate that the <i> BA2</i>/<i>LAX2</i> gene family&mdash;comprising <i>BA2 </i>/<i>LAX2</i>, <i>BA2</i>/<i>LAX2-Like1</i> (<i>BA2</i>/<i>LAX2L1</i>), and <i>BA2</i>/<i> LAX2-Like2</i> (<i>BA2</i>/<i>LAX2L2</i>)&mdash;is restricted to monocots, and shows evidence of two independent gene duplication events. During its evolution, the <i>BA2</i>/<i>LAX2</i> gene family appears to have gained a PDZ Protein Binding motif, which may allow it to interact with other proteins besides <i>BA1</i>/<i> LAX1</i>. My expression analyses show that <i>BA2</i>/<i> LAX2</i> genes are expressed during multiple stages of inflorescence development, and this expression is conserved across multiple grass species. A unique floral expression pattern appears to have evolved at the base of the Joinvilleaceae lineage. My study further supports the hypothesis that <i> BA2</i>/<i>LAX2</i> genes are functioning in multiple AM pathways. </p>