Characterization and developmental regulation of a gene expressed specifically in the skeletogenic lineage of the sea urchin embryo

The sea urchin embryonic skeleton, or spicule, is deposited by mesenchymal progeny of four precursor cells, the micromeres, which are determined to the skeletogenic pathway by a process known as cytoplasmic localization. A gene encoding one of the major products of the skeletogenic mesenchyme, a prominent 50 kD protein of the spicule matrix, has been characterized in detail. cDNA clones were first isolated by antibody screening of a phage expression library, followed by isolation of homologous genomic clones. The gene, known as SM50, is single copy in the sea urchin genome, is divided into two exons of 213 and 1682 bp, and is expressed only in skeletogenic cells. Transcripts are first detectable at the 120 cell stage, shortly after the segregation of the skeletogenic precursors from the rest of the embryo. The SM50 open reading frame begins within the first exon, is 450 amino acids in length, and contains a loosely repeated 13 amino acid motif rich in acidic residues which accounts for 45% of the protein and which is possibly involved in interaction with the mineral phase of the spicule.

The important cis-acting regions of the SM50 gene necessary for proper regulation of expression were identified by gene transfer experiments. A 562 bp promoter fragment, containing 438 bp of 5' promoter sequence and 124 bp of the SM50 first exon (including the SM50 initiation codon), was both necessary and sufficient to direct high levels of expression of the bacterial chloramphenicol acetyltransferase (CAT) reporter gene specifically in the skeletogenic cells. Removal of promoter sequences between positions -2200 and -438, and of transcribed regions downstream of +124 (including the SM50 intron), had no effect on the spatial or transcriptional activity of the transgenes.

Regulatory proteins that interact with the SM50 promoter were identified by the gel retardation assay, using bulk embryo mesenchyme blastula stage nuclear proteins. Five protein binding sites were identified and mapped to various degrees of resolution. Two sites are homologous, may be enhancer elements, and at least one is required for expression. Two additional sites are also present in the promoter of the aboral ectoderm specific cytoskeletal actin gene CyIIIa; one of these is a CCAA T element, the other a putative repressor element. The fifth site overlaps the binding site of the putative repressor and may function as a positive regulator by interfering with binding of the repressor. All of the proteins are detectable in nuclear extracts prepared from 64 cell stage embryos, a stage just before expression of SM50 is initiated, as well as from blastula and gastrula stage; the putative enhancer binding protein may be maternal as well.

Rate and Microstructure Effects on the Dynamics of Carbon Nanotube Foams

Soft hierarchical materials often present unique functional properties that are sensitive to the geometry and organization of their micro- and nano-structural features across different lengthscales. Carbon Nanotube (CNT) foams are hierarchical materials with fibrous morphology that are known for their remarkable physical, chemical and electrical properties. Their complex microstructure has led them to exhibit intriguing mechanical responses at different length-scales and in different loading regimes. Even though these materials have been studied for mechanical behavior over the past few years, their response at high-rate finite deformations and the influence of their microstructure on bulk mechanical behavior and energy dissipative characteristics remain elusive.

In this dissertation, we study the response of aligned CNT foams at the high strain-rate regime of 10² - 10⁴ s^-1. We investigate their bulk dynamic response and the fundamental deformation mechanisms at different lengthscales, and correlate them to the microstructural characteristics of the foams. We develop an experimental platform, with which to study the mechanics of CNT foams in high-rate deformations, that includes direct measurements of the strain and transmitted forces, and allows for a full field visualization of the sample’s deformation through high-speed microscopy.

We synthesize various CNT foams (e.g., vertically aligned CNT (VACNT) foams, helical CNT foams, micro-architectured VACNT foams and VACNT foams with microscale heterogeneities) and show that the bulk functional properties of these materials are highly tunable either by tailoring their microstructure during synthesis or by designing micro-architectures that exploit the principles of structural mechanics. We also develop numerical models to describe the bulk dynamic response using multiscale mass-spring models and identify the mechanical properties at length scales that are smaller than the sample height.

The ability to control the geometry of microstructural features, and their local interactions, allows the creation of novel hierarchical materials with desired functional properties. The fundamental understanding provided by this work on the key structure-function relations that govern the bulk response of CNT foams can be extended to other fibrous, soft and hierarchical materials. The findings can be used to design materials with tailored properties for different engineering applications, like vibration damping, impact mitigation and packaging.

I. The rate and mechanism of the vapor-phase reaction between water and parts-per-million concentrations of nitrogen dioxide. II. The rate and mechanism of the air oxidation of parts-per-million concentrations of nitric oxide in the presence of water vapor.

Part I

A study of the thermal reaction of water vapor and parts-per-million concentrations of nitrogen dioxide was carried out at ambient temperature and at atmospheric pressure. Nitric oxide and nitric acid vapor were the principal products. The initial rate of disappearance of nitrogen dioxide was first order with respect to water vapor and second order with respect to nitrogen dioxide. An initial third-order rate constant of 5.5 (± 0.29) x 10⁴ liter² mole^-2 sec^-1 was found at 25˚C. The rate of reaction decreased with increasing temperature. In the temperature range of 25˚C to 50˚C, an activation energy of -978 (± 20) calories was found.

The reaction did not go to completion. From measurements as the reaction approached equilibrium, the free energy of nitric acid vapor was calculated. This value was -18.58 (± 0.04) kilocalories at 25˚C.

The initial rate of reaction was unaffected by the presence of oxygen and was retarded by the presence of nitric oxide. There were no appreciable effects due to the surface of the reactor. Nitric oxide and nitrogen dioxide were monitored by gas chromatography during the reaction.

Part II

The air oxidation of nitric oxide, and the oxidation of nitric oxide in the presence of water vapor, were studied in a glass reactor at ambient temperatures and at atmospheric pressure. The concentration of nitric oxide was less than 100 parts-per-million. The concentration of nitrogen dioxide was monitored by gas chromatography during the reaction.

For the dry oxidation, the third-order rate constant was 1.46 (± 0.03) x 10⁴ liter² mole^-2 sec^-1 at 25˚C. The activation energy, obtained from measurements between 25˚C and 50˚C, was -1.197 (±0.02) kilocalories.

The presence of water vapor during the oxidation caused the formation of nitrous acid vapor when nitric oxide, nitrogen dioxide and water vapor combined. By measuring the difference between the concentrations of nitrogen dioxide during the wet and dry oxidations, the rate of formation of nitrous acid vapor was found. The third-order rate constant for the formation of nitrous acid vapor was equal to 1.5 (± 0.5) x 10⁵ liter² mole^-2 sec^-1 at 40˚C. The reaction rate did not change measurably when the temperature was increased to 50˚C. The formation of nitric acid vapor was prevented by keeping the concentration of nitrogen dioxide low.

Surface effects were appreciable for the wet tests. Below 35˚C, the rate of appearance of nitrogen dioxide increased with increasing surface. Above 40˚C, the effect of surface was small.

The rate and mechanism of the partial oxidation of acetaldehyde by PPM concentrations of nitrogen dioxide

The thermal reaction between nitrogen dioxide and acetaldehyde in the gas phase was investigated at room temperature and atmospheric pressure. The initial rate of disappearance of nitrogen dioxide was 1.00 ± 0.03 order with respect to nitrogen dioxide and 1.00 ± 0.07 order with respect to acetaldehyde. An initial second order rate constant of (8.596 ± 0.189) x 10^-3 1.mole^-1 sec^-1 was obtained at 22.0 ± 0.1 °C and a total pressure of one atmosphere. The activation energy of the reaction was 12,900 cal/mole in the temperature range between 22°C and 122°C.

The products of the reaction were nitric oxide, carbon dioxide, methyl nitrite, nitromethane and a trace amount of trans-dimeric nitrosomethane. The addition of nitric oxide increased the rate of formation of nitromethane and decreased the rate of formation of methyl nitrite. There were no measurable surface effects due to the addition of glass wool or glass beads to the reactor.

Reactants and products were analyzed by gas chromatography. A mechanism was proposed incorporating the principal features of the reaction.

Deep near-infrared spectroscopy of high-redshift galaxies: the physical growth of passive systems

The assembly history of massive galaxies is one of the most important aspects of galaxy formation and evolution. Although we have a broad idea of what physical processes govern the early phases of galaxy evolution, there are still many open questions. In this thesis I demonstrate the crucial role that spectroscopy can play in a physical understanding of galaxy evolution. I present deep near-infrared spectroscopy for a sample of high-redshift galaxies, from which I derive important physical properties and their evolution with cosmic time. I take advantage of the recent arrival of efficient near-infrared detectors to target the rest-frame optical spectra of z > 1 galaxies, from which many physical quantities can be derived. After illustrating the applications of near-infrared deep spectroscopy with a study of star-forming galaxies, I focus on the evolution of massive quiescent systems.

Most of this thesis is based on two samples collected at the W. M. Keck Observatory that represent a significant step forward in the spectroscopic study of z > 1 quiescent galaxies. All previous spectroscopic samples at this redshift were either limited to a few objects, or much shallower in terms of depth. Our first sample is composed of 56 quiescent galaxies at 1 < z < 1.6 collected using the upgraded red arm of the Low Resolution Imaging Spectrometer (LRIS). The second consists of 24 deep spectra of 1.5 < z < 2.5 quiescent objects observed with the Multi-Object Spectrometer For Infra-Red Exploration (MOSFIRE). Together, these spectra span the critical epoch 1 < z < 2.5, where most of the red sequence is formed, and where the sizes of quiescent systems are observed to increase significantly.

We measure stellar velocity dispersions and dynamical masses for the largest number of z > 1 quiescent galaxies to date. By assuming that the velocity dispersion of a massive galaxy does not change throughout its lifetime, as suggested by theoretical studies, we match galaxies in the local universe with their high-redshift progenitors. This allows us to derive the physical growth in mass and size experienced by individual systems, which represents a substantial advance over photometric inferences based on the overall galaxy population. We find a significant physical growth among quiescent galaxies over 0 < z < 2.5 and, by comparing the slope of growth in the mass-size plane dlogR_e/dlogM_∗ with the results of numerical simulations, we can constrain the physical process responsible for the evolution. Our results show that the slope of growth becomes steeper at higher redshifts, yet is broadly consistent with minor mergers being the main process by which individual objects evolve in mass and size.

By fitting stellar population models to the observed spectroscopy and photometry we derive reliable ages and other stellar population properties. We show that the addition of the spectroscopic data helps break the degeneracy between age and dust extinction, and yields significantly more robust results compared to fitting models to the photometry alone. We detect a clear relation between size and age, where larger galaxies are younger. Therefore, over time the average size of the quiescent population will increase because of the contribution of large galaxies recently arrived to the red sequence. This effect, called progenitor bias, is different from the physical size growth discussed above, but represents another contribution to the observed difference between the typical sizes of low- and high-redshift quiescent galaxies. By reconstructing the evolution of the red sequence starting at z ∼ 1.25 and using our stellar population histories to infer the past behavior to z ∼ 2, we demonstrate that progenitor bias accounts for only half of the observed growth of the population. The remaining size evolution must be due to physical growth of individual systems, in agreement with our dynamical study.

Finally, we use the stellar population properties to explore the earliest periods which led to the formation of massive quiescent galaxies. We find tentative evidence for two channels of star formation quenching, which suggests the existence of two independent physical mechanisms. We also detect a mass downsizing, where more massive galaxies form at higher redshift, and then evolve passively. By analyzing in depth the star formation history of the brightest object at z > 2 in our sample, we are able to put constraints on the quenching timescale and on the properties of its progenitor.

A consistent picture emerges from our analyses: massive galaxies form at very early epochs, are quenched on short timescales, and then evolve passively. The evolution is passive in the sense that no new stars are formed, but significant mass and size growth is achieved by accreting smaller, gas-poor systems. At the same time the population of quiescent galaxies grows in number due to the quenching of larger star-forming galaxies. This picture is in agreement with other observational studies, such as measurements of the merger rate and analyses of galaxy evolution at fixed number density.