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Dynamic Regulations of Co-Translational Protein Targeting by the Signal Recognition Particle Receptor in E. coli and Human
Get PDFThe Signal Recognition Particle (SRP) and its receptor (SR) co-translationally deliver the majority of secretory and membrane proteins to their membrane destinations. SRP recognizes and binds to the cargo, and SR recruits the transit complex to the target membrane. Precise timing and coordination are encoded into the interactions between SRP and SR in response to the cargo and environment to enable efficient and accurate targeting. This dissertation explores the regulation mechanisms of SRP membrane recruitment by SR in both bacteria and eukaryotes.
The bacterial SR is a peripheral membrane protein and recruits SRP by adjusting its membrane-binding modes. A complete kinetic model of SRP membrane recruitment by the SR was established based on direct observations and quantitative analyses of protein-membrane binding events at single-molecule resolution. Biochemical and cell biological examinations validated the physiological significance of the newly discovered dynamic membrane-binding mode of the SR. The two-stage SR membrane binding mechanism ensures both speed and specificity in bacterial co-translational protein targeting.
In eukaryotes, a proper initial recognition of SRP sets the right course for membrane delivery of the transit complex, as eukaryotic SR is anchored on the endoplasmic reticulum. A Molecular Recognition Feature (MoRF) in the disordered linker domain of eukaryotic SR was identified and characterized. The SR MoRF element enables rapid SRP-SR assembly in response to the ribosomal content of the cargo. The stimulation by SR MoRF is only present in eukaryotes and is likely a functional replacement of the tetraloop in bacterial SRP RNA whose stimulatory role was abolished during evolution.</p
Measurement of R(D) and R(D*) Using Semileptonic Tags and Hadronic τ Decays
Get PDFWe present a measurement of R(D(*)) = B(B → D̅(*) τντ)/B(B → D̅(*) ℓνℓ) using semileptonic tagging and hadronic τ decays on the 433 fb-1 of data collected at the Υ(4S) resonance using the BaBar detector at the PEP-II collider. We obtain a high statistics data sample using loose selection criteria. The signal is extracted by performing a 2-dimensional fit of the component densities to the kernel density estimate of the data, which is made computationally tractable by algorithmic improvements and speedups provided by graphics processing units. We obtain two distinct central values based on the model used to represent the BB̅ background densities: R(D) = 0.231 ± 0.028 ± 0.028 and R(D*) = 0.127 ± 0.019 ± 0.031 with a correlation of 0.06 and R(D) = 1.454 ± 0.028 ± 0.028 and R(D*) = 1.507 ± 0.019 ± 0.031 with a correlation of 0.06. The region encompassed by the two results are consistent with both the Standard Model prediction and the world average.</p
Computational Methods for Gravitational Wave Physics: Spectral Cauchy-Characteristic Extraction and Tidal Splicing
Get PDFAs the aLIGO and Virgo detectors continue to improve their sensitivity for observing gravitational waves from merging compact binaries, they will require ever more precise theoretical predictions to extract a detailed understanding of the physics governing these merging systems. This thesis discusses advancements within computing the gravitational waveforms along two avenues of research: the continued development of a spectral Cauchy-Characteristic Extraction (CCE) code and the presentation of a novel method called 'Tidal Splicing' for generating waveforms for binary neutron star (BNS), black hole-neutron star (BHNS), and even Beyond GR systems.
Due to the finite extents of typical 3+1 simulations of merging binaries, the waveforms they generate can suffer from near-zone effects and lingering gauge ambiguities. CCE was developed in order evolve radiating gravitational waves as they propagate outward to future null infinity, allowing studies connecting the dynamical spacetime of binary evolutions to effects seen by distant observers, such as superkicks, and angluar and linear momentum fluxes. A recent spectral version of CCE showed promising improvements in accuracy and efficiency over the older finite-differencing code, PittNull. However, lingering issues with the numerics and implementation of the theory prevented it from wide spread use. We detail the developments updated its initial release and demonstrate the enhancement in accuracy they yield beyond the capabilities of PittNull.
The method of Tidal Splicing enhances the inexpensive Post-Newtonian (PN) tidal corrections with BBH waveforms from numerical simulations to generate waveforms corresponding to inpsiraling BNS or BHNS systems. This leverages the accuracy of numerical BBH waveforms to effectively replace the corresponding unknown PN terms. In addition, by picking individual terms in the PN tidal expansions to include, then comparing with existing numerical simulations, we are able to probe the significance of each contribution to the total difference in evolution between BBH and BNS or BHNS inspirals. We also demonstrate how the splicing concepts used for tidal effects can extended in order to model waveforms with corrections according to theories beyond GR using an example case of a resonating ultra-compact object.</p
Passive and Active Control of Radiative Heat Flow
Get PDFMaterials that control the absorption and emission of thermal radiation have attracted renewed interest for energy applications. Materials of interest include those with static optical properties that vary with photon wavelength in a desired manner as well as those with dynamic properties that can be actively tuned by external stimuli. The research in this thesis focuses on creating materials in both categories.
First, we examine selective absorbers for solar thermal energy conversion with high absorptivity in solar wavelengths and low emissivity in infrared wavelengths. Achieving stagnation temperatures exceeding 200 °C with unconcentrated sunlight, pertinent to technologies like industrial process heat, air conditioning, and electricity generation, requires better spectrally selective absorbers with ultra-low thermal emittance. Current state-of-art surfaces are based on ceramic-metal mixtures and patterned metal or metal-dielectric structures. Semiconductor based selective surfaces with near zero absorption below the bandgap offer the potential for lower thermal emittance than that achieved with such surfaces that employ metals in the primary absorbing medium. In this thesis, we report a semiconductor-based multilayer selective absorber that exploits the sharp drop in optical absorption at the band gap energy to achieve a measured absorptance of 76% at solar wavelengths and a low emittance of approximately 5% at thermal wavelengths. In field tests, we obtain a peak temperature of 225 °C, comparable to that achieved with state-of-the-art selective surfaces. With straightforward optimization to improve solar absorption, our work shows the potential for unconcentrated solar thermal systems to reach stag- nation temperatures exceeding 300 °C, higher than any available selective surface. Our surface would eliminate the need for solar concentrators for mid-temperature solar applications such as supplying process heat.
Second, we theoretically propose and experimentally implement a thermal switch for near-field radiative transfer. In the field of active thermal materials for manipulating heat flow in a controllable and reversible manner, numerous approaches to perform thermal switching have been reported. However, they typically suffer from various limitations, including small switching ratio or requiring large temperature differentials. We report the experimental implementation of a scheme to electrostatically control near-field radiative transfer in a graphene field effect heterostructure. We measure a maximum heat flux modulation of 4 ± 3% and an absolute heat flux modulation rate of 24 ± 7 mWm−2 per V bias. Employing gate dielectrics with lower surface warp and higher dielectric breakdown strength as well as reducing conductive losses would enable modulations up to 100%, substantially exceeding the switching ratios achievable by other methods. Our work paves the way for electrostatic control of near-field radiative transfer using two-dimensional materials.</p
Synthesis, Characterization, and Reactivity of "Reverse Pyridine" Bis(phosphinite) Pincer Complexes
Get PDFIn chapter 1 the synthesis of several pincer complexes based on a "reverse pyridine" bis(phosphinite) ligand motif is described. Coordination of a borane Lewis acid to the parent ligand is described, as well as factors influencing the subsequent metalation of both the "free base" ligand and borane-adduct ligand. The effect of Lewis acid coordination on the spectroscopic and electrochemical properties of the resulting metal complexes is discussed. The reversibility of Lewis acid binding for some complexes is also described.
In chapter 2, an unusual method for carbon-oxygen and carbon-sulfur bond cleavage mediated by alkoxide salts and using silanes as reductants is described.</p
Analysis on Vector Bundles over Noncommutative Tori
Get PDFNoncommutative geometry is the study of noncommutative algebras, especially C*-algebras, and their geometric interpretation as topological spaces. One C*-algebra particularly important in physics is the noncommutative n-torus, the irrational rotation C*-algebra AΘ with n unitary generators U1, . . . , Un which satisfy UkUj = e2πiθj,kUjUk and Uj* = Uj-1, where Θ ∈ Mn(ℝ) is skew-symmetric with upper triangular entries that are irrational and linearly independent over ℚ. We focus on two projects: an analytically detailed derivation of the pseudodifferential calculus on noncommutative tori, and a proof of an index theorem for vector bundles over the noncommutative two torus. We use Raymond's definition of an oscillatory integral with Connes' construction of pseudodifferential operators to rederive the calculus in more detail, following the strategy of the derivations in Wong's book on pseudodifferential operators. We then define the corresponding analog of Sobolev spaces on noncommutative tori, for which we prove analogs of the Sobolev and Rellich lemmas, and extend all of these results to vector bundles over noncommutative tori. We extend Connes and Tretkoff's analog of the Gauss-Bonnet theorem for the noncommutative two torus to an analog of the McKean-Singer index theorem for vector bundles over the noncommutative two torus, proving a rearrangement lemma where a self-adjoint idempotent e appears in the denominator but does not commute with the k2 already there from the rearrangement lemma proven by Connes and Tretkoff.</p
New Insights into Circumstellar Disk Evolution in the Era of ALMA
Get PDFIn recent decades, it has become clear that solar systems such as our own form within the circumstellar disks of gas and dust that surround young stars. Thus, to understand how these solar systems come to be, it is necessary to study the conditions within these disks. Until recently, such studies have required a focus on intrinsically brighter and younger disks that are easier to observe. However, a full picture of planet formation requires the characterization of older disks to determine how these systems change over time. The unprecedented capabilities of the Atacama Large Millimeter/submillimeter Array (ALMA) now present the opportunity to study populations of these disks in detail for the first time. In this thesis, I present a study of over 100 such disks in the 5-11 Myr old Upper Scorpius OB Association (Upper Sco) using ALMA, as well as Keck Observatory, with the aim of comparing the properties of these systems to younger disks in order to shed new light on disk evolution.
Following background discussion on disks and their evolution, ALMA measurements of the continuum and CO line fluxes of these disks at 0.88 mm are reported in Chapter 3. The continuum fluxes are used to show that the majority of these systems contain less than 1 M⊕ of dust. It is then shown that dust masses around these stars are on average a factor of ∼4.5 lower than their younger counterparts in the Taurus star-forming region, placing important constraints on the mass evolution of these systems. Finally, constraints on depletion of gas in these disks are discussed using the CO measurements.
The spatial distributions of the gas and dust within these Upper Sco disks are modeled in Chapter 4. The radial extents of gas and dust are measured and compared, with several systems showing evidence of the gas being more extended. The sizes of the dust disks are compared to younger systems, showing that these disks shrink by a factor of approximately three as they age. These results suggest that dust evolves from the outside-in within disks, perhaps through radial drift. Despite this evolution, dust disks in Upper Sco fall on the same correlation between size and millimeter luminosity as their younger counterparts. This implies a link between the radial structures of disks of different ages, perhaps indicating that these systems are composed of optically-thick dust substructure.
Of course, an understanding of planetary system formation would be incomplete without accounting for the presence of stellar companions, which are common around young stars and are expected to shorten disk lifetimes by truncating their sizes. As such, Chapter 5 presents a search for stellar companions in the Upper Sco disk sample analyzed in Chapters 3 and 4. Using adaptive optics imaging and aperture masking observations with the NIRC2 instrument on the Keck II telescope, stellar companions are identified in 27 of 112 systems. It is then shown that the companion fraction of systems with disks is lower than those without, confirming the harmful effects of stellar companions on disks seen in younger systems. However, the fraction of disk systems in Upper Sco with a close companion is shown to match that of younger disks in Taurus. This indicates that these effects occur within the first ∼Myr of disk evolution, after which stellar companions have little to no effect. Additionally, while the millimeter luminosities of disks with stellar companions are observed to be lower than those around single stars in Taurus, there exists no such difference in Upper Sco. This provides further support for outside-in dust evolution, as the shrinking of disks around single stars would cause them by the 5-11 Myr age of Upper Sco to match the sizes and brightnesses of truncated disks in binary systems.
Taken together, the results presented in this thesis show the masses and radial extents of the dust-component of circumstellar disks decrease with age. This thesis concludes by summarizing these results and discussing their link within a scenario of outside-in dust evolution involving radial drift and dust substructure. To close, potential avenues are presented to continue the study of disk evolution with ALMA.</p
Optical Imaging of Dopamine Dynamics and Decoding its Role in Arousal and Salience
Get PDFDopamine (DA) is a key neuromodulator in the brain that can exert a profound impact on brain physiology and cognitive functions. There is consensus that DA plays critical roles in reward prediction error, reinforcement learning, and motor control, and that dysregulation of DA signaling is common in many neuropsychiatric diseases, such as Parkinson’s disease, drug addiction, and depression. Although the tools to study the functional roles of DA have considerably expanded with novel genetic tools and optical imaging methods, we are still limited in our ability to record or visualize DA release in vivo with long-term stability and high spatiotemporal resolution. This is an unmet need in DA research, as DA release at the post-synaptic sites can be decoupled from DA cell body firing due to local circuit interaction and influence from other afferent activities. In parallel, there is growing evidence that DA is functionally heterogeneous beyond its classically described roles for reward and movement, based on its anatomical location, projection target, electrophysiological properties, and response patterns to stimuli with motivational valence. Pharmacological and genetic studies have provided indirect evidence that DA can promote strong behavioral arousal and signal salience, but the precise neural substrates for these functions remain largely unknown. Towards this end, my thesis work has been focused on 1) developing and characterizing optical tools to visualize DA release in vivo and 2) utilizing such optical and genetic tools to study the overlooked, sparse DA populations in the dorsal midbrain, demonstrating that they are functionally unique DA cells for broadcasting arousal and salience signals to the forebrain targets.
As neuromodulatory systems exert profound influences on brain function, understanding how these systems modify the operating mode of target circuits requires spatiotemporally precise measurement of neuromodulator release. Towards this goal, in Chapter II, my colleagues and I developed dLight1, an intensity-based genetically encoded DA indicator, to enable optical recording of DA dynamics with high spatiotemporal resolution in behaving mice. We demonstrated the utility of dLight1 by imaging DA dynamics simultaneously with pharmacological manipulation, electrophysiological or optogenetic stimulation, and calcium imaging of local neuronal activity. dLight1 enabled chronic tracking of learning-induced changes in millisecond DA transients in mouse striatum. Further, we used dLight1 to image spatially distinct, functionally heterogeneous DA transients relevant to learning and motor control in mouse cortex. We also validated our sensor design platform for developing norepinephrine, serotonin, melatonin, and opioid neuropeptide indicators. Together, this tool provides a unique opportunity to optically monitor DA release dynamics in vivo with long-term stability and unprecedented spatiotemporal resolution.
In Chapter III, I have characterized the functional roles of sparse DA populations in the dorsal raphe nucleus (DRN) and discovered that these neurons play key roles in promoting behavioral arousal. I first demonstrated that DRNDA neurons are activated by diverse forms of motivationally salient stimuli, irrespective of valence. Simultaneous fiber photometry and polysomnographic recordings showed that DRNDA neuronal activity is correlated with distinct sleep-wake states, showing highest activities during wakefulness over sleep states. Optogenetic activation of DRNDA neurons was sufficient to cause immediate sleep-to-wake transitions and promote longer wakefulness upon sustained stimulation. On contrary, DRNDA inhibition via chemogenetics reduced wakefulness and promoted non-rapid eye movement sleep, even in the presence of ethologically relevant salient stimuli. Taken together, this pinpoints DRNDA neurons as the critical contributor of arousal-promoting DA system in the brain.
In Chapter IV, I further characterized the encoding dynamics of DRNDA neurons during classical conditioning tasks where mice learned the association between neutral cues and outcomes with positive or negative outcomes. DRNDA neurons developed phasic, positive responses to cues predicting both positive and negative unconditioned stimuli across learning, suggesting that these populations track motivational salience rather than valence. In addition, DRNDA neurons encoded unsigned prediction error, demonstrating higher neuronal activity to unexpected reward or punishment over fully expected outcomes. Collectively with Chapter III, these results expand on the existing literature on functionally heterogeneous roles of DA in the brain and propose that DRNDA neurons play critical roles in signaling arousal and motivational salience to the forebrain regions to coordinate appropriate behavior, depending on the nature of environmental stimuli.</p
mRNA Splicing-Mediated Gene Expression Regulation in Innate Immunity
Get PDFAt the heart of an inflammatory response lies a tightly regulated gene expression program. Perturbations to this finely tuned response can result in unchecked or inappropriately scaled inflammation, shifting the balance from protective to destructive immunity. A variety of post-transcriptional mechanisms play a role in the fine-tuning of an inflammatory gene expression program. One such mechanism involves unproductive RNA splicing, whereby alternative splicing can frameshift the transcript or introduce a premature termination codon (PTC). These effects render the transcript nonfunctional and/or subject it to nonsense-mediated decay.
We observed such an event in Irf7, the master regulator of the type I interferon response. We found a single intron was consistently retained at a level much greater than other introns in the Irf7 transcript. In an effort to understand trans-acting factors that regulate this retention, we used RNA-antisense purification followed by mass spectrometry (RAP-MS) to identify the factor BUD13 as a highly enriched protein on Irf7 transcripts. Deficiency in BUD13 was associated with increased retention, decreased mature Irf7 transcript and protein levels, and consequently a dampened type I interferon response, which compromised the ability of BUD13-deficient macrophages to withstand vesicular stomatitis virus (VSV) infection.
Beyond this intron retention event in Irf7, we identified a variety of other unproductive splicing events in a number of important genes involved with the innate immune response. This unproductive splicing was not restricted to intron retention events. For example, we identified a frequently used alternative splice site in the crucial murine antiviral response gene, oligoadenylate synthetase 1g (Oas1g) that led to both a frameshift and incorporation of a PTC. Genome editing was used to remove the alternative splice site in a macrophage cell line, which led to both increased Oas1g expression and improved viral clearance. We hypothesize these events exist as a means of mitigation for what might otherwise be an inappropriately scaled response. In doing so, they represent a previously underappreciated layer of gene expression regulation in innate immunity.</p
Dynamics, Mechanics and Stability of Physical Gels
Get PDFFrom the commercial products that we encounter in our daily lives to the mucous that lines our gut, gels assembled by the reversible association of polymers or colloids are a ubiquitous, important and fascinating class of soft materials. The dual solid and fluid-like (viscoelastic) properties of associative polymer gels render them useful in a number of applications including as tissue-regeneration scaffolds, drug delivery vectors and organic electronics and batter technologies. However, there remains a number of open questions regarding the microscopic origins of many of the dynamical and mechanical properties that make these materials so appealing. The wide range of length and timescales in physical gels present a formidable challenge towards the formulation of a complete microscopic dynamical and rheological portrait. My work has focused on the development of microscopically-informed and experimentally verifiable explanations for some of the fundamental dynamical and mechanical properties of associative gels. I first present our viewpoint, informed by computer simulation and experiment, on the origin of the long-time self-diffusivity of telechelic polymer gels. Our perspective and resulting theory compare favorably with experiments. Shearing an associative polymer gel is found to result in the emergence of new diffusive modes with applied shear that are can destabilize homogeneous flow for gels sufficiently close to the two phase boundary. This finding motivates the idea that nonequilibrium forcing may promote the relaxation of arrested colloidal materials, such as a colloidal gel, closer to their thermodynamic ground state. The driving force need not be externally applied. The induced collective motion in colloidal gels subject to internal driving forces (such as the presence of a small fraction of self-propelling colloids) can drive the system from a state of arrested metastablity to a state of lower free energy. I conclude by showing that the internal stress generated by the self-propelling particles -- the active stress -- is not a "true" stress, but rather an equivalent stress analogous to the dynamic pressure of fluids in a gravitational field. The importance of this finding is demonstrated in resolving the perplexing finding of a negative surface tension in phase separated active materials.</p