VictorJuiz/thesis huggingface dataset:
Dimerization quality control as a molecular mechanism for ensuring proper complex formation

Cellular signaling relies on many proteins that form dimers and the correct pairing of each dimer is critical for their activity. Despite the fact that there are numerous recurrent and modular dimerization domains, including BTB domains and leucine zippers, whether cells have mechanisms to detect and eliminate dimers of aberrant composition is not known. Here, by searching for regulators of the BTB family, we discovered a quality control pathway that ensures functional dimerization [dimerization quality control (DQC)]. Key to this network is the E3 ligase SCF-FBXL17, which selectively binds and ubiquitylates BTB dimers of aberrant composition to trigger their clearance by proteasomal degradation. Underscoring the physiological importance of DQC, SCF-FBXL17 is required for the differentiation, function, and survival of neural crest and neuronal cells in Xenopus laevis. Thus, metazoan organisms actively monitor BTB dimerization./nAfter identifying DQC, one key question that remained was how a single enzyme, SCF-FBXL17, could recognize hundreds of aberrant heterodimers but leave active homodimers intact. In order to understand how DQC can distinguish complexes based on their composition, we used cryo-electron microscopy and X-ray crystallography to determine structures of aberrant BTBs alone and bound to SCF-FBXL17. We find that SCF-FBXL17 encircles a monomeric BTB domain through a multipartite interface that can accommodate significant sequence variation. Distinct from most quality control systems, SCF-FBXL17 does not detect protein misfolding, but probes dimer stability through a domain-swapped and rapidly evolving β-strand within BTB domains. Our findings suggest that SCF-FBXL17 uses a mechanism of subunit exchange to establish quality control of complex composition, and we predict that other quality control proteins may similarly monitor other recurrent oligomerization domains.

Advisor: Rape, Michael
Something new under the sun: transposon sequencing and phylogenomics shed light on unstudied photosynthetic genes

The core machinery required for oxygenic photosynthesis is well conserved from cyanobacteria to land plants and in various algal clades, and has been studied intensely for decades. Much remains to be learned, however, both about the regulation and assembly of the core machinery, and about a host of optional factors associated with it only in particular lineages or under particular conditions. I sought to identify unstudied genes required for long-term acclimation to high light intensity in the cyanobacterium Synechococcus elongatus PCC 7942 by simultaneously comparing the growth rates of mutants in every nonessential gene in the genome under low, normal, and high light via RB-TnSeq (growth of a pooled transposon library, followed by sequencing and quantification of DNA barcodes). This was the first study to investigate low and high light conditions with high enough resolution to as- sign fitness values to nearly every gene. It confirmed that high light mainly damages the Photosystem II reaction center, and that synthesis and recycling of chlorophyll from damaged PSII subunits is essential for acclimation. It also identified several unexpected genes with large fitness effects. The include a mys- terious intrinsically disordered protein with homology to phasins, an unusual transcriptional regulator that may physically protect DNA from oxidative damage, and an ABC transporter that takes up amino acids. Next I grew the transposon library under a range of fluctuating light conditions to determine which genes are important for fitness only when fluctuations include higher highs or lower lows, alternate quickly or slowly, include gradual or sudden low-to-high transitions, or mimic the patterns found in dense bioreactor cultures. Many genes were beneficial only under certain conditions but not others, and some were also found to be detrimental (knocking them out increased fitness) under certain con- ditions. The genes identified are implicated in diverse cellular processes relevant to photosynthesis, in- cluding: the stringent response to dark periods, alternative electron sinks (plastoquinone reduction, glycogen synthesis) needed during sudden shifts to high light, and regulation of transcription. As part of a separate line of inquiry into genes conserved across photosynthetic species but missing from other branches of life, I also developed a scripting language that partially automates comparisons between large numbers of genomes using a variety of popular sequence search and ortholog-finding programs: BLAST, BLAST reciprocal best hits, HMMER, DIAMOND, MMSeqs2, OrthoFinder, SonicParanoid, and others. It reproducibly installs all the programs and runs them as needed to per- form searches as described in a concise, human-readable domain-specific language designed to facilitate sharing and incremental improvement of the search algorithms.

Advisor: Niyogi, Krishna
Realization of Integrated Coherent LiDAR

LiDAR (Light Detection and Ranging) captures high-definition real-time 3D images of the surrounding environment through active sensing with infrared lasers. It has unique advantages that can compensate the fundamental limitations in camera-based 3D imaging via vision algorithms or RADARs, which makes it an important sensing modality to guarantee robust autonomy in self-driving cars. However, high price tag of existing commercial LiDAR modules based on mechanical beam scanners and intensity-based detection scheme makes them unusable in the context of mass produced consumer products./nThe focus of thesis is on the integrated coherent LiDAR with optical phased array-based solid-state beam steering, which has great potential to dramatically bring down the cost of a LiDAR module. It begins with an overview of LiDAR implementation options and system requirements in the context of autonomous vehicles, which leads us to conclude that beam-steering coherent FMCW LiDAR in optical C-band is indeed the best implementation strategy to realize low-cost automotive LiDARs. Motivated by this observation, a quantitative framework for evaluating FMCW LiDAR performance is also introduced to predict the design that satisfies car-grade performance requirements. Then the thesis presents the silicon implementation results from our single-chip optical phased array and integrated coherent LiDAR prototype. Our implementations leverage the 3D heterogeneous integration platform, where custom silicon photonics and nanoscale CMOS fabricated at a 300 mm wafer facility are combined at the wafer-scale to minimize the unit cost without I/O density issues. After discussing remaining challenges and possible ways to enhance the operating range and system reliability, this thesis finally addresses the problem of fundamental trade-off between phase noise and wavelength tuning in FMCW laser source, and present circuit- and algorithm-level techniques to enable FMCW measurements beyond inherent laser coherence range limit.

Advisor: Stojanovic, Vladimir
Hybrid Aesthetics: Bridging Material Practices and Digital Fabrication through Computational Crafting Proxies

Creative technologies like digital fabrication led to the rise of the Maker movement, engendering grassroots innovation in education, manufacturing, and healthcare. Today, these creative technologies stand at a crossroads – despite a significant rise in participation, a deeper engagement with design and material is absent from traditional computer-aided design workflows. In this thesis, I will motivate the need for creative technologies to support the morphogenetic model of making, a thinking and working style characteristic of how practitioners work with physical materials but difficult to access in digital design tools. To communicate my findings, I introduce the concept of a Crafting Proxy, an intermediary between a practitioner and a material that can be used to facilitate the interpretation, manipulation, and evaluation of a material as a part of a creative process. In these works, I employ a Research through Design (RtD) methodology to construct intermediate-level knowledge around the design, implementation, and evaluation of Crafting Proxies. I’ll demonstrate how Crafting Proxies can be enacted within physical materials, physical tools, and physical practices to support morphogenetic workflows in domains such as light and heater design, and metalworking. As a result, this work contributes a design method for creating crafting proxies and a set of design principles that inform how new materials and digital fabrication technologies can foreground the existing knowledge and practices of material practitioners and generate new forms and aesthetics that can alter the trajectory of the Maker movement towards a New Making Renaissance.

Advisor: Paulos, Eric
High-dimensional and causal inference

High-dimensional and causal inference are topics at the forefront of statistical research. This thesis is a unified treatment of three contributions to these literatures. The first two contributions are to the theoretical statistical literature; the third puts the techniques of causal inference into practice in policy evaluation./nIn Chapter 2, we suggest a broadly applicable remedy for the failure of Efron’s bootstrap in high dimensions is to modify the bootstrap so that data vectors are broken into blocks and the blocks are resampled independently of one another. Cross-validation can be used effectively to choose the optimal block length. We show both theoretically and in numerical studies that this method restores consistency and has superior predictive performance when used in combination with Breiman’s bagging procedure. This chapter is joint work with Peter Hall and Hugh Miller./nIn Chapter 3, we investigate regression adjustment for the modified outcome (RAMO). An equivalent procedure is given in Rubin and van der Laan [2007] and then in Luedtke and van der Laan [2016]; philosophically similar ideas appear to originate in Miller [1976]. We establish new guarantees when the procedure is applied in designed experiments (where the propensity score is known a priori) and confirm that the procedure is doubly robust. RAMO can be implemented in only a few lines of code and it can be immediately combined with existing regression models, including random forests and deep neural networks, used in classical prediction problems. This chapter is joint work with Bin Yu and Jasjeet Sekhon./nIn Chapter 4, we investigate the specific deterrent effect of traffic citations. In Queensland, Australia many speeding and red-light running offenses are detected by traffic cameras and drivers are notified of the citation, not at the time they commit the offense, but when the citation notice is delivered by mail about two weeks later. We use a regression discontinuity design to assess whether the chance of crashing or recidivism changes at the moment of notification. We analyzed a population of nearly 3 million drivers who committed camera-detected offenses. We conclude that there is not a significant change in the incidence of crashes but there is a marked decrease in recidivism of about 25%. This chapter is joint work with David Studdert and Jeremy Goldhaber-Fiebert.

Advisor: Yu, Bin
Sekhon, Jasjeet S
Mechanisms of mTORC1 signal regulation by the Rag GTPases

The decision of whether to allocate resources toward cellular growth or toward quality control is a matter of cellular life and death; disruption of growth pathways is an emerging driving force in diseases ranging from cancer to neurodegeneration. In mammalian cells, the protein kinase activity of mTORC1 promotes cellular anabolism and impedes cellular catabolism, ultimately achieving a balance that dictates the rate of cell growth. In response to nutrient levels, mTORC1 is activated upon recruitment to the lysosome, an organelle whose role as a nutrient sensing integrator has recently come into focus. The Rag GTPases are required for mTORC1 recruitment to the lysosome, but the mechanisms via which the Rags sense nutrients and precisely couple the degree of mTORC1 lysosomal recruitment levels to the level of available nutrients were unknown./nI report new live-imaging and reconstitution approaches that enabled the discovery that when the Rags transition from their inactive nucleotide binding state to their active nucleotide state in response to nutrient stimulation, they also loosen their binding affinity for their lysosomal scaffold, Ragulator. The resulting spatial cycling between the lysosome and the cytoplasm ultimately limits mTORC1 accumulation on its Rag-Ragulator lysosomal scaffold, and promotes rapid responsiveness of mTORC1./nNext, I asked whether the nucleotide states of the two Rag GTPase domains are coordinated. Prior work had established that a complex of Folliculin (FLCN) and FLCN-interacting protein 2 (FLCN:FNIP2) serves as a RagC-specific GTPase-activating protein (GAP) and thus has a positive role in mTORC1 stimulation. However, genetic evidence placed FLCN as a tumor suppressor, suggesting a negative role. I reconstituted the “Lysosomal Folliculin Complex” (LFC), a supercomplex composed of Ragulator, inactive-loaded Rags, and FLCN:FNIP2 that localizes to lysosomes. I discovered that in the LFC, FLCN:FNIP2 clamps Rags in their inactive state (RagAGDP:RagCGTP) by directly inhibiting nucleotide exchange in RagA, concomitant with inhibition of its RagC GAP activity, a conclusion reinforced by a high-resolution (3.6 Å) structure of the LFC. Thus, when nutrients are low, FLCN:FNIP2 is able to maintain the Rag heterodimer in its inactive state, but, in response to a rise in nutrients, FLCN:FNIP2 is converted into a functional GAP./nFinally, by assessing newly available structures of active nucleotide-bound and inactive nucleotide-bound Rag heterodimers, along with recent structural information about Rag interactors, I was able to assemble an integrated structure-guided model of the Rag-mediated cycle of recruitment and activation of mTORC1 at the lysosome. My findings increase our understanding of the molecular logic of nutrient sensing and point to new opportunities for manipulating mTORC1 signaling in disease contexts.

Advisor: Zoncu, Roberto
Investigating the roles of ribosome-associated proteins in translational regulation of gene expression

The ribosome has long been regarded as the protein producing center of the cell, yet we are still defining its role as a target of gene regulation in the cell. Towards this end, I developed a proteomics-based approach to uncover novel ribosome-associated proteins that regulate translation in the cell. I also worked with the Kopito Lab at Stanford University to understand why a core ribosomal protein, RPL26, is post-translationally modified by a ubiquitin-like protein, UFM1. Additionally, I collaborated with the Corn Lab at UC Berkeley and ETH Zurich to describe how double-stranded DNA damage leads to the depletion of core ribosomal proteins, RPL40 and RPS27A. These projects demonstrate that ribosome composition is neither static nor homogeneous but can be altered depending on the environmental and cellular context.

Advisor: Ingolia, Nicholas
Corn, Jacob
Investigation of terminal alkene formation by acyl-CoA dehydrogenases in the biosynthesis of complex natural products

Polyketides are a class of natural products known for their chemical complexity and bioactive properties. They are biosynthesized by an assembly line-like system termed polyketide synthases (PKSs), in which each module consists of various enzymatic domains, each of which has a single catalytic function. The polyketide is extended by two carbons and selectively reduced by each module. These biosynthetic principles make PKSs attractive engineering targets, because a simple swap of one enzymatic domain could change the final polyketide structure and effect an improved or novel bioactivity. However, gaping holes in our understanding of polyketide biosynthesis still exist. In particular, the activities of polyketide-associated enzymes remain relatively under-characterized and are generally not the primary focus of engineering efforts. These enzymes, which include glycosyl transferases, cytochrome P450s, and many others, often are responsible for imparting bioactive functional groups into the polyketide backbone or after the assembly of the “naked” polyketide. Many of these non-canonical functional groups discovered within polyketides are therefore the most desirable engineering targets. In this dissertation, I describe my efforts to characterize a family of terminal alkene-forming enzymes which were originally identified in a polyketide biosynthetic cluster. I show that the enzymes are regioselective and that regioselectivity is controlled by a shift in the protein structure. Finally, I show that these enzymes are widespread in not only polyketide biosynthetic gene clusters, but also in other natural product clusters in the genomes of diverse Actinomycetes.

Advisor: Keasling, Jay D
On Lyric's Minor Commons

On Lyric’s Minor Commons studies how minoritized writers use lyric poems to create alternative forms of collectivity. I argue that poets like Amiri Baraka, Frank O’Hara, M. NourbeSe Philip, and Jack Spicer use lyric poetry’s undetermined multiplicity of voicing and reading for various social and political aims. These poems become spaces for politics: they conjure and mobilize collectives of action and feeling. My argument complicates generic and historicist critiques that associate lyric voice with the reinforcement of humanism. While lyric is often considered to be the genre of individual subjective experience, I read lyric as the genre of the collective who can voice or read it. For the minoritized poets I discuss in this dissertation, lyric’s collective is not the unmarked hegemonic universal, but another commons. The politics of difference often emphasizes the individual relative to the collective; my readings show how the “black poem” or the “queer poem” offer a commons of their own. These poems are minor in their position vis-à-vis the unmarked hegemonic universal, and also minor in their alternative prefiguring of a different form of sociality without a coercive and majoritarian impulse. By calling these invocations commons rather than particularisms, I argue that these writers figure minoritized identities as forms of collectivity rather than fixed identifications./nI begin with a reconsideration of the Romantic lyric, a primal scene for the association of lyric with the unmarked and abstract subject. Through reading Dorothy Wordsworth’s Journals in apposition to some of William Wordsworth’s poems, I suggest a version of lyric reading that does not merely reproduce the subject, but instead exposes language to alternative reiteration through a contingent holding. Lyric’s attempt to capture experience in language, I argue, makes that experience into a commons. I then turn to more specific invocations by three mid-century U.S. poets, Amiri Baraka, Jack Spicer, and Frank O’Hara. Baraka and Spicer use lyric poems to call collective subjectivities—queerness and blackness, respectively—into being. Poems, for them, become utopian queer or black spaces par excellence: as Baraka writes, “Let the world be a black poem.” I complicate this utopian impulse by asking whether poems can offer security against despair. Across O’Hara’s bright sociality, and Spicer’s alienation, I find their poems offer various and contradictory sites of entry that account for how experiences of joy and negativity interfuse one another. The dissertation concludes with a discussion of M. NourbeSe Philip’s Zong!, whose commons is also internally differentiated, speculative, and without assurance. This poem, I argue, assembles a universality of aesthetic judgment as an ecstatic community that is radically indeterminate, yet posited nevertheless. Throughout these investigations I emphasize how the collective is not a lamentable or optional feature for these poets: rather than using lyric to plumb the depths of personal experience, they use it to imagine and create various emergent commons.

Advisor: Hejinian, Lyn
The Influence of Environmental Exposures on Glucocorticoid Receptor Signaling

Glucocorticoids (GCs) are hormones secreted in response to psychological stress. GCs have systemic effects on the endocrine, metabolic, cardiovascular, immune, reproductive, and central nervous systems. The physiological effects of GCs are mediated by the glucocorticoid receptor (GR), which is expressed in nearly every cell of the body. Environmental chemicals that disrupt GR signaling and/or cortisol homeostasis could adversely affect human health. A major challenge in identifying environmental chemicals that alter GR signaling in humans is a lack of adequate screening methods. The overall objective of this dissertation is to investigate the impact of environmental exposures on GR signaling and human GC levels. Chapter 1 discusses regulation of GC levels and presents evidence that environmental chemicals modulate GR signaling. Chapter 2 provides the field with a new bioassay to assess the endocrine disrupting effects of environmental chemicals on GR signaling. Results from this work demonstrate that this bioassay can be used to identify environmental chemicals that modulate GR activation and to screen serum samples for differences in total GC levels. Chapter 3 examines the persistent effect of early-life arsenic exposure on GC levels in a human population study. This is the first epidemiology study to investigate associations between early-life environmental exposures and GC levels in adulthood. Lastly, Chapter 4 evaluates prediction models of GR ligand mixtures. As a whole, this research aims to inform risk assessment of endocrine disrupting chemicals by developing methods to evaluate their impact on GR signaling and human health.

Advisor: Smith, Martyn T