Developing molecular tools for probing and modulating genomic spatial adjacency

In addition to the vast information encoded in DNA sequence, the genome has physical features that are also essential for its function, including its organization in threedimensional space. The development of high-throughput technology has greatly advanced our understanding of the spatial organization of the genome but has also raised more questions. In this thesis, we developed molecular tools to address the remaining challenges regarding the interplay between genomic organization and function. By breaking down the subject from the global architecture of the genome into an ensemble of spatially adjacent chromatin segments, we came up with different methods covering various aspects. We demonstrated in Paper I that global spatial information can be transferred in the format of DNA sequence encoding pairwise spatial proximity between two distinct molecular objects. We have shown that by growing network from pairwise relationship encoded in DNA sequence, spatial features at a global scale can be recovered. The results from this work highlighted the potential of using pairwise adjacency as a fundamental unit for recording the spatial organization of complex molecular systems. The high programmability and versatility of nucleic acids make them an ideal medium for encoding this information. With the aim of studying the pairwise relationship between genomic DNA in cells, we devised a CRISPR-dCas9 system for different purposes by leveraging its high programmability for genome targeting. In Paper III, we have shown that the re-designed guide RNA can direct dCas9 to a pair of genomic loci, inducing DNA contacts. This system can be applied as a modulation tool to introduce pairwise contacts for decoding functional implications in cells. In Paper IV, we developed a method for the direct detection of pairwise interactions between genomic loci at the single-cell level in situ. This method is achieved by conjugating oligonucleotide tags to Cas9 and using the tags for probing the spatial adjacency between a pair of genomic loci targeted by Cas9 Meanwhile, we developed an efficient method to fabricate and purify DNA origami with modifications in Paper II. This method makes the production of functionalized nanostructures more time and material-efficient compared to established techniques. The ease of production allows broader applications of functionalized nanostructures, including characterizing the effect of nanoscale distance on biochemical assays, as shown in Paper IV

R3^3 Prompting: Review, Rephrase and Resolve for Chain-of-Thought Reasoning in Large Language Models under Noisy Context

With the help of Chain-of-Thought (CoT) prompting, Large Language Models (LLMs) have achieved remarkable performance on various reasoning tasks. However, most of them have been evaluated under noise-free context and the dilemma for LLMs to produce inaccurate results under the noisy context has not been fully investigated. Existing studies utilize trigger sentences to encourage LLMs to concentrate on the relevant information but the trigger has limited effect on final answer prediction. Inspired by interactive CoT method, where intermediate reasoning steps are promoted by multiple rounds of interaction between users and LLMs, we propose a novel prompting method, namely R$^3$ prompting, for CoT reasoning under noisy context. Specifically, R$^3$ prompting interacts with LLMs to perform key sentence extraction, variable declaration and answer prediction, which corresponds to a thought process of reviewing, rephrasing and resolving. The responses generated at the last interaction will perform as hints to guide toward the responses of the next interaction. Our experiments show that R$^3$ prompting significantly outperforms existing CoT prompting methods on five reasoning tasks under noisy context. With GPT-3.5-turbo, we observe 3.7% accuracy improvement on average on the reasoning tasks under noisy context compared to the most competitive prompting baseline. More analyses and ablation studies show the robustness and generalization of R$^3$ prompting method in solving reasoning tasks in LLMs under noisy context

Improving Zero-shot Visual Question Answering via Large Language Models with Reasoning Question Prompts

Zero-shot Visual Question Answering (VQA) is a prominent vision-language task that examines both the visual and textual understanding capability of systems in the absence of training data. Recently, by converting the images into captions, information across multi-modalities is bridged and Large Language Models (LLMs) can apply their strong zero-shot generalization capability to unseen questions. To design ideal prompts for solving VQA via LLMs, several studies have explored different strategies to select or generate question-answer pairs as the exemplar prompts, which guide LLMs to answer the current questions effectively. However, they totally ignore the role of question prompts. The original questions in VQA tasks usually encounter ellipses and ambiguity which require intermediate reasoning. To this end, we present Reasoning Question Prompts for VQA tasks, which can further activate the potential of LLMs in zero-shot scenarios. Specifically, for each question, we first generate self-contained questions as reasoning question prompts via an unsupervised question edition module considering sentence fluency, semantic integrity and syntactic invariance. Each reasoning question prompt clearly indicates the intent of the original question. This results in a set of candidate answers. Then, the candidate answers associated with their confidence scores acting as answer heuristics are fed into LLMs and produce the final answer. We evaluate reasoning question prompts on three VQA challenges, experimental results demonstrate that they can significantly improve the results of LLMs on zero-shot setting and outperform existing state-of-the-art zero-shot methods on three out of four data sets. Our source code is publicly released at \url{https://github.com/ECNU-DASE-NLP/RQP}

Multisite phosphorylation is required for sustained interaction with GRKs and arrestins during rapid -opioid receptor desensitization

Copyright © 2018 The Authors. G protein receptor kinases (GRKs) and -arrestins are key regulators of -opioid receptor (MOR) signaling and trafficking. We have previously shown that high-efficacy opioids such as DAMGO stimulate a GRK2/3-mediated multisite phosphorylation of conserved C-terminal tail serine and threonine residues, which facilitates internalization of the receptor. In contrast, morphine-induced phosphorylation of MOR is limited to Ser375 and is not sufficient to drive substantial receptor internalization. We report how specific multisite phosphorylation controlled the dynamics of GRK and -arrestin interactions with MOR and show how such phosphorylation mediated receptor desensitization. We showed that GRK2/3 was recruited more quickly than was -arrestin to a DAMGO-activated MOR. -Arrestin recruitment required GRK2 activity and MOR phosphorylation, but GRK recruitment also depended on the phosphorylation sites in the C-terminal tail, specifically four serine and threonine residues within the 370TREHPSTANT379 motif. Our results also suggested that other residues outside this motif participated in the initial and transient recruitment of GRK and -arrestins. We identified two components of high-efficacy agonist desensitization of MOR: a sustained component, which required GRK2-mediated phosphorylation and a potential soluble factor, and a rapid component, which was likely mediated by GRK2 but independent of receptor phosphorylation. Elucidating these complex receptor-effector interactions represents an important step toward a mechanistic understanding of MOR desensitization that leads to the development of tolerance and dependence

Organizational slack and firm's internationalization: A longitudinal study of high-technology firms

We employ both organization theory and economic theory for exploring how the effects of high-discretion organizational slack and low-discretion organizational slack impact the degree of firm's internationalization. We tested hypotheses using longitudinal data (2000-2005) from 179 high-technology publicly listed companies in Taiwan. The results indicate a U-shaped relationship exists between high-discretion organizational slack and the internationalization of firms. In contrast, low-discretion organizational slack presents a positive linear relationship with the internationalization of firms.Organizational slack High-discretion organizational slack Low-discretion organizational slack Internationalization

Estimating the Effect of Vibration Mixing Process on Air Pore Size Distributions in Concrete Using Digital Image Analysis

Concrete is a typical porous material, in which the air voids entrained or entrapped during the mixing process have a significant impact on the material’s strength and durability. An automatic methodology based on digital image analysis was used to examine the influence of a novel mixing process with vibration on the entrapped air pore size and distribution of concrete in this paper. The volume of permeable spaces and porosity in hardened concrete are found to be greatly reduced when using the vibration mixing process compared to the reference concrete. Meanwhile, the quantity of air pores and their specific surface area are positively associated with the vibration acceleration, while the average equivalent pore diameter decreases. The findings of the analysis of variance (ANOVA) reveal that the population means for porosity, quantity, and pore size are significantly different when utilizing the vibration or non-vibration mixing processes. Furthermore, the pore size distribution curves show that the vibration mixing process significantly modified the pore structure by reducing the number of larger size pores and increasing the amount of small size pores. This may be attributed to a series of changes in the bubbles during the vibration mixing process. In addition, the findings of freeze-thaw resistance and water penetration resistance reveal that, owing to the vibration mixing process, the impermeability and durability of the concrete are significantly improved

Estimating the Effect of Vibration Mixing Process on Air Pore Size Distributions in Concrete Using Digital Image Analysis

Concrete is a typical porous material, in which the air voids entrained or entrapped during the mixing process have a significant impact on the material’s strength and durability. An automatic methodology based on digital image analysis was used to examine the influence of a novel mixing process with vibration on the entrapped air pore size and distribution of concrete in this paper. The volume of permeable spaces and porosity in hardened concrete are found to be greatly reduced when using the vibration mixing process compared to the reference concrete. Meanwhile, the quantity of air pores and their specific surface area are positively associated with the vibration acceleration, while the average equivalent pore diameter decreases. The findings of the analysis of variance (ANOVA) reveal that the population means for porosity, quantity, and pore size are significantly different when utilizing the vibration or non-vibration mixing processes. Furthermore, the pore size distribution curves show that the vibration mixing process significantly modified the pore structure by reducing the number of larger size pores and increasing the amount of small size pores. This may be attributed to a series of changes in the bubbles during the vibration mixing process. In addition, the findings of freeze-thaw resistance and water penetration resistance reveal that, owing to the vibration mixing process, the impermeability and durability of the concrete are significantly improved

The internationalization and performance of a firm: Moderating effect of a firm's behavior

Empirical studies show mixed results regarding the relationship between a firm's internationalization and performance. One reason is that these studies used various economic market firms as their research sample. Thus, this study on a firm's internationalization is based on newly industrialized economies to shed additional light on this relationship. According to the firm behavioral theory, the moderating impact of a firm's internal organizational slack and attainment discrepancy are critical for enhancing a firm's performance in the internationalization process. We tested these hypotheses using longitudinal data (2000-2005) from 179 high-technology listed companies in Taiwan. We extended the behavioral theory of the firm to explain the moderating effect on the internationalization-performance model. We found that firms tend to exhibit positive performance in internationalization when they have a higher level of organizational slack and attainment discrepancy.Internationalization Performance Organizational slack Attainment discrepancy

Solid Phase Synthesis of DNA Nanostructures in Heavy Liquid

Introduction of the solid phase method to synthesize biopolymers has revolutionized the field of biological research by enabling efficient production of peptides and oligonucleotides. One of the advantages of this method is the ease of removal of excess production materials from the desired product, as it is immobilized on solid substrate. The DNA origami method utilizes the nature of nucleotide base-pairing to construct well-defined objects at the nanoscale, and has become a potent tool for manipulating matter in the fields of chemistry, physics, and biology. Here, the development of an approach to synthesize DNA nanostructures directly on magnetic beads, where the reaction is performed in heavy liquid to maintain the beads in suspension is reported. It is demonstrated that the method can achieve high folding yields of up to 90% for various DNA shapes, comparable to standard folding. At the same time, this establishes an easy, fast, and efficient way to further functionalize the DNA origami in one-pot, as well as providing a built-in purification method for easy removal of excess by-products such as non-integrated DNA strands and residual functionalization molecules.Peer reviewe