Investigating spatial distribution and dynamics of membrane proteins in polymer-tethered lipid bilayer systems using single molecule-sensitive imaging techniques

Indiana University-Purdue University Indianapolis (IUPUI)Plasma membranes are complex supramolecular assemblies comprised of lipids and membrane proteins. Both types of membrane constituents are organized in highly dynamic patches with profound impact on membrane functionality, illustrating the functional importance of plasma membrane fluidity. Exemplary, dynamic processes of membrane protein oligomerization and distribution are of physiological and pathological importance. However, due to the complexity of the plasma membrane, the underlying regulatory mechanisms of membrane protein organization and distribution remain elusive. To address this shortcoming, in this thesis work, different mechanisms of dynamic membrane protein assembly and distribution are examined in a polymer-tethered lipid bilayer system using comple-mentary confocal optical detection techniques, including 2D confocal imaging and single molecule-sensitive confocal fluorescence intensity analysis methods [fluorescence correlation spectroscopy (FCS) autocorrelation analysis and photon counting histogram (PCH) method]. Specifically, this complementary methodology was applied to investigate mechanisms of membrane protein assembly and distribution, which are of significance in the areas of membrane biophysics and cellular mechanics. From the membrane biophysics perspective, the role of lipid heterogeneities in the distribution and function of membrane proteins in the plasma membrane has been a long-standing problem. One of the most well-known membrane heterogeneities are known as lipid rafts, which are domains enriched in sphingolipids and cholesterol (CHOL). A hallmark of lipid rafts is that they are important regulators of membrane protein distribution and function in the plasma membrane. Unfortunately, progress in deciphering the mechanisms of raft-mediated regulation of membrane protein distribution has been sluggish, largely due to the small size and transient nature of raft domains in cellular membranes. To overcome this challenge, the current thesis explored the distribution and oligomerization of membrane proteins in raft-mimicking lipid mixtures, which form stable coexisting CHOL-enriched and CHOL-deficient lipid domains of micron-size, which can easily be visualized using optical microscopy techniques. In particular, model membrane experiments were designed, which provided insight into the role of membrane CHOL level versus binding of native ligands on the oligomerization state and distribution of GPI-anchored urokinase plasminogen activator receptor (uPAR) and the transmembrane protein αvβ3 integrin. Experiments on uPAR showed that receptor oligomerization and raft sequestration are predominantly influenced by the binding of natural ligands, but are largely independent of CHOL level changes. In contrast, through a presumably different mechanism, the sequestration of αvβ3 integrin in raft-mimicking lipid mixtures is dependent on both ligand binding and CHOL content changes without altering protein oligomerization state. In addition, the significance of membrane-embedded ligands as regulators of integrin sequestration in raft-mimicking lipid mixtures was explored. One set of experiments showed that ganglioside GM3 induces dimerization of α5β1 integrins in a CHOL-free lipid bilayer, while addition of CHOL suppresses such a dimerization process. Furthermore, GM3 was found to recruit α5β1 integrin into CHOL-enriched domains, illustrating the potential sig-nificance of GM3 as a membrane-associated ligand of α5β1 integrin. Similarly, uPAR was observed to form complexes with αvβ3 integrin in a CHOL dependent manner, thereby causing the translocation of the complex into CHOL-enriched domains. Moreover, using a newly developed dual color FCS and PCH assay, the composition of uPAR and integrin within complexes was determined for the first time. From the perspective of cell mechanics, the characterization of the dynamic assembly of membrane proteins during formation of cell adhesions represents an important scientific problem. Cell adhesions play an important role as force transducers of cellular contractile forces. They may be formed between cell and extracellular matrix, through integrin-based focal adhesions, as well as between different cells, through cadherin-based adherens junctions (AJs). Importantly, both types of cell adhesions act as sensitive force sensors, which change their size and shape in response to external mechanical signals. Traditionally, the correlation between adhesion linker assembly and external mechanical cues was investigated by employing polymeric substrates of adjustable substrate stiffness containing covalently attached linkers. Such systems are well suited to mimic the mechanosensitive assembly of focal adhesions (FAs), but fail to replicate the rich dynamics of cell-cell linkages, such as treadmilling of adherens junctions, during cellular force sensing. To overcome this limitation, the 2D confocal imaging methodology was applied to investigate the dynamic assembly of N-cadherin-chimera on the surface of a polymer-tethered lipid multi-bilayer in the presence of plated cells. Here, the N-cadherin chimera-functionalized polymer-tethered lipid bilayer acts as a cell surface-mimicking cell substrate, which: (i) allows the adjustment of substrate stiffness by changing the degree of bilayer stacking and (ii) enables the free assembly of N-cadherin chimera linkers into clusters underneath migrating cells, thereby forming highly dynamic cell-substrate linkages with remarkable parallels to adherens junctions. By applying the confocal methodology, the dynamic assembly of dye-labeled N-cadherin chimera into clusters was monitored underneath adhered cells. Moreover, the long-range mobility of N-cadherin chimera clusters was analyzed by tracking the cluster positions over time using a MATLAB-based multiple-particle tracking method. Disruption of the cytoskeleton organization of plated cells confirmed the disassembly of N-cadherin chimera clusters, emphasizing the important role of the cytoskeleton of migrating cells during formation of cadherin-based cell-substrate linkages. Size and dynamics of N-cadherin chimera clusters were also analyzed as a function of substrate stiffness

A WRF-UCM-SOLWEIG framework of 10m resolution to quantify the intra-day impact of urban features on thermal comfort

City-scale outdoor thermal comfort diagnostics are essential for understanding actual heat stress. However, previous research primarily focused on the street scale. Here, we present the WRF-UCM-SOLWEIG framework to achieve fine-grained thermal comfort mapping at the city scale. The background climate condition affecting thermal comfort is simulated by the Weather Research and Forecasting (WRF) model coupled with the urban canopy model (UCM) at a local-scale (500m). The most dominant factor, mean radiant temperature, is simulated using the Solar and Longwave Environmental Irradiance Geometry (SOLWEIG) model at the micro-scale (10m). The Universal Thermal Climate Index (UTCI) is calculated based on the mean radiant temperature and local climate parameters. The influence of different ground surface materials, buildings, and tree canopies is simulated in the SOLWEIG model using integrated urban morphological data. We applied this proposed framework to the city of Guangzhou, China, and investigated the intra-day variation in the impact of urban morphology during a heat wave period. Through statistical analysis, we found that the elevation in UTCI is primarily attributed to the increase in the fraction of impervious surface (ISF) during daytime, with a maximum correlation coefficient of 0.80. Tree canopy cover has a persistent cooling effect during the day. Implementing 40% of tree cover can reduce the daytime UTCI by 1.5 to 2.0 K. At nighttime, all urban features have a negligible contribution to outdoor thermal comfort. Overall, the established framework provides essential input data and references for studies and urban planners in the practice of urban (micro)climate diagnostics and planning

BPKD: Boundary Privileged Knowledge Distillation For Semantic Segmentation

Current knowledge distillation approaches in semantic segmentation tend to adopt a holistic approach that treats all spatial locations equally. However, for dense prediction, students' predictions on edge regions are highly uncertain due to contextual information leakage, requiring higher spatial sensitivity knowledge than the body regions. To address this challenge, this paper proposes a novel approach called boundary-privileged knowledge distillation (BPKD). BPKD distills the knowledge of the teacher model's body and edges separately to the compact student model. Specifically, we employ two distinct loss functions: (i) edge loss, which aims to distinguish between ambiguous classes at the pixel level in edge regions; (ii) body loss, which utilizes shape constraints and selectively attends to the inner-semantic regions. Our experiments demonstrate that the proposed BPKD method provides extensive refinements and aggregation for edge and body regions. Additionally, the method achieves state-of-the-art distillation performance for semantic segmentation on three popular benchmark datasets, highlighting its effectiveness and generalization ability. BPKD shows consistent improvements across a diverse array of lightweight segmentation structures, including both CNNs and transformers, underscoring its architecture-agnostic adaptability. The code is available at \url{https://github.com/AkideLiu/BPKD}.Comment: 17 pages, 9 figures, 9 table

TextPainter: Multimodal Text Image Generation with Visual-harmony and Text-comprehension for Poster Design

Text design is one of the most critical procedures in poster design, as it relies heavily on the creativity and expertise of humans to design text images considering the visual harmony and text-semantic. This study introduces TextPainter, a novel multimodal approach that leverages contextual visual information and corresponding text semantics to generate text images. Specifically, TextPainter takes the global-local background image as a hint of style and guides the text image generation with visual harmony. Furthermore, we leverage the language model and introduce a text comprehension module to achieve both sentence-level and word-level style variations. Besides, we construct the PosterT80K dataset, consisting of about 80K posters annotated with sentence-level bounding boxes and text contents. We hope this dataset will pave the way for further research on multimodal text image generation. Extensive quantitative and qualitative experiments demonstrate that TextPainter can generate visually-and-semantically-harmonious text images for posters.Comment: Accepted to ACM MM 2023. Dataset Link: https://tianchi.aliyun.com/dataset/16003

Continuous Beam Steering Through Broadside Using Asymmetrically Modulated Goubau Line Leaky-Wave Antennas

Goubau line is a single-conductor transmission line, featuring easy integration and low-loss transmission properties. Here, we propose a periodic leaky-wave antenna (LWA) based on planar Goubau transmission line on a thin dielectric substrate. The leaky-wave radiations are generated by introducing periodic modulations along the Goubau line. In this way, the surface wave, which is slow-wave mode supported by the Goubau line, achieves an additional momentum and hence enters the fast-wave region for radiations. By employing the periodic modulations, the proposed Goubau line LWAs are able to continuously steer the main beam from backward to forward within the operational frequency range. However, the LWAs usually suffer from a low radiation efficiency at the broadside direction. To overcome this drawback, we explore both transversally and longitudinally asymmetrical modulations to the Goubau line. Theoretical analysis, numerical simulations and experimental results are given in comparison with the symmetrical LWAs. It is demonstrated that the asymmetrical modulations significantly improve the radiation efficiency of LWAs at the broadside. Furthermore, the measurement results agree well with the numerical ones, which experimentally validates the proposed LWA structures. These novel Goubau line LWAs, experimentally demonstrated and validated at microwave frequencies, show also great potential for millimeter-wave and terahertz systems

AutoPoster: A Highly Automatic and Content-aware Design System for Advertising Poster Generation

Advertising posters, a form of information presentation, combine visual and linguistic modalities. Creating a poster involves multiple steps and necessitates design experience and creativity. This paper introduces AutoPoster, a highly automatic and content-aware system for generating advertising posters. With only product images and titles as inputs, AutoPoster can automatically produce posters of varying sizes through four key stages: image cleaning and retargeting, layout generation, tagline generation, and style attribute prediction. To ensure visual harmony of posters, two content-aware models are incorporated for layout and tagline generation. Moreover, we propose a novel multi-task Style Attribute Predictor (SAP) to jointly predict visual style attributes. Meanwhile, to our knowledge, we propose the first poster generation dataset that includes visual attribute annotations for over 76k posters. Qualitative and quantitative outcomes from user studies and experiments substantiate the efficacy of our system and the aesthetic superiority of the generated posters compared to other poster generation methods.Comment: Accepted for ACM MM 202

Reciprocal polarization imaging of complex media

The vectorial evolution of polarized light interaction with a medium can reveal its microstructure and anisotropy beyond what can be obtained from scalar light interaction. Anisotropic properties (diattenuation, retardance, and depolarization) of a complex medium can be quantified by polarization imaging by measuring the Mueller matrix. However, polarization imaging in the reflection geometry, ubiquitous and often preferred in diverse applications, has suffered a poor recovery of the medium's anisotropic properties due to the lack of suitable decomposition of the Mueller matrices measured inside a backward geometry. Here, we present reciprocal polarization imaging of complex media after introducing reciprocal polar decomposition for backscattering Mueller matrices. Based on the reciprocity of the optical wave in its forward and backward scattering paths, the anisotropic diattenuation, retardance, and depolarization of a complex medium are determined by measuring the backscattering Mueller matrix. We demonstrate reciprocal polarization imaging in various applications for quantifying complex non-chiral and chiral media (birefringence resolution target, tissue sections, and glucose suspension), uncovering their anisotropic microstructures with remarkable clarity and accuracy. We also highlight types of complex media that Lu-Chipman and differential decompositions of backscattering Mueller matrices lead to erroneous medium polarization properties, whereas reciprocal polar decomposition recovers properly. Reciprocal polarization imaging will be instrumental in imaging complex media from remote sensing to biomedicine and will open new applications of polarization optics in reflection geometry