Dynamic Cellular Environment Research Articles

DNA-based ForceChrono probes for deciphering single-molecule force dynamics in living cells

Understanding cellular force transmission dynamics is crucial in mechanobiology. We developed the DNA-based ForceChrono probe to measure force magnitude, duration, and loading rates at the single-molecule level within living cells. The ForceChrono probe circumvents the limitations of invitro single-molecule force spectroscopy by enabling direct measurements within the dynamic cellular environment. Our findings reveal integrin force loading rates of 0.5-2 pN/s and durations ranging from tens of seconds in nascent adhesions to approximately 100s in mature focal adhesions. The probe's robust and reversible design allows for continuous monitoring of these dynamic changes as cells undergo morphological transformations. Additionally, by analyzing how mutations, deletions, or pharmacological interventions affect these parameters, we can deduce the functional roles of specific proteins or domains in cellular mechanotransduction. The ForceChrono probe provides detailed insights into the dynamics of mechanical forces, advancing our understanding of cellular mechanics and the molecular mechanisms of mechanotransduction.

CORDAX web server: an online platform for the prediction and 3D visualization of aggregation motifs in protein sequences.

Proteins, the molecular workhorses of biological systems, execute a multitude of critical functions dictated by their precise three-dimensional structures. In a complex and dynamic cellular environment, proteins can undergo misfolding, leading to the formation of aggregates that take up various forms, including amorphous and ordered aggregation in the shape of amyloid fibrils. This phenomenon is closely linked to a spectrum of widespread debilitating pathologies, such as Alzheimer's disease, Parkinson's disease, type-II diabetes, and several other proteinopathies, but also hampers the engineering of soluble agents, as in the case of antibody development. As such, the accurate prediction of aggregation propensity within protein sequences has become pivotal due to profound implications in understanding disease mechanisms, as well as in improving biotechnological and therapeutic applications. We previously developed Cordax, a structure-based predictor that utilizes logistic regression to detect aggregation motifs in protein sequences based on their structural complementarity to the amyloid cross-beta architecture. Here, we present a dedicated web server interface for Cordax. This online platform combines several features including detailed scoring of sequence aggregation propensity, as well as 3D visualization with several customization options for topology models of the structural cores formed by predicted aggregation motifs. In addition, information is provided on experimentally determined aggregation-prone regions that exhibit sequence similarity to predicted motifs, scores, and links to other predictor outputs, as well as simultaneous predictions of relevant sequence propensities, such as solubility, hydrophobicity, and secondary structure propensity. The Cordax webserver is freely accessible at https://cordax.switchlab.org/.

Enabling Delay-Guaranteed Congestion Control With One-Bit Feedback in Cellular Networks

Unexpected large packet delays are often observed in cellular networks due to huge network queuing caused by excessive traffic coming into the network. To deal with the large queue problem, many congestion control algorithms try to find out how much traffic the network can accommodate, either by measuring network performance or by directly providing explicit information. However, due to the nature of the control in which queue growth should be observed or the necessity to modify the overall network architecture, existing algorithms are experiencing difficulties in keeping queues within a strict bound. In this paper, we propose a novel congestion control algorithm based on simple feedback, ECLAT which can provide bounded queuing delay using only one-bit signaling already available in traditional network architecture. To do so, a base station or a router running ECLAT 1) calculates how many packets each flow should transmit and 2) analyzes when congestion feedback needs to be forwarded to adjust the flow’s packet transmission to the desired rate. Our extensive experiments in our testbed demonstrate that ECLAT achieves strict queuing delay bounds, even in the dynamic cellular network environment.

Simultaneous Nanorheometry and Nanothermometry Using Intracellular Diamond Quantum Sensors.

The viscoelasticity of the cytoplasm plays a critical role in cell morphology, cell division, and intracellular transport. Viscoelasticity is also interconnected with other biophysical properties, such as temperature, which is known to influence cellular bioenergetics. Probing the connections between intracellular temperature and cytoplasmic viscoelasticity provides an exciting opportunity for the study of biological phenomena, such as metabolism and disease progression. The small length scales and transient nature of changes in these parameters combined with their complex interdependencies pose a challenge for biosensing tools, which are often limited to a single readout modality. Here, we present a dual-mode quantum sensor capable of performing simultaneous nanoscale thermometry and rheometry in dynamic cellular environments. We use nitrogen-vacancy centers in diamond nanocrystals as biocompatible sensors for in vitro measurements. We combine subdiffraction resolution single-particle tracking in a fluidic environment with optically detected magnetic resonance spectroscopy to perform simultaneous sensing of viscoelasticity and temperature. We use our sensor to demonstrate probing of the temperature-dependent viscoelasticity in complex media at the nanoscale. We then investigate the interplay between intracellular forces and the cytoplasmic rheology in live cells. Finally, we identify different rheological regimes and reveal evidence of active trafficking and details of the nanoscale viscoelasticity of the cytoplasm.

Recency effect and self-regulating design of mobile cellular systems in the context of interlaced generations: network bandwidth, power efficiency, and connection reliability

The mobile cellular system is undergoing a generation-interlaced evolution from 2G/3G/4G to 5G, with various signals and protocols coexisting for a long period. A crucial but challenging problem that emerges is how to ensure high performance (fast transmission), high energy efficiency (low power consumption), and high reliability (stable connection) for the services in a large, heterogeneous, and highly dynamic cellular environment. This problem simultaneously involves mobile phone manufacturers, network service providers, and OS/application developers. In this paper, we comprehensively review the representative related works in recent years, and learn that a series of performance anomalies in cellular systems is in fact attributed to the “recency effect" at the core mechanisms. Upon selecting which mobile network to use, there exist generation biases and unconscious performance-first tendencies affected by public opinions and commercial promotions. The biases and tendencies lead to knock-on effects like the “loss-making migration" towards 5G, the “saturated hunger" of network capacity, and the “ping-pong jumping" among base station connections. To overcome the above shortcomings, we propose self-regulating techniques based on 4G/5G dual-connectivity and Time-Inhomogenous Markov Process, which reduce the failure rate of 5G devices by 40% and the average failure duration of all devices by 36%. We also discuss other insights and innovations based on large-scale industrial practice.

Designing bioresponsive nanomaterials for intracellular self-assembly.

Supramolecular assemblies are essential components of living organisms. Cellular scaffolds, such as the cytoskeleton or the cell membrane, are formed via secondary interactions between proteins or lipids and direct biological processes such as metabolism, proliferation and transport. Inspired by nature’s evolution of function through structure formation, a range of synthetic nanomaterials has been developed in the past decade, with the goal of creating non-natural supramolecular assemblies inside living mammalian cells. Given the intricacy of biological pathways and the compartmentalization of the cell, different strategies can be employed to control the assembly formation within the highly crowded, dynamic cellular environment. In this Review, we highlight emerging molecular design concepts aimed at creating precursors that respond to endogenous stimuli to build nanostructures within the cell. We describe the underlying reaction mechanisms that can provide spatial and temporal control over the subcellular formation of synthetic nanostructures. Showcasing recent advances in the development of bioresponsive nanomaterials for intracellular self-assembly, we also discuss their impact on cellular function and the challenges associated with establishing structure–bioactivity relationships, as well as their relevance for the discovery of novel drugs and imaging agents, to address the shortfall of current solutions to pressing health issues.

Probing the Hidden Sensitivity of Intrinsically Disordered Proteins to Their Chemical Environment

Intrinsically disordered protein-regions (IDRs) play pivotal roles in signaling and regulatory pathways. A low number of intramolecular bonds and a high degree of surface area exposure make IDRs highly sensitive to changes in their surrounding solution, a situation that can be imagined as a constantly evolving “tug-of-war” between intramolecular and protein-solution interactions. Despite the vital importance of IDRs to cellular function, how they respond to fluctuations in the chemically dynamic cellular environment has not been thoroughly studied. To investigate changes in IDR structure in response to changing chemical surroundings, we developed a FRET-based high-throughput approach. We have used this approach, which we call “solution-space scanning,” to test four naturally occurring IDRs and three sequence scrambles of a naturally occurring IDR in over 180 biologically-compatible solution conditions. We found each sequence to have its own unique set of concentration-dependent responses to different solutes, which we term the sequence's “solution-space fingerprint.” We see that these fingerprints depend on IDR sequence and amino acid composition, but not on length. In live cells, using the same FRET constructs, the four naturally occurring IDRs showed global dimension trends in line with our high-throughput in vitro assay results. Our results underline the idea that IDRs must be understood not only in terms of their sequence, but also in the context of their environment. Based on these findings, we propose a general mechanism by which IDRs are able to function as sensors and actuators within cells.

Three-dimensional bioprinting of aneurysm-bearing tissue structure for endovascular deployment of embolization coils

Various types of embolization devices have been developed for the treatment of cerebral aneurysms. However, it is challenging to properly evaluate device performance and train medical personnel for device deployment without the aid of functionally relevant models. Current in vitro aneurysm models suffer from a lack of key functional and morphological features of brain vasculature that limit their applicability for these purposes. These features include the physiologically relevant mechanical properties and the dynamic cellular environment of blood vessels subjected to constant fluid flow. Herein, we developed three-dimensionally (3D) printed aneurysm-bearing vascularized tissue structures using gelatin-fibrin hydrogel of which the inner vessel walls were seeded with human cerebral microvascular endothelial cells (hCMECs). The hCMECs readily exhibited cellular attachment, spreading, and confluency all around the vessel walls, including the aneurysm walls. Additionally, the in vitro platform was directly amenable to flow measurements via particle image velocimetry, enabling the direct assessment of the vascular flow dynamics for comparison to a 3D computational fluid dynamics model. Detachable coils were delivered into the printed aneurysm sac through the vessel using a microcatheter and static blood plasma clotting was monitored inside the aneurysm sac and around the coils. This biomimetic in vitro aneurysm model is a promising method for examining the biocompatibility and hemostatic efficiency of embolization devices and for providing hemodynamic information which would aid in predicting aneurysm rupture or healing response after treatment.

Biomarkers in traumatic brain injury: new concepts.

Traumatic brain injury is a multifaceted condition that encompasses a spectrum of injuries: contusions, axonal injuries in specific brain regions, edema, and hemorrhage. Brain injury determines a broad clinical and disability spectrum due to the implication of various cellular pathways, genetic phenotypes, and environmental factors. It is challenging to predict patient outcomes, to appropriately evaluate the patients, to determine a suitable treatment strategy and rehabilitation program, and to communicate with patient relatives. Biomarkers detected from body fluids are potential evaluation tools for traumatic brain injury patients. These may serve as internal indicators of cerebral damage, delivering valuable information about the dynamic cellular, biochemical, and molecular environments. The diagnostic and prognostic value of biomarkers tested both in animal models of traumatic brain injury is still under question, despite a considerable scientific literature. Recent publications emphasize that a more realistic approach involves combining multiple types of biomarkers with other investigative tools (imaging, outcome scales, and genetic polymorphisms). Additionally, there is increasing interest in the use of biomarkers as tools for treatment monitoring and as surrogate outcome variables to facilitate the design of distinct randomized controlled trials. This review highlights the latest available evidence regarding biomarkers in adults after traumatic brain injury and discusses new approaches in the evaluation of this patient group.

Signal integration and information transfer in an allosterically regulated network

A biological reaction network may serve multiple purposes, processing more than one input and impacting downstream processes via more than one output. These networks operate in a dynamic cellular environment in which the levels of network components may change within cells and across cells. Recent evidence suggests that protein concentration variability could explain cell fate decisions. However, systems with multiple inputs, multiple outputs, and changing input concentrations have not been studied in detail due to their complexity. Here, we take a systems biochemistry approach, combining physiochemical modeling and information theory, to investigate how cyclooxygenase-2 (COX-2) processes simultaneous input signals within a complex interaction network. We find that changes in input levels affect the amount of information transmitted by the network, as does the correlation between those inputs. This, and the allosteric regulation of COX-2 by its substrates, allows it to act as a signal integrator that is most sensitive to changes in relative input levels.

How is mechanobiology involved in mesenchymal stem cell differentiation toward the osteoblastic or adipogenic fate?

Mechanobiology plays a major role in transducing physical cues from the dynamic cellular environment into biochemical modifications that promote cell-specific differentiation paths. Mesenchymal stem cells in the bone marrow or in other mesenchymal tissues will differentiate according to the expression of transcription factors (TFs) that govern their lineage commitment. The favoring of either osteogenic or adipogenic differentiation relies on TF expression as well as mechanical properties of the cells' niche that are translated into the activation of certain signaling pathways. Physical factors can induce significant shifts in bipotential lineage commitment between osteogenesis and adipogenesis. The stiffness of the extracellular matrix (ECM) surrounding a cell, varying greatly from rigid environments close to the bone surface to softer regions in the bone marrow, can influence the path of differentiation. Additionally, mechanical loading through exercise appears to favor osteogenesis whereas disuse conditions seem to promote adipogenesis.

“Exploring the Regulation of Photosynthesis in Single‐Cell Lineages at Sub‐Cellular Resolution”

Photosynthesis is regulated in response to many diverse stimuli including light intensity and wavelength, nutrient availability, and temperature. Understanding how photosynthesis is regulated is critical if we are to increase photosynthetic efficiency for the benefit of society and the environment. While decades of research have revealed the structures and mechanisms of isolated photosynthetic complexes at molecular resolution, little is known about how these complexes interact and are regulated in the context of the dynamic cellular environment.To investigate how photosynthesis is regulated in single‐cells, we utilized long‐term, quantitative time‐lapse fluorescence microscopy to visualize the growth dynamics of wild‐type and mutant cyanobacterial strains following diverse environmental, chemical, and genetic perturbations. Quantitative image processing with custom algorithms enabled us to track individual cells across single‐cell derived lineages for multiple generations and monitor cellular physiology at sub‐cellular resolution.Our results suggest that mechanical interactions with the physical environment can lead to attenuation of photosynthesis and can play a major role in governing cellular fitness and productivity in cyanobacteria. The mechanistic basis of this phenomenon was investigated using mathematical modeling, electron microscopy, mutational analysis, and CRISPRi‐based dynamic gene regulation. These results provide insight into how photosynthetic organisms navigate the physical environment and a new perspective on source‐sink regulation of photosynthesis.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Biomimetic shear stress and nanoparticulate drug delivery

Fluidic shear stress generated by fluid flow in human body contributes to the interactions between nanoparticles (NPs) and cells and may affect cellular distribution and delivery of NPs. Furthermore, different NPs properties may differently influence the cellular delivery and targeting of NPs under the in vivo dynamic environments. Thus, we reviewed and discussed how fluidic shear stress affects to drug delivery and cellular distribution of diverse NPs under the biomimetic microfluidic shear stress. Among different physicochemical properties of NPs, size, shape, material type and surface functionality, and surface charge of NPs are critical factors to cellular uptake in presence of fluidic shear stress in dynamic cellular environment. From previous studies, it suggested that fluidic shear stress stimulated specific endocytosis and prompted cell signaling pathway. The cellular interactions between NPs and cells in drug delivery should be carefully investigated in presence of shear stress which is one of critical factors to show the difference of in vitro and in vivo drug distribution and therapy. The shear-activated drug delivery using NPs were also the scope of this review.

Site-specific azide-acetyllysine photochemistry on epigenetic readers for interactome profiling.

Chemical modifications on DNA, RNA and histones are recognized by an array of 'reader' modules to regulate transcriptional programming and cell fate. However, identification of reader-specific interacting partners in a dynamic cellular environment remains a significant challenge. Herein, we report a chemoproteomic approach termed 'interaction-based protein profiling' (IBPP) to characterize novel interacting partners of potentially any reader protein. IBPP harnesses a photosensitive amino acid introduced into the hydrophobic pocket of a reader module to crosslink and enrich transient interacting partners that are inaccessible to traditional methods. Using bromodomain-containing protein 4 (BRD4) as a paradigm, we engineer an 'aromatic cage' of the bromodomain to introduce 4-azido-l-phenylalanine (pAzF) without compromising its ability to recognize acetylated lysine residues in histone proteins. We establish the binding efficiency, substrate specificity and crosslinking ability of the engineered 'reader' module in biochemical assays. Applying IBPP, we uncovered novel acetylated interacting partners of BRD4, such as transcription factors, expanding on its previously unappreciated role in diverse biological processes. By setting up an azide-acetyllysine photoreaction deep inside the bromodomain aromatic cage as a means to detect protein acetylation, our approach provides a potentially general platform for rapid and unbiased profiling of interacting partners of diverse epigenetic readers whose functions in eukaryotic gene regulation remain convoluted.

Cognitive spectrum management in dynamic cellular environments: A case-based Q-learning approach

This paper examines how novel cellular system architectures and intelligent spectrum management techniques can be used to play a key role in accommodating the exponentially increasing demand for mobile data capacity in the near future. A significant challenge faced by the artificial intelligence methods applied to such flexible wireless communication systems is their dynamic nature, e.g. network topologies that change over time. This paper proposes an intelligent case-based Q-learning method for dynamic spectrum access (DSA) which improves and stabilises the performance of cognitive cellular systems with dynamic topologies. The proposed approach is the combination of classical distributed Q-learning and a novel implementation of case-based reasoning which aims to facilitate a number of learning processes running in parallel. Large scale simulations of a stadium small cell network show that the proposed case-based Q-learning approach achieves a consistent improvement in the system quality of service (QoS) under dynamic and asymmetric network topology and traffic load conditions. Simulations of a secondary spectrum sharing scenario show that the cognitive cellular system that employs the proposed case-based Q-learning DSA scheme is able to accommodate a 28-fold increase in the total primary and secondary system throughput, but with no need for additional spectrum and with no degradation in the primary user QoS.

How Bacteria Subvert Animal Cell Structure and Function.

Bacterial pathogens encode a wide variety of effectors and toxins that hijack host cell structure and function. Of particular importance are virulence factors that target actin cytoskeleton dynamics critical for cell shape, stability, motility, phagocytosis, and division. In addition, many bacteria target organelles of the general secretory pathway (e.g., the endoplasmic reticulum and the Golgi complex) and recycling pathways (e.g., the endolysosomal system) to establish and maintain an intracellular replicative niche. Recent research on the biochemistry and structural biology of bacterial effector proteins and toxins has begun to shed light on the molecular underpinnings of these host-pathogen interactions. This exciting work is revealing how pathogens gain control of the complex and dynamic host cellular environments, which impacts our understanding of microbial infectious disease, immunology, and human cell biology.

Optimization of Worker Assignment in Dynamic Cellular Manufacturing System Using Genetic Algorithm

Cellular manufacturing system (CMS) involves a number of machine cells, where each cell is responsible for the processing of families of similar parts. The dynamic cellular manufacturing environment refers to anticipated changes of demand or production process for multiple time periods. This paper applied with the meta-heuristics for the design of Dynamic Cellular Manufacturing System (DCMS) using genetic algorithm (GA) to minimize holding cost, back order cost, machine cost and salary cost. It is shown that the PSO algorithm is efficient in finding out good quality solutions for the cellular manufacturing in a dynamic environment.

In vitro micro-physiological immune-competent model of the human skin.

Skin allergy, in particular, allergic contact dermatitis and irritant contact dermatitis, are common occupational and environmental health problems affecting the quality of life of a significant proportion of the world population. Since all new ingredients to be incorporated into a product are potential skin allergens, it is essential that these ingredients be first tested for their allergenic potential. However, despite the considerable effort using animal models to understand the underlying mechanism of skin sensitization, to date, the molecular and cellular responses due to skin contact with sensitizers are still not fully understood. To replace animal testing and to improve the prediction of skin sensitization, significant attention has been directed to the use of reconstructed organotypic in vitro models of human skin. Here we describe a miniaturized immune competent in vitro model of human skin based on 3D co-culture of immortalized human keratinocytes (HaCaT) as a model of the epidermis barrier and human leukemic monocyte lymphoma cell line (U937) as a model of human dendritic cells. The biological model was fitted in a microfluidic-based cell culture system that provides a dynamic cellular environment that mimics the in vivo environment of skin. The dynamic perfusion of culture media significantly improved the tight junction formation as evidenced by measuring higher values of TEER compared to static culture. This setting also maintained the high viability of cells over extended periods of time up to 17 days. The perfusion-based culture also allows growth of the cells at the air-liquid interface by exposing the apical side of the cells to air while providing the cell nutrients through a basolateral fluidic compartment. The microsystem has been evaluated to investigate the effect of the chemical and physical (UV irradiation) stimulation on the skin barrier (i.e. the TJ integrity). Three-tiered culture differential stimulation allowed the investigation of the role of the keratinocyte layer as a protection barrier to chemical/biological hazards.

Study of mRNA turnover never decays.

Study of mRNA turnover never decays.

Spatiotemporal mapping of three dimensional rotational dynamics of single ultrasmall gold nanorods.

Spatiotemporal mapping of the position and orientation of nano-machinery inside complex and dynamic cellular environments is essential for the detailed understanding of many bio-physical processes. For the genuine observation of such biomolecular dynamics with high signal to noise ratio and reduced disturbance from the labeling probes, reduction in the size of nano-bio labels and simplification of techniques for their observation are important. Here we achieve this using polarized dark field scattering micro-spectroscopy (PDFSMS), in its simplest form so that it is deployable in several experiments. We not only locate tiny gold nanorods (GNRs) of size 30 (length) × 10 nm (diameter) inside HEK293 cells but also demonstrate mapping of their in-situ polarization patterns using a novel method. Real time observations of rotating GNR with DFSMS and PDFSMS are used to resolve in-plane and out-of-plane rotational modes of GNR. We have shown that PDFSMS itself can provide complete information about the state of GNR. A step ahead, we demonstrate the application of PDFSMS to track three dimensional rotational dynamics of transferrin-conjugated GNRs inside live HEK293 cells. These first-time observations of the three dimensional intracellular rotational dynamics of tiny GNRs using PDFSMS present a new landmark in single particle scattering spectroscopy.