scholarly journals Protein nanomechanics in biological context

2021 ◽  
Author(s):  
Jorge Alegre-Cebollada
Keyword(s):  
Single Molecule ◽  
Current Knowledge ◽  
Biological Systems ◽  
Mechanical Forces ◽  
Loss Of Function ◽  
Living Matter ◽  
Conventional View ◽  
Form And Function ◽  
Biological Context

AbstractHow proteins respond to pulling forces, or protein nanomechanics, is a key contributor to the form and function of biological systems. Indeed, the conventional view that proteins are able to diffuse in solution does not apply to the many polypeptides that are anchored to rigid supramolecular structures. These tethered proteins typically have important mechanical roles that enable cells to generate, sense, and transduce mechanical forces. To fully comprehend the interplay between mechanical forces and biology, we must understand how protein nanomechanics emerge in living matter. This endeavor is definitely challenging and only recently has it started to appear tractable. Here, I introduce the main in vitro single-molecule biophysics methods that have been instrumental to investigate protein nanomechanics over the last 2 decades. Then, I present the contemporary view on how mechanical force shapes the free energy of tethered proteins, as well as the effect of biological factors such as post-translational modifications and mutations. To illustrate the contribution of protein nanomechanics to biological function, I review current knowledge on the mechanobiology of selected muscle and cell adhesion proteins including titin, talin, and bacterial pilins. Finally, I discuss emerging methods to modulate protein nanomechanics in living matter, for instance by inducing specific mechanical loss-of-function (mLOF). By interrogating biological systems in a causative manner, these new tools can contribute to further place protein nanomechanics in a biological context.

Unfolding single RNA molecules: bridging the gap between equilibrium and non-equilibrium statistical thermodynamics

2005 ◽  
Vol 38 (4) ◽  
pp. 291-301 ◽  
Author(s):  
Carlos Bustamante
Keyword(s):  
Single Molecule ◽  
Molecular Motors ◽  
Biological Systems ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Fluctuation Theorems ◽  
Rna Molecules ◽  
The Times ◽  
Non Equilibrium

During the last 15 years, scientists have developed methods that permit the direct mechanical manipulation of individual molecules. Using this approach, they have begun to investigate the effect of force and torque in chemical and biochemical reactions. These studies span from the study of the mechanical properties of macromolecules, to the characterization of molecular motors, to the mechanical unfolding of individual proteins and RNA. Here I present a review of some of our most recent results using mechanical force to unfold individual molecules of RNA. These studies make it possible to follow in real time the trajectory of each molecule as it unfolds and characterize the various intermediates of the reaction. Moreover, if the process takes place reversibly it is possible to extract both kinetic and thermodynamic information from these experiments at the same time that we characterize the forces that maintain the three-dimensional structure of the molecule in solution. These studies bring us closer to the biological unfolding processes in the cell as they simulate in vitro, the mechanical unfolding of RNAs carried out in the cell by helicases. If the unfolding process occurs irreversibly, I show here that single-molecule experiments can still provide equilibrium, thermodynamic information from non-equilibrium data by using recently discovered fluctuation theorems. Such theorems represent a bridge between equilibrium and non-equilibrium statistical mechanics. In fact, first derived in 1997, the first experimental demonstration of the validity of fluctuation theorems was obtained by unfolding mechanically a single molecule of RNA. It is perhaps a sign of the times that important physical results are these days used to extract information about biological systems and that biological systems are being used to test and confirm fundamental new laws in physics.

scholarly journals Drosophila CLIP-190 and mammalian CLIP-170 display reduced microtubule plus end association in the nervous system

2015 ◽  
Vol 26 (8) ◽  
pp. 1491-1508 ◽  
Author(s):  
Robin Beaven ◽  
Nikola S. Dzhindzhev ◽  
Yue Qu ◽  
Ines Hahn ◽  
Federico Dajas-Bailador ◽  
...  
Keyword(s):  
Nervous System ◽  
Current Knowledge ◽  
Coiled Coil ◽  
Growth Dynamics ◽  
Loss Of Function ◽  
Axon Extension ◽  
Coiled Coil Domain ◽  
C Terminus

Axons act like cables, electrically wiring the nervous system. Polar bundles of microtubules (MTs) form their backbones and drive their growth. Plus end–tracking proteins (+TIPs) regulate MT growth dynamics and directionality at their plus ends. However, current knowledge about +TIP functions, mostly derived from work in vitro and in nonneuronal cells, may not necessarily apply to the very different context of axonal MTs. For example, the CLIP family of +TIPs are known MT polymerization promoters in nonneuronal cells. However, we show here that neither Drosophila CLIP-190 nor mammalian CLIP-170 is a prominent MT plus end tracker in neurons, which we propose is due to low plus end affinity of the CAP-Gly domain–containing N-terminus and intramolecular inhibition through the C-terminus. Instead, both CLIP-190 and CLIP-170 form F-actin–dependent patches in growth cones, mediated by binding of the coiled-coil domain to myosin-VI. Because our loss-of-function analyses in vivo and in culture failed to reveal axonal roles for CLIP-190, even in double-mutant combinations with four other +TIPs, we propose that CLIP-190 and -170 are not essential axon extension regulators. Our findings demonstrate that +TIP functions known from nonneuronal cells do not necessarily apply to the regulation of the very distinct MT networks in axons.

scholarly journals KLF17 promotes human naïve pluripotency but is not required for its establishment

2020 ◽  
Author(s):  
Rebecca A. Lea ◽  
Afshan McCarthy ◽  
Stefan Boeing ◽  
Kathy K. Niakan
Keyword(s):  
Zinc Finger Protein ◽  
Current Knowledge ◽  
Species Conservation ◽  
Ectopic Expression ◽  
Human Embryos ◽  
Related Factor ◽  
Loss Of Function ◽  
Blastocyst Development ◽  
Summary Statement

AbstractCurrent knowledge of the transcriptional regulation of human pluripotency is incomplete, with lack of inter-species conservation observed. Single-cell transcriptomics of human embryos previously enabled us to identify transcription factors, including the zinc-finger protein KLF17, that are enriched in the human epiblast and naïve hESCs. Here we show that KLF17 is expressed coincident with the known pluripotency factors NANOG and SOX2 across human blastocyst development. We investigate the function of KLF17 in pluripotency using primed and naïve hESCs for gain- and loss-of-function analyses. We find that ectopic expression of KLF17 in primed hESCs is sufficient to induce a naïve-like transcriptome and that KLF17 can drive transgene-mediated resetting to naïve pluripotency. This implies a role for KLF17 in establishing naïve pluripotency. However, CRISPR-Cas9-mediated knockout studies reveal that KLF17 is not required for naïve pluripotency acquisition in vitro. Transcriptome analysis of naïve hESCs identifies subtle effects on metabolism and signalling following KLF17 loss of function, and possible redundancy with the related factor, KLF5. Overall, we show that KLF17 is sufficient, but not necessary, for naïve pluripotency under the given in vitro conditions.Summary statementInvestigating KLF17 in human pluripotency reveals that it is sufficient, but not necessary, to establish naïve hESCs. We posit that KLF17 is a peripheral regulator, like KLF2 in the mouse.

scholarly journals KLF17 promotes human naïve pluripotency but is not required for its establishment

Development ◽  
2021 ◽  
Author(s):  
Rebecca A. Lea ◽  
Afshan McCarthy ◽  
Stefan Boeing ◽  
Todd Fallesen ◽  
Kay Elder ◽  
...  
Keyword(s):  
Zinc Finger Protein ◽  
Current Knowledge ◽  
Species Conservation ◽  
Ectopic Expression ◽  
Human Embryos ◽  
Loss Of Function ◽  
Blastocyst Development ◽  
Finger Protein ◽  
The Given

Current knowledge of the transcriptional regulation of human pluripotency is incomplete, with lack of inter-species conservation observed. Single-cell transcriptomics analysis of human embryos previously enabled us to identify transcription factors, including the zinc-finger protein KLF17, that are enriched in the human epiblast and naïve hESCs. Here we show that KLF17 is expressed coincident with the known pluripotency-associated factors NANOG and SOX2 across human blastocyst development. We investigate the function of KLF17 using primed and naïve hESCs for gain- and loss-of-function analyses. We find that ectopic expression of KLF17 in primed hESCs is sufficient to induce a naïve-like transcriptome and that KLF17 can drive transgene-mediated resetting to naïve pluripotency. This implies a role for KLF17 in establishing naïve pluripotency. However, CRISPR-Cas9-mediated knockout studies reveal that KLF17 is not required for naïve pluripotency acquisition in vitro. Transcriptome analysis of naïve hESCs identifies subtle effects on metabolism and signalling pathways following KLF17 loss-of-function, and possible redundancy with other KLF paralogues. Overall, we show that KLF17 is sufficient, but not necessary, for naïve pluripotency under the given in vitro conditions.

scholarly journals DYT1 dystonia patient-derived fibroblasts have increased deformability and susceptibility to damage by mechanical forces

2018 ◽  
Author(s):  
Navjot Kaur Gill ◽  
Chau Ly ◽  
Paul H. Kim ◽  
Cosmo A. Saunders ◽  
Loren G. Fong ◽  
...  
Keyword(s):  
Human Fibroblasts ◽  
Mechanical Stretch ◽  
Mechanical Forces ◽  
Linc Complex ◽  
Loss Of Function ◽  
Mouse Fibroblasts ◽  
Dyt1 Dystonia ◽  
Sense And Respond ◽  
Aaa Protein

AbstractDYT1 dystonia is a neurological movement disorder that is caused by a loss-of-function mutation in the DYT1/TOR1A gene, which encodes torsinA, the luminal ATPase-associated (AAA+) protein. TorsinA is required for the assembly of functional linker of nucleoskeleton and cytoskeleton (LINC) complexes, and consequently the mechanical integration of the nucleus and the cytoskeleton. Despite the potential implications of altered mechanobiology in dystonia pathogenesis, the role of torsinA in regulating cellular mechanical phenotype, or mechanotype, in DYT1 dystonia remains unknown. Here, we define the mechanotype of mouse fibroblasts lacking functional torsinA as well as human fibroblasts isolated from DYT1 dystonia patients. We find that the deletion of torsinA or the expression of torsinA containing the DYT1 dystonia-causing ΔE302/303 (ΔE) mutation results in a more deformable cellular mechanotype. We observe a similar increased deformability of mouse fibroblasts that lack lamina-associated polypeptide 1 (LAP1), which interacts with and stimulates the ATPase activity of torsinA in vitro; as well as with depletion of the LINC complex proteins, Sad1/UNC-84 (SUN)1 and SUN2, lamin A/C, or lamin B1. Moreover, we report that DYT1 dystonia patient-derived fibroblasts are more compliant than fibroblasts isolated from unafflicted individuals. DYT1 fibroblasts also exhibit increased nuclear strain and decreased viability following mechanical stretch. Taken together, our results support a model where the physical connectivity between the cytoskeleton and nucleus contributes to cellular mechanotype. These findings establish the foundation for future mechanistic studies to understand how altered cellular mechanotype may contribute to DYT1 dystonia pathogenesis; this may be particularly relevant in the context of how neurons sense and respond to mechanical forces during traumatic brain injury, which is known to be a major cause of acquired dystonia.

scholarly journals Of numbers and movement – understanding transcription factor pathogenesis by advanced microscopy

2020 ◽  
Vol 13 (12) ◽  
pp. dmm046516
Author(s):  
Julia M. T. Auer ◽  
Jack J. Stoddart ◽  
Ioannis Christodoulou ◽  
Ana Lima ◽  
Kassiani Skouloudaki ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Molecule ◽  
Molecular Mechanisms ◽  
Gene Dosage ◽  
Super Resolution ◽  
Loss Of Function ◽  
Developmental Defects ◽  
Tf Gene

ABSTRACTTranscription factors (TFs) are life-sustaining and, therefore, the subject of intensive research. By regulating gene expression, TFs control a plethora of developmental and physiological processes, and their abnormal function commonly leads to various developmental defects and diseases in humans. Normal TF function often depends on gene dosage, which can be altered by copy-number variation or loss-of-function mutations. This explains why TF haploinsufficiency (HI) can lead to disease. Since aberrant TF numbers frequently result in pathogenic abnormalities of gene expression, quantitative analyses of TFs are a priority in the field. In vitro single-molecule methodologies have significantly aided the identification of links between TF gene dosage and transcriptional outcomes. Additionally, advances in quantitative microscopy have contributed mechanistic insights into normal and aberrant TF function. However, to understand TF biology, TF-chromatin interactions must be characterised in vivo, in a tissue-specific manner and in the context of both normal and altered TF numbers. Here, we summarise the advanced microscopy methodologies most frequently used to link TF abundance to function and dissect the molecular mechanisms underlying TF HIs. Increased application of advanced single-molecule and super-resolution microscopy modalities will improve our understanding of how TF HIs drive disease.

scholarly journals Age and Sex Are Critical Factors in Ischemic Stroke Pathology

Endocrinology ◽  
2018 ◽  
Vol 159 (8) ◽  
pp. 3120-3131 ◽  
Author(s):  
Meaghan Roy-O’Reilly ◽  
Louise D McCullough
Keyword(s):  
Ischemic Stroke ◽  
Stroke Risk ◽  
Current Knowledge ◽  
Clinical Care ◽  
Interactive Effect ◽  
Loss Of Function ◽  
Biological Variables ◽  
Ischemic Stroke Risk ◽  
Age And Sex

Abstract Ischemic stroke is a devastating brain injury resulting in high mortality and substantial loss of function. Understanding the pathophysiology of ischemic stroke risk, mortality, and functional loss is critical to the development of new therapies. Age and sex have a complex and interactive effect on ischemic stroke risk and pathophysiology. Aging is the strongest nonmodifiable risk factor for ischemic stroke, and aged stroke patients have higher mortality and morbidity and poorer functional recovery than their young counterparts. Importantly, patient age modifies the influence of patient sex in ischemic stroke. Early in life, the burden of ischemic stroke is higher in men, but stroke becomes more common and debilitating for women in elderly populations. The profound effects of sex and age on clinical ischemic stroke are mirrored in the results of experimental in vivo and in vitro studies. Here, we review current knowledge on the influence of age and sex in the incidence, mortality, and functional outcome of ischemic stroke in clinical populations. We also discuss the experimental evidence for sex and age differences in stroke pathophysiology and how a better understanding of these biological variables can improve clinical care and enhance development of novel therapies.

scholarly journals Trophoblast development

Reproduction ◽  
2012 ◽  
Vol 143 (3) ◽  
pp. 231-246 ◽  
Author(s):  
Peter L Pfeffer ◽  
David J Pearton
Keyword(s):  
Regulatory Networks ◽  
Current Knowledge ◽  
Embryonic Stem ◽  
Loss Of Function ◽  
Expression Studies ◽  
Gene Regulatory ◽  
Gene Expression Studies ◽  
Function Models

This review summarises current knowledge about the specification, commitment and maintenance of the trophoblast lineage in mice and cattle. Results from gene expression studies, in vivo loss-of-function models and in vitro systems using trophoblast and embryonic stem cells have been assimilated into a model seeking to explain trophoblast ontogeny via gene regulatory networks. While trophoblast differentiation is quite distinct between cattle and mice, as would be expected from their different modes of implantation, recent studies have demonstrated that differences arise much earlier during trophoblast development.

Modulation of Matrix Metalloproteinases by Plant-Derived Products

2020 ◽  
Vol 20 ◽  
Author(s):  
Nur Najmi Mohamad Anuar ◽  
Nurul Iman Natasya Zulkafali ◽  
Azizah Ugusman
Keyword(s):  
Clinical Trials ◽  
Natural Products ◽  
Matrix Metalloproteinases ◽  
Animal Studies ◽  
Current Knowledge ◽  
In Vitro Models ◽  
In Vivo Studies ◽  
Extracellular Matrix Components

: Matrix metalloproteinases (MMPs) are a group of zinc-dependent metallo-endopeptidase that are responsible towards the degradation, repair and remodelling of extracellular matrix components. MMPs play an important role in maintaining a normal physiological function and preventing diseases such as cancer and cardiovascular diseases. Natural products derived from plants have been used as traditional medicine for centuries. Its active compounds, such as catechin, resveratrol and quercetin, are suggested to play an important role as MMPs inhibitors, thereby opening new insights into their applications in many fields, such as pharmaceutical, cosmetic and food industries. This review summarises the current knowledge on plant-derived natural products with MMP-modulating activities. Most of the reviewed plant-derived products exhibit an inhibitory activity on MMPs. Amongst MMPs, MMP-2 and MMP-9 are the most studied. The expression of MMPs is inhibited through respective signalling pathways, such as MAPK, NF-κB and PI3 kinase pathways, which contribute to the reduction in cancer cell behaviours, such as proliferation and migration. Most studies have employed in vitro models, but a limited number of animal studies and clinical trials have been conducted. Even though plant-derived products show promising results in modulating MMPs, more in vivo studies and clinical trials are needed to support their therapeutic applications in the future.

Bone Marrow Niches for Skeletal Progenitor Cells and their Inhabitants in Health and Disease

2019 ◽  
Vol 14 (4) ◽  
pp. 305-319 ◽  
Author(s):  
Marietta Herrmann ◽  
Franz Jakob
Keyword(s):  
Bone Marrow ◽  
Progenitor Cells ◽  
Progenitor Cell ◽  
Adult Stem Cells ◽  
Current Knowledge ◽  
Cell Populations ◽  
Surface Markers ◽  
Bone Marrow Niches

The bone marrow hosts skeletal progenitor cells which have most widely been referred to as Mesenchymal Stem or Stromal Cells (MSCs), a heterogeneous population of adult stem cells possessing the potential for self-renewal and multilineage differentiation. A consensus agreement on minimal criteria has been suggested to define MSCs in vitro, including adhesion to plastic, expression of typical surface markers and the ability to differentiate towards the adipogenic, osteogenic and chondrogenic lineages but they are critically discussed since the differentiation capability of cells could not always be confirmed by stringent assays in vivo. However, these in vitro characteristics have led to the notion that progenitor cell populations, similar to MSCs in bone marrow, reside in various tissues. MSCs are in the focus of numerous (pre)clinical studies on tissue regeneration and repair.Recent advances in terms of genetic animal models enabled a couple of studies targeting skeletal progenitor cells in vivo. Accordingly, different skeletal progenitor cell populations could be identified by the expression of surface markers including nestin and leptin receptor. While there are still issues with the identity of, and the overlap between different cell populations, these studies suggested that specific microenvironments, referred to as niches, host and maintain skeletal progenitor cells in the bone marrow. Dynamic mutual interactions through biological and physical cues between niche constituting cells and niche inhabitants control dormancy, symmetric and asymmetric cell division and lineage commitment. Niche constituting cells, inhabitant cells and their extracellular matrix are subject to influences of aging and disease e.g. via cellular modulators. Protective niches can be hijacked and abused by metastasizing tumor cells, and may even be adapted via mutual education. Here, we summarize the current knowledge on bone marrow skeletal progenitor cell niches in physiology and pathophysiology. We discuss the plasticity and dynamics of bone marrow niches as well as future perspectives of targeting niches for therapeutic strategies.

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