Abstract

Nowadays, it is an established concept that the capability to reach a specialised cell identity via differentiation, as in the case of multi- and pluripotent stem cells, is not only determined by biochemical factors, but that also physical aspects of the microenvironment play a key role; interpreted by the cell through a force-based signalling pathway called mechanotransduction. However, the intricate ties between the elements involved in mechanotransduction, such as the extracellular matrix, the glycocalyx, the cell membrane, Integrin adhesion complexes, Cadherin-mediated cell/cell adhesion, the cytoskeleton, and the nucleus, are still far from being understood in detail. Here we report what is currently known about these elements in general and their specific interplay in the context of multi- and pluripotent stem cells. We furthermore merge this overview to a more comprehensive picture, that aims to cover the whole mechanotransductive pathway from the cell/microenvironment interface to the regulation of the chromatin structure in the nucleus. Ultimately, with this review we outline the current picture of the interplay between mechanotransductive cues and epigenetic regulation and how these processes might contribute to stem cell dynamics and fate.

Introduction

One of the basic principles enabling multicellular life is the differentiation of stem cells (SCs), into specialised cells with a defined identity, shape, and function ( McBeath et al., 2004 ) ( Tewary et al., 2018 ). SCs are defined as cells that are clonogenic, which defines their capability of both, self-renewing and retaining multilineage differentiation potency. In vertebrates, the Zygote represents the earliest stem cell (totipotent) in ontogeny, from which originate the pluripotent stem cells (PSCs) of the inner cell mass of the blastocyst ( Dean et al., 2003 , Smith, 2017 ). Soon thereafter, PSCs continue to expand during embryo development becoming progressively committed in their cell fate and acquiring a more restricted potency ( Dean et al., 2003 , Osorno et al., 2012 , Smith, 2017 ). Even at the end of embryogenesis, when the organism is fully formed, and its organs are composed of functionally mature cells, some SCs are still present. These cells, which possess a more limited differentiating potential (multipotent), in turn, give rise to a progeny of precursor cells and, finally, to functionally mature cells. Some examples are represented by the hematopoietic SCs in the bone marrow, the intestinal SCs in the gut’s crypts, the epithelial SCs in the epidermis or the mesenchymal stem cells (MSCs) which play an important role in the maintenance and regeneration of bones and cartilage ( Barker, 2014 , Evans et al., 2013 , Guan et al., 2012 , Tewary et al., 2018 , Watt, 2002 , Weissman, 2000 ).

The inherent high level of plasticity characterising SCs is at the base of their capability to respond precisely to appropriate stimuli with differentiative behaviour. Understanding this essential principle of multicellular life; i.e., the regulation of cell identity and fate, is one of the most important fields of research in modern cell biology, primarily due to its impact on the development of new therapeutic approaches in regenerative medicine. Such knowledge can directly benefit stem cell bioengineering by leveraging the SC plasticity and control over SC behaviour for the application in tissue engineering, patient-specific disease modelling and cell therapies ( Tewary et al., 2018 ).

Conclusions

In recent years there has been a fascinating expansion of our knowledge about the interplay between mechanotransductive signalling and chromatin regulation, and how this impacts on multi- and pluripotent stem cell dynamics, which we try to highlight in this review.

However, what shapes the life of a cell and its identity, from the outside (biophysical and structural cues of the cellular microenvironment) and from the inside (nuclear and chromatin organisation) and how it is connected (mechanotransduction and epigenetics) are highly relevant biological issues that are yet far from being understood in detail. Starting from the stem cell/microenvironment interface, a much more detailed understanding of the nanoscale events in this interface is necessary to unravel 1) what the SC perceives of the biophysical nature and the 3D adhesion site configuration and 2) how the immanent nanoscale information is interpreted and converted, through force-based actions, into a remodelling at the level of the IACs, cell/cell adhesions, cytoskeleton and nucleus.

1- Introduction

2- Mechanisms regulating MSC differentiation

3- Pericytes as pre-programmed MSC precursors in vivo

4- Discussion and future considerations

References

Abstract

The increasing application of approaches that allow tracing of individual cells over time, together with transcriptomic and epigenomic analyses is changing the way resident stromal stem cells (mesenchymal stem cells) are viewed. Rather than being a defined, homogeneous cell population as described following in vitro expansion, in vivo, these cells are highly programmed according to their resident tissue location. This programming is evidenced by different epigenetic landscapes and gene transcription signatures in cells before any in vitro expansion. This has potentially profound implications for the heterotypic use of these cells in therapeutic tissue engineering applications.

Introduction

The prototypical mesenchymal stem cell (MSC), isolated from bone marrow (BM) was described over 3 decades ago by Friedenstein [1]. This characterisation was later expanded to include a cell type, isolated from connective tissue stroma that can exhibit stem cell properties in vitro [2, 3]. These observations led many researchers to identify cells in multiple organs that had a similar immunophenotype and characteristic s in vitro of MSCs in that they could be differentiated to form osteoblast -like, chondrocyte -like, and adipocyte -like cells. Tissues included, connective tissue of teeth [4, 5] , muscle [6], adipose [7], dermis [8], heart and liver [9]. These cell populations expressed a number of the by now established MSC gene markers and all shared some common characteristics including morphology, adherence to tissue culture plastic, and trilineage differentiation [9, 10].

Background: Virtually all eukaryotic cells contain spatially distinct genomes, a single nuclear genome that harbours the vast majority of genes and much smaller genomes found in mitochondria present at thousands of copies per cell. To generate a coordinated gene response to various environmental cues, the genomes must communicate with each another. Much of this bi-directional crosstalk relies on epigenetic processes, including DNA, RNA, and histone modification pathways. Crucially, these pathways, in turn depend on many metabolites generated in specific pools throughout the cell, including the mitochondria. They also involve the transport of metabolites as well as the enzymes that catalyse these modifications between nuclear and mitochondrial genomes. Scope of review: This study examines some of the molecular mechanisms by which metabolites influence the activity of epigenetic enzymes, ultimately affecting gene regulation in response to metabolic cues. We particularly focus on the subcellular localisation of metabolite pools and the crosstalk between mitochondrial and nuclear proteins and RNAs. We consider aspects of mitochondrial-nuclear communication involving histone proteins, and potentially their epigenetic marks, and discuss how nuclear-encoded enzymes regulate mitochondrial function through epitranscriptomic pathways involving various classes of RNA molecules within mitochondria. Major conclusions: Epigenetic communication between nuclear and mitochondrial genomes occurs at multiple levels, ultimately ensuring a coordinated gene expression response between different genetic environments. Metabolic changes stimulated, for example, by environmental factors, such as diet or physical activity, alter the relative abundances of various metabolites, thereby directly affecting the epigenetic machinery. These pathways, coupled to regulated protein and RNA transport mechanisms, underpin the coordinated gene expression response. Their overall importance to the fitness of a cell is highlighted by the identification of many mutations in the pathways we discuss that have been linked to human disease including cancer.

Introduction

To fit within the confined space of a eukaryotic nucleus, a cell’s genome must be efficiently compacted, but in a highly ordered manner that maintains accessibility to the genetic information. This is achieved by DNA complexing with histone proteins to form nucleosome structures, which in turn further compact to form chromatin that ultimately packages entire chromosomes. Importantly, chromatin is not an inert packaging structure but rather an instructive scaffold capable of responding to various cues to regulate access of the DNA to different cellular machineries. This accessibility is fundamentally regulated by ATP-dependent chromatin remodelling activities and by covalent modification of both DNA and histone proteins. These modifications are commonly referred to as “epigenetic” modifications. Furthermore, in addition to epigenetics, the emerging field of epitranscriptomics is now adding numerous modifications of RNA molecules to the “modification repertoire,” thereby expanding our current view of epigenetics and how it regulates all DNA processes, in particular gene expression [1]. DNA and histone modifications regulate gene expression by modulating chromatin accessibility and/or providing binding sites for regulatory proteins. To date, more than 100 post-translational modifications (PTMs) have been detected on histones, including acetylation, methylation, ubiquitylation, phosphorylation, citrullination, and O-GlcNAcylation. Of these, acetylation and methylation are by far the best-studied and most characterised histone modifications (for review, see [2,3]).

Abstract

We review recent advances in the DNA sequencing method based on measurements of transverse electrical currents. Device configurations proposed in the literature are classified according to whether the molecular fingerprints appear as the major (Mode I) or perturbing (Mode II) current signals. Scanning tunneling microscope and tunneling electrode gap configurations belong to the former category, while the nanochannels with or without an embedded nanopore belong to the latter. The molecular sensing mechanisms of Modes I and II roughly correspond to the electron tunneling and electrochemical gating, respectively. Special emphasis will be given on the computer simulation studies, which have been playing a critical role in the initiation and development of the field. We also highlight low-dimensional nanomaterials such as carbon nanotubes, graphene, and graphene nanoribbons that allow the novel Mode II approach. Finally, several issues in previous computational studies are discussed, which points to future research directions toward more reliable simulation of electrical current DNA sequencing devices.

Introduction

The sequencing of DNA not only has significant scientific implications in the context of deciphering the fundamental code of life but also represents an enormous opportunity to improve the well-being of humankind by ushering in a new era of personal or precision medicine (Mardis, 2011; Shendure and Aiden, 2012; Rabbani et al., 2014). A decade after the completion of the Human Genome Project in 2003, effort is now devoted into the development of next-generation DNA sequencing technologies that can meet the ‘$1000 genome’ goal set by National Institute of Health. In this endeavor, in contrast to the second-generation DNA sequencing technologies that still require polymerase chain reaction amplification and fluorescent labeling as in the first-generation counterpart, the newly-emerged third-generation DNA sequencing technologies propose single molecule detection based on changes in ionic or electrical currents

Abstract

Ultra-thin solid-state nanopore with good wetting property is strongly desired to achieve high spatial resolution for DNA sequencing applications. Atomic thick hexagonal boron nitride (h-BN) layer provides a promising two-dimensional material for fabricating solid-state nanopores. Due to its good oxidation resistance, the hydrophilicity of h-BN nanopore device can be significantly improved by UV-Ozone treatment. The contact angle of a KCl-TE droplet on h-BN layer can be reduced from 57° to 26° after the treatment. Abundant DNA translocation events have been observed in such devices, and strong DNA-nanopore interaction has been revealed in pores smaller than 10 nm in diameter. The 1/f noise level is closely related to the area of suspended h-BN layer, and it is significantly reduced in smaller supporting window. The demonstrated performance in h-BN nanopore paves the way towards base discrimination in a single DNA molecule.

Introduction

As one of the most promising concepts, DNA sequencing with nanopore has attracted a lot of attention since 19961. DNA translocation1,2,3 and base discrimination4 has been demonstrated in biological nanopores made of specific proteins. Solid-state nanopores, however, are strongly preferred for higher stability and easier device integration. The first solid-state nanopores are fabricated in silicon nitride (Si3N4) membranes that are tens of nanometers thick5, and significant progress has been made in detection of DNA translocation6,7, and DNA motion control8 in such nanopore devices. To achieve base discrimination in a single DNA molecule, the effective thickness of a solid-state nanopore has to be reduced down to sub-nanometer. For such propose, atomic thick graphene layers has been employed in fabricating nanopore devices featured with high resolution and geometrical sensitivity9,10,11,12. On the other hand, the hydrophilicity of nanopore is practically important to avoid clogging due to strong nonspecific hydrophobic interaction with DNA. Recently, the hydrophilicity of graphene nanopores has been improved by atomic-layer deposition of titanium dioxide11 and self-assembled hydrophilic coating13, which however increases the effective thickness of the pores. New types of atomic thick nanopores with high hydrophilicity are thus still strongly desired.

Major depressive disorder is characterized by persistence of sadness, loss of interest or pleasure, feelings of guilt or low self-worth, and other symptoms, including sleep and circadian changes, changes in weight or appetite, and diminished concentration. More than 300 million people worldwide are living with depression, making it the leading global cause of disability.1 Depression can affect people at any age, and can persist or reoccur throughout the lifespan. Previous experience of depression increases the likelihood of future episodes. Women are approximately 50% more likely than men to experience depression, with increased risk associated with puberty, peripartum periods, and menopause. As with other psychiatric disorders, depression is a complex, heterogeneous syndrome. The causes of depression cannot be pinned down to any one biological or external factor, and instead result from the interplay of genetic, neurobiological, environmental, and cultural factors. The genetic heritability of depression is estimated to be from 35%–40%.2,3 While heritable genetic factors clearly play a role, a significant proportion of depression risk is attributed to life experience and environment, particularly previous and/or recent stress experience. In the last decade, there has been a considerable focus on the role of epigenetics as a possible bridge between genes and experience in the pathophysiology of depression. The epigenetic landscape, originally envisioned by Waddington more than 60 years ago, sits beautifully at the intersection of genetics, development, environmental influence, and inheritance.4 The word “epigenetic” means “above the genome” and epigenetic mechanisms refer to structural and molecular factors that alter expression of genes without altering the sequence of DNA. These include many types of chemical modifications to histone proteins, DNA, and RNA, as well as alterations in other regulatory proteins and noncoding RNAs.5–7 While epigenetic mechanisms were previously defined as stable through cell division, the dynamic nature of epigenetic modifications is now understood. As primary regulators of gene expression, epigenetic mechanisms are ideal molecular candidates to flexibly encode and translate environmental factors that increase risk for depression and other highly complex psychiatric syndromes.

The advancements in the cellular and molecular biology have played a tremendous role in our understanding of the complex clinical disorders. Epigenetics is one of the fields, which has been studied extensively in various areas of health research like neurosciences, cancer medicine etc. The term epigenetics was coined in 1942 which refers to changes in the gene expression that results from external or environmental factors, and such changes are not encoded in DNA sequence[1]. The DNA encodes the secrets of life in two specific layers of information. The first layer of information which forms the basis of our genetic code is called the coding region of DNA. This coding region comprises only 2% portion of total DNA and remaining 98% region is called noncoding region. The genetic code is translated to different mRNAs and is responsible for creating all the proteins that we need to function. The second layer which is above this genetic code known as the epigenetic layer is responsible for deciding when, where and which gene to express. This epigenetic layer comprises the different epigenetic mechanisms including DNA Methylation, Histone Modifications or non-coding RNAs. Epigenetic mechanisms are crucial for cellular differentiation, and this process is highly conserved and controlled at the cellular level in each particular organism. When such highly conserved process is affected by the external environment directly or indirectly, it leads to change in the expression of genes and thus function. For example, differentiation of human CD4+ T cells is influenced by the locally acting inflammatory substances[2]. These inflammatory substances or interleukins further activate transcription factors that bind to specific methylated or demethylated sites in the promoter regions of specific genes in a developing T cell DNA and decide the fate of lineage of T cell development[3]. The increase in DNA methylation close to the promoter region of the specific gene results in decreased expression and vice versa[3]. Similarly, in pathological conditions like periodontitis, the local immune response is decided by these locally acting transcription factors and methylation or demethylation of specific genes[4].

Multiple evolutionarily conserved T cell populations with innate characteristics are present in mice and humans that differ in their antigen receptor expression and their functional properties [1,2]. In mice, invariant Natural Killer T cells (iNKT), which express an invariant Va14-Ja18 T cell receptor alpha (TCRa) chain that can pair with a limited number of TCRb chains (Vb2, Vb7 and Vb8.1–8.2), are the most abundant innate T cell population. These cells recognize glycolipid antigens presented by the non-classical, non-polymorphic Major Histocompatibility Complex (MHC) I family molecule CD1D. The majority of iNKT cells follow the same developmental trajectories in the thymus as conventional T cells; however, iNKT cells mature in the thymus and acquire a phenotype reminiscent of antigen-experienced T cells during development. The frequency and number of iNKT effector subsets varies between naı¨ve mouse strains and human individuals and influences the fate of bystander immune cells, even at the steady state [3–5]. Due to their poised state, iNKT cells produce a vast amount of a variety of cytokines rapidly after activation, thereby influencing the functions of a number of innate and adaptive immune cells. iNKT cells have been implicated as positive or negative regulators of a wide range of diseased states, including microbial infection, cancer and autoimmunity and they are being targeted in immune therapies and vaccination strategies [6,7].