MicroRNA-205 promotes hair regeneration by modulating mechanical properties of hair follicle stem cells.

Stiffness and actomyosin contractility are intrinsic mechanical properties of animal cells required for the shaping of tissues. However, whether tissue stem cells (SCs) and progenitors located within SC niche have different mechanical properties that modulate their size and function remains unclear. Here, we show that hair follicle SCs in the bulge are stiff with high actomyosin contractility and resistant to size change, whereas hair germ (HG) progenitors are soft and periodically enlarge and contract during quiescence. During activation of hair follicle growth, HGs reduce contraction and more frequently enlarge, a process that is associated with weakening of the actomyosin network, nuclear YAP accumulation, and cell cycle reentry. Induction of miR-205, a novel regulator of the actomyosin cytoskeleton, reduces actomyosin contractility and activates hair regeneration in young and old mice. This study reveals the control of tissue SC size and activities by spatiotemporally compartmentalized mechanical properties and demonstrates the possibility to stimulate tissue regeneration by fine-tuning cell mechanics.

Structural alterations of Lamin A protein in dilated cardiomyopathy.

Lamin A protein, encoded by the LMNA gene, belongs to the type V intermediate filament protein family and is a major nuclear protein component of higher metazoan organisms, including humans. Lamin A along with B-type lamins impart structural rigidity to the nucleus by forming a lamina that is closely apposed to the inner nuclear membrane and is also present as a filamentous network in the interior of the nucleus. A vast number of mutations that lead to a diverse array of at least 11 diseases in humans, collectively termed laminopathies, are being gradually uncovered in the LMNA gene. Dilated cardiomyopathy (DCM) is one such laminopathy in which ventricular dilation leads to an increase in systolic and diastolic volumes, resulting in cardiac arrhythmia and ultimately myocardial infarction. The point mutations in lamin A protein span the entire length of the protein, with a slight preponderance in the central α-helical coiled-coil forming domain. In this work, we have focused on three such important mutations that had been previously observed in DCM-afflicted patients producing severe symptoms. This is the first report to show that these mutations entail significant alterations in the secondary and tertiary structure of the protein, hence perturbing the intrinsic self-association behavior of lamin A protein. Comparison of the enthalpy changes accompanying the deoligomerization process for the wild type and the mutants suggests a difference in the energetics of their self-association. This is further corroborated by the formation of the aggregates of different size and distribution formed inside the nuclei of transfected cells.

Mechanotransduction through adhesion molecules: Emerging roles in regulating the stem cell niche.

Stem cells have been shown to play an important role in regenerative medicine due to their proliferative and differentiation potential. The challenge, however, lies in regulating and controlling their potential for this purpose. Stem cells are regulated by growth factors as well as an array of biochemical and mechanical signals. While the role of biochemical signals and growth factors in regulating stem cell homeostasis is well explored, the role of mechanical signals has only just started to be investigated. Stem cells interact with their niche or to other stem cells via adhesion molecules that eventually transduce mechanical cues to maintain their homeostatic function. Here, we present a comprehensive review on our current understanding of the influence of the forces perceived by cell adhesion molecules on the regulation of stem cells. Additionally, we provide insights on how this deeper understanding of mechanobiology of stem cells has translated toward therapeutics.

Scanning electron microscopy of murine skin ultrathin sections and cultured keratinocytes.

Generating high-quality electron microscopy images of the skin and keratinocytes can be challenging. Here we describe a simple protocol for scanning electron microscopy (SEM) of murine skin. The protocol enables characterization of the ultrastructure of the epidermis, dermis, hair follicles, basement membrane, and cell-cell junctions. We detail the specific steps for sample preparation and highlight the critical need for proper orientation of the sample for ultrathin sectioning. We also describe the isolation and preparation of primary keratinocyte monolayers for SEM. For complete details on the use and execution of this protocol, please refer to Biswas et al. (2021).

Mechanical instability of adherens junctions overrides intrinsic quiescence of hair follicle stem cells.

Vinculin, a mechanotransducer associated with both adherens junctions (AJs) and focal adhesions (FAs), plays a central role in force transmission through cell-cell and cell-substratum contacts. We generated the conditional knockout (cKO) of vinculin in murine skin that results in the loss of bulge stem cell (BuSC) quiescence and promotes continual cycling of the hair follicles. Surprisingly, we find that the AJs in vinculin cKO cells are mechanically weak and impaired in force generation despite increased junctional expression of E-cadherin and α-catenin. Mechanistically, we demonstrate that vinculin functions by keeping α-catenin in a stretched/open conformation, which in turn regulates the retention of YAP1, another potent mechanotransducer and regulator of cell proliferation, at the AJs. Altogether, our data provide mechanistic insights into the hitherto-unexplored regulatory link between the mechanical stability of cell junctions and contact-inhibition-mediated maintenance of BuSC quiescence.

Novel linkage of LMNA Single Nucleotide Polymorphism with Dilated Cardiomyopathy in an Indian case study.

BACKGROUND: Dilated Cardiomyopathy (DCM) is one of the most commonly encountered heart diseases reported globally. It is characterized by enlarged ventricles with impaired systolic and diastolic functions. Mutations in LMNA gene are one of the causative factors to precipitate the disease. However, association of SNPs of LMNA with DCM in particular has not been well documented. METHOD: Here we present a limited and restricted case study of patients from south eastern part of India afflicted with idiopathic DCM and conduction defects. By using next generation sequencing we have sequenced the exons of LMNA gene from genomic DNA isolated from patients. RESULT: We have identified the linkage of 8 different LMNA SNPs with idiopathic DCM viz. rs121117552, rs538089, rs505058, rs4641, rs646840, rs534807, rs80356803 and rs7339. These SNPs are scattered throughout the gene with prevalence for the region encoding the central rod domain of lamin A/C. CONCLUSION: Most of these SNPs in LMNA were previously reported to be involved in various disorders other than DCM. We conclude that, variation in LMNA is one of the major underlying genetic causes for the pathogenesis of DCM, as observed in few Indian populations.

Viscoelastic behavior of human lamin A proteins in the context of dilated cardiomyopathy.

Lamins are intermediate filament proteins of type V constituting a nuclear lamina or filamentous meshwork which lines the nucleoplasmic side of the inner nuclear membrane. This protein mesh provides a supporting scaffold for the nuclear envelope and tethers interphase chromosome to the nuclear periphery. Mutations of mainly A-type lamins are found to be causative for at least 11 human diseases collectively termed as laminopathies majority of which are characterised by aberrant nuclei with altered structural rigidity, deformability and poor mechanotransduction behaviour. But the investigation of viscoelastic behavior of lamin A continues to elude the field. In order to address this problem, we hereby present the very first report on viscoelastic properties of wild type human lamin A and some of its mutants linked with Dilated cardiomyopathy (DCM) using quantitative rheological measurements. We observed a dramatic strain-softening effect on lamin A network as an outcome of the strain amplitude sweep measurements which could arise from the large compliance of the quasi-cross-links in the network or that of the lamin A rods. In addition, the drastic stiffening of the differential elastic moduli on superposition of rotational and oscillatory shear stress reflect the increase in the stiffness of the laterally associated lamin A rods. These findings present a preliminary insight into distinct biomechanical properties of wild type lamin A protein and its mutants which in turn revealed interesting differences.