@article {2242, title = {Mechanical instability of adherens junctions overrides intrinsic quiescence of hair follicle stem cells.}, journal = {Dev Cell}, volume = {56}, year = {2021}, month = {2021 Mar 22}, pages = {761-780.e7}, abstract = {

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.

}, issn = {1878-1551}, doi = {10.1016/j.devcel.2021.02.020}, author = {Biswas, Ritusree and Banerjee, Avinanda and Lembo, Sergio and Zhao, Zhihai and Lakshmanan, Vairavan and Lim, Ryan and Le, Shimin and Nakasaki, Manando and Kutyavin, Vassily and Wright, Graham and Palakodeti, Dasaradhi and Ross, Robert S and Jamora, Colin and Vasioukhin, Valeri and Jie, Yan and Raghavan, Srikala} } @article {2310, title = {Scanning electron microscopy of murine skin ultrathin sections and cultured keratinocytes.}, journal = {STAR Protoc}, volume = {2}, year = {2021}, month = {2021 Sep 17}, pages = {100729}, abstract = {

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).

}, issn = {2666-1667}, doi = {10.1016/j.xpro.2021.100729}, author = {Banerjee, Avinanda and Biswas, Ritusree and Lim, Ryan and Pasolli, Hilda Amalia and Raghavan, Srikala} } @article {2200, title = {tRNA-derived fragments (tRFs): establishing their turf in post-transcriptional gene regulation.}, journal = {Cell Mol Life Sci}, year = {2021}, month = {2021 Jan 02}, abstract = {

Transfer RNA (tRNA)-derived fragments (tRFs) are an emerging class of conserved small non-coding RNAs that play important roles in post-transcriptional gene regulation. High-throughput sequencing of multiple biological samples have identified heterogeneous species of tRFs with distinct functionalities. These small RNAs have garnered a lot of scientific attention due to their ubiquitous expression and versatility in regulating various biological processes. In this review, we highlight our current understanding of tRF biogenesis and their regulatory functions. We summarize the diverse modes of biogenesis through which tRFs are generated and discuss the mechanism through which different tRF species regulate gene expression and the biological implications. Finally, we conceptualize research areas that require focus to strengthen our understanding of the biogenesis and function of tRFs.

}, issn = {1420-9071}, doi = {10.1007/s00018-020-03720-7}, author = {Krishna, Srikar and Raghavan, Srikala and DasGupta, Ramanuj and Palakodeti, Dasaradhi} } @article {2201, title = {Staying connected under tension.}, journal = {Science}, volume = {370}, year = {2020}, month = {2020 11 27}, pages = {1036-1037}, keywords = {Mechanotransduction, Cellular, Microfilament Proteins}, issn = {1095-9203}, doi = {10.1126/science.abf2782}, author = {Raghavan, Srikala and Vasioukhin, Valeri} } @article {1740, title = {Dynamic expression of tRNA-derived small RNAs define cellular states.}, journal = {EMBO Rep}, volume = {20}, year = {2019}, month = {2019 Jul}, pages = {e47789}, abstract = {

Transfer RNA (tRNA)-derived small RNAs (tsRNAs) have recently emerged as important regulators of protein translation and shown to have diverse biological functions. However, the underlying cellular and molecular mechanisms of tsRNA function in the context of dynamic cell-state transitions remain unclear. Expression analysis of tsRNAs in distinct heterologous cell and tissue models of stem vs. differentiated states revealed a differentiation-dependent enrichment of 5{\textquoteright}-tsRNAs. We report the identification of a set of 5{\textquoteright}-tsRNAs that is upregulated in differentiating mouse embryonic stem cells (mESCs). Notably, interactome studies with differentially enriched 5{\textquoteright}-tsRNAs revealed a switch in their association with "effector" RNPs and "target" mRNAs in different cell states. We demonstrate that specific 5{\textquoteright}-tsRNAs can preferentially interact with the RNA-binding protein, Igf2bp1, in the RA-induced differentiated state. This association influences the transcript stability and thereby translation of the pluripotency-promoting factor, c-Myc, thus providing a mechanistic basis for how 5{\textquoteright}-tsRNAs can modulate stem cell states in mESCs. Together our study highlights the role of 5{\textquoteright}-tsRNAs in defining distinct cell states.

}, issn = {1469-3178}, doi = {10.15252/embr.201947789}, author = {Krishna, Srikar and Yim, Daniel Gr and Lakshmanan, Vairavan and Tirumalai, Varsha and Koh, Judice Ly and Park, Jung Eun and Cheong, Jit Kong and Low, Joo Leng and Lim, Michelle Js and Sze, Siu Kwan and Shivaprasad, Padubidri and Gulyani, Akash and Raghavan, Srikala and Palakodeti, Dasaradhi and DasGupta, Ramanuj} } @article {1736, title = {Unraveling the ECM-Immune Cell Crosstalk in Skin Diseases.}, journal = {Front Cell Dev Biol}, volume = {7}, year = {2019}, month = {2019}, pages = {68}, abstract = {

The extracellular matrix (ECM) is a complex network of proteins and proteoglycans secreted by keratinocytes, fibroblasts and immune cells. The function of the skin ECM has expanded from being a scaffold that provides structural integrity, to a more dynamic entity that is constantly remodeled to maintain tissue homeostasis. The ECM functions as ligands for cell surface receptors such as integrins, dystroglycans, and toll-like receptors (TLRs) and regulate cellular signaling and immune cell dynamics. The ECM also acts as a sink for growth factors and cytokines, providing critical cues during epithelial morphogenesis. Dysregulation in the organization and deposition of ECMs lead to a plethora of pathophysiological conditions that are exacerbated by aberrant ECM-immune cell interactions. In this review, we focus on the interplay between ECM and immune cells in the context of skin diseases and also discuss state of the art therapies that target the key molecular players involved.

}, issn = {2296-634X}, doi = {10.3389/fcell.2019.00068}, author = {Bhattacharjee, Oindrila and Ayyangar, Uttkarsh and Kurbet, Ambika S and Ashok, Driti and Raghavan, Srikala} } @article {1148, title = {Isolating Immune Cells from Mouse Embryonic Skin.}, journal = {Methods Mol Biol}, year = {2018}, month = {2018 May 24}, abstract = {

Skin is the primary barrier against the external environment and develops a robust immune network for its surveillance. The origin of the resident immune cells of the skin has become a focus of interest over past a decade. Fate mapping studies have revealed that the macrophages home into the skin as early as E12.5 and are derived from the yolk sac and fetal liver. The resident γδT cells are born in the thymus and home to the skin by E16.5. Recent work from our lab has shown that the embryonic macrophages can actively remodel the extracellular matrix in skin suggesting that the skin immune system can be activated long before exposure to foreign antigens. In this chapter, we present a detailed protocol for isolating monocytes, macrophages, and epidermal dendritic T cell populations from embryonic skin.

}, issn = {1940-6029}, doi = {10.1007/7651_2018_148}, author = {Kurbet, Ambika S and Raghavan, Srikala} } @article {1160, title = {Cytoplasmic poly (A)-binding protein critically regulates epidermal maintenance and turnover in the planarian .}, journal = {Development}, volume = {144}, year = {2017}, month = {2017 09 01}, pages = {3066-3079}, abstract = {

Identifying key cellular events that facilitate stem cell function and tissue organization is crucial for understanding the process of regeneration. Planarians are powerful model system to study regeneration and stem cell (neoblast) function. Here, using planaria, we show that the initial events of regeneration, such as epithelialization and epidermal organization are critically regulated by a novel cytoplasmic poly A-binding protein, SMED-PABPC2. Knockdown leads to defects in epidermal lineage specification, disorganization of epidermis and ECM, and deregulated wound healing, resulting in the selective failure of neoblast proliferation near the wound region. Polysome profiling suggests that epidermal lineage transcripts, including , are translationally regulated by SMED-PABPC2 Together, our results uncover a novel role for SMED-PABPC2 in the maintenance of epidermal and ECM integrity, critical for wound healing and subsequent processes for regeneration.

}, keywords = {Animals, Cell Lineage, Cell Proliferation, Cytoplasm, Epidermis, Epithelium, Extracellular Matrix, Gene Knockdown Techniques, Homeostasis, Models, Biological, Planarians, Poly(A)-Binding Protein I, Regeneration, RNA, Messenger, Wound Healing}, issn = {1477-9129}, doi = {10.1242/dev.152942}, author = {Bansal, Dhiru and Kulkarni, Jahnavi and Nadahalli, Kavana and Lakshmanan, Vairavan and Krishna, Srikar and Sasidharan, Vidyanand and Geo, Jini and Dilipkumar, Shilpa and Pasricha, Renu and Gulyani, Akash and Raghavan, Srikala and Palakodeti, Dasaradhi} } @article {1168, title = {Sterile Inflammation Enhances ECM Degradation in Integrin β1 KO Embryonic Skin.}, journal = {Cell Rep}, volume = {16}, year = {2016}, month = {2016 09 20}, pages = {3334-3347}, abstract = {

Epidermal knockout of integrin β1 results in complete disorganization of the basement membrane (BM), resulting in neonatal lethality. Here, we report that this disorganization is exacerbated by an early\ embryonic inflammatory response involving the recruitment of tissue-resident and monocyte-derived macrophages to the dermal-epidermal junction, associated with increased matrix metalloproteinase activity. Remarkably, the skin barrier in the integrin β1 knockout animals is intact, suggesting that this inflammatory response is initiated in a sterile environment. We demonstrate that the molecular mechanism involves de novo expression of integrin αvβ6 in the basal epidermal cells, which activates a TGF-β1 driven inflammatory cascade resulting in upregulation of dermal NF-κB in a Tenascin C-dependent manner. Importantly, treatment of β1 KO embryos in utero with small molecule inhibitors of TGF-βR1 and NF-κB results in marked rescue of the BM defects and amelioration of immune response, revealing an unconventional immuno-protective role for integrin β1 during BM remodeling.

}, keywords = {Animals, Extracellular Matrix, Inflammation, Integrin beta1, Macrophages, Mice, Mice, Knockout, Signal Transduction, Skin}, issn = {2211-1247}, doi = {10.1016/j.celrep.2016.08.062}, author = {Kurbet, Ambika S and Hegde, Samarth and Bhattacharjee, Oindrila and Marepally, Srujan and Vemula, Praveen K and Raghavan, Srikala} }