Regulation of ERK basal and pulsatile activity control proliferation and exit from the stem cell compartment in mammalian epidermis

Fluctuation in signal transduction pathways is frequently observed during mammalian development. However, its role in regulating stem cells has not been explored. Here we tracked spatiotemporal ERK MAPK dynamics in human epidermal stem cells. While stem cells and differentiated cells were distinguished by high and low stable basal ERK activity, respectively, we also found cells with pulsatile ERK activity. Transitions from Basalhi-Pulselo(stem) to Basalhi-Pulsehi, Basalmid-Pulsehi, and Basallo-Pulselo(differentiated) cells occurred in expanding keratinocyte colonies and in response to differentiation stimuli. Pharmacological inhibition of ERK induced differentiation only when cells were in the Basalmid-Pulsehistate. Basal ERK activity and pulses were differentially regulated by DUSP10 and DUSP6, leading us to speculate that DUSP6-mediated ERK pulse down-regulation promotes initiation of differentiation, whereas DUSP10-mediated down-regulation of mean ERK activity promotes and stabilizes postcommitment differentiation. Levels of MAPK1/MAPK3 transcripts correlated with DUSP6 and DUSP10 transcripts in individual cells, suggesting that ERK activity is negatively regulated by transcriptional and posttranslational mechanisms. When cells were cultured on a topography that mimics the epidermal−dermal interface, spatial segregation of mean ERK activity and pulses was observed. In vivo imaging of mouse epidermis revealed a patterned distribution of basal cells with pulsatile ERK activity, and down-regulation was linked to the onset of differentiation. Our findings demonstrate that ERK MAPK signal fluctuations link kinase activity to stem cell dynamics.

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ERK basal and pulsatile activity are differentially regulated in mammalian epidermis to control proliferation and exit from the stem cell compartment

10.1101/2020.04.12.038406 ◽

2020 ◽

Cited By ~ 1

Author(s):

Toru Hiratsuka ◽

Ignacio Bordeu ◽

Gunnar Pruessner ◽

Fiona M. Watt

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Control Cell ◽

Spatial Segregation ◽

Mouse Epidermis ◽

Cell Compartment ◽

Erk Mapk ◽

Differentiated Cells ◽

Stem Cell Compartment ◽

Erk Activity

AbstractFluctuation in signal transduction pathways is frequently observed during mammalian development. However, its role in regulating stem cells has not been explored. Here we tracked spatiotemporal ERK MAPK dynamics in human epidermal stem cells. While stem cells and differentiated cells were distinguished by high and low stable basal ERK activity, respectively, we also found cells with pulsatile ERK activity. Transitions from Basalhi-Pulselo (stem) to Basalhi-Pulsehi, Basalmid-Pulsehi, and Basallo-Pulselo (differentiated) cells occurred in expanding keratinocyte colonies and in response to a range of differentiation stimuli. Pharmacological inhibition of ERK induced differentiation only when cells were in the Basalmid-Pulsehi state. Basal ERK activity and pulses were differentially regulated by DUSP10 and DUSP6, leading us to speculate that DUSP6-mediated ERK pulse downregulation promotes initiation of differentiation whereas DUSP10-mediated downregulation of mean ERK activity promotes and stabilizes post-commitment differentiation. Quantification of MAPK1/3, DUSP6 and DUSP10 transcripts in individual cells demonstrated that ERK activity is controlled both transcriptionally and post-transcriptionally. When cells were cultured on a topography that mimics the epidermal-dermal interface, spatial segregation of mean ERK activity and pulses was observed. In vivo imaging of mouse epidermis revealed a patterned distribution of basal cells with pulsatile ERK activity and downregulation was linked to the onset of differentiation. Our findings demonstrate that ERK MAPK signal fluctuations link kinase activity to stem cell dynamics.SignificanceUnderstanding how intracellular signaling cascades control cell fate is a key issue in stem cell biology. Here we show that exit from the stem cell compartment in mammalian epidermis is characterised by pulsatile ERK MAPK activity. Basal activity and pulses are differentially regulated by DUSP10 and DUSP6, two phosphatases that have been shown previously to regulate differentiation commitment in the epidermis. ERK activity is controlled both transcriptionally and post-transcriptionally. Spatial segregation of mean ERK activity and pulses is observed both in reconstituted human epidermis and in mouse epidermis. Our findings demonstrate the tight spatial and temporal regulation of ERK MAPK expression and activity in mammalian epidermis.

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Differentiation-related expression of a major 64K corneal keratin in vivo and in culture suggests limbal location of corneal epithelial stem cells.

The Journal of Cell Biology ◽

10.1083/jcb.103.1.49 ◽

1986 ◽

Vol 103 (1) ◽

pp. 49-62 ◽

Cited By ~ 830

Author(s):

A Schermer ◽

S Galvin ◽

T T Sun

Keyword(s):

Stem Cells ◽

Strong Support ◽

Transitional Zone ◽

Basal Cells ◽

Epithelial Stem Cells ◽

Corneal Epithelial ◽

Differentiated Cells ◽

Limbal Epithelium ◽

Cell Layers

In this paper we present keratin expression data that lend strong support to a model of corneal epithelial maturation in which the stem cells are located in the limbus, the transitional zone between cornea and conjunctiva. Using a new monoclonal antibody, AE5, which is highly specific for a 64,000-mol-wt corneal keratin, designated RK3, we demonstrate that this keratin is localized in all cell layers of rabbit corneal epithelium, but only in the suprabasal layers of the limbal epithelium. Analysis of cultured corneal keratinocytes showed that they express sequentially three major keratin pairs. Early cultures consisting of a monolayer of "basal" cells express mainly the 50/58K keratins, exponentially growing cells synthesize additional 48/56K keratins, and postconfluent, heavily stratified cultures begin to express the 55/64K corneal keratins. Cell separation experiments showed that basal cells isolated from postconfluent cultures contain predominantly the 50/58K pair, whereas suprabasal cells contain additional 55/64K and 48/56K pairs. Basal cells of the older, postconfluent cultures, however, can become AE5 positive, indicating that suprabasal location is not a prerequisite for the expression of the 64K keratin. Taken together, these results suggest that the acidic 55K and basic 64K keratins represent markers for an advanced stage of corneal epithelial differentiation. The fact that epithelial basal cells of central cornea but not those of the limbus possess the 64K keratin therefore indicates that corneal basal cells are in a more differentiated state than limbal basal cells. These findings, coupled with the known centripetal migration of corneal epithelial cells, strongly suggest that corneal epithelial stem cells are located in the limbus, and that corneal basal cells correspond to "transient amplifying cells" in the scheme of "stem cells----transient amplifying cells----terminally differentiated cells."

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Mammalian genes induce partially reprogrammed pluripotent stem cells in non-mammalian vertebrate and invertebrate species

eLife ◽

10.7554/elife.00036 ◽

2013 ◽

Vol 2 ◽

Cited By ~ 41

Author(s):

Ricardo Antonio Rosselló ◽

Chun-Chun Chen ◽

Rui Dai ◽

Jason T Howard ◽

Ute Hochgeschwender ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Pluripotent Stem Cells ◽

Mammalian Species ◽

Embryonic Stem ◽

Model Organisms ◽

Differentiated Cells ◽

Transcription Factor Genes ◽

Induced Pluripotent

Cells are fundamental units of life, but little is known about evolution of cell states. Induced pluripotent stem cells (iPSCs) are once differentiated cells that have been re-programmed to an embryonic stem cell-like state, providing a powerful platform for biology and medicine. However, they have been limited to a few mammalian species. Here we found that a set of four mammalian transcription factor genes used to generate iPSCs in mouse and humans can induce a partially reprogrammed pluripotent stem cell (PRPSCs) state in vertebrate and invertebrate model organisms, in mammals, birds, fish, and fly, which span 550 million years from a common ancestor. These findings are one of the first to show cross-lineage stem cell-like induction, and to generate pluripotent-like cells for several of these species with in vivo chimeras. We suggest that the stem-cell state may be highly conserved across a wide phylogenetic range.

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Designing and Engineering Stem Cell Niches

MRS Bulletin ◽

10.1557/mrs2010.527 ◽

2010 ◽

Vol 35 (8) ◽

pp. 591-596 ◽

Cited By ~ 5

Author(s):

Ana I. Teixeira ◽

Ola Hermanson ◽

Carsten Werner

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Biology ◽

Chemical Engineering ◽

Polyglycolic Acid ◽

Differentiated Cells ◽

Extracellular Matrix Components ◽

Stem Cell Niches

AbstractStem cells have received a lot of attention due to great promises in medical treatment, for example, by replacing lost and sick cells and re-constituting cell populations. There are several classes of stem cells, including embryonic, fetal, and adult tissue specific. More recently, the generation of so-called induced pluripotent stem (iPS) cells from differentiated cells has been established. Common criteria for all types of stem cells include their ability to self-renew and to retain their ability to differentiate in response to specific cues. These characteristics, as well as the instructive steering of the cells into differentiation, are largely dependent on the microenvironment surrounding the cells. Such “stem cell friendly” microenvironments, provided by structural and biochemical components, are often referred to as niches. Biomaterials offer attractive solutions to engineer functional stem cell niches and to steer stem cell state and fatein vitroas well asin vivo. Among materials used so far, promising results have been achieved with low-toxicity and biodegradable polymers, such as polyglycolic acid and related materials, as well as other polymers used as structural “scaffolds” for engineering of extracellular matrix components. To improve the efficiency of stem cell control and the design of the biomaterials, interfaces among stem cell research, developmental biology, regenerative medicine, chemical engineering, and materials research are rapidly developing. Here we provide an introduction to stem cell biology and principles of niche engineering and give an overview of recent advancements in stem cell niche engineering from two stem cell systems—blood and brain.

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Serum-Free Manufacturing of Mesenchymal Stem Cell Tissue Rings Using Human-Induced Pluripotent Stem Cells

Stem Cells International ◽

10.1155/2019/5654324 ◽

2019 ◽

Vol 2019 ◽

pp. 1-11 ◽

Cited By ~ 1

Author(s):

Tackla S. Winston ◽

Kantaphon Suddhapas ◽

Chenyan Wang ◽

Rafael Ramos ◽

Pranav Soman ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Regenerative Medicine ◽

Induced Pluripotent Stem Cells ◽

Pluripotent Stem Cells ◽

Culture Conditions ◽

Serum Free ◽

Differentiated Cells ◽

Induced Pluripotent

Combination of stem cell technology and 3D biofabrication approaches provides physiological similarity to in vivo tissues and the capability of repairing and regenerating damaged human tissues. Mesenchymal stem cells (MSCs) have been widely used for regenerative medicine applications because of their immunosuppressive properties and multipotent potentials. To obtain large amount of high-quality MSCs without patient donation and invasive procedures, we differentiated MSCs from human-induced pluripotent stem cells (hiPSC-MSCs) using serum-free E6 media supplemented with only one growth factor (bFGF) and two small molecules (SB431542 and CHIR99021). The differentiated cells showed a high expression of common MSC-specific surface markers (CD90, CD73, CD105, CD106, CD146, and CD166) and a high potency for osteogenic and chondrogenic differentiation. With these cells, we have been able to manufacture MSC tissue rings with high consistency and robustness in pluronic-coated reusable PDMS devices. The MSC tissue rings were characterized based on inner diameter and outer ring diameter and observed cell-type-dependent tissue contraction induced by cell-matrix interaction. Our approach of simplified hiPSC-MSC differentiation, modular fabrication procedure, and serum-free culture conditions has a great potential for scalable manufacturing of MSC tissue rings for different regenerative medicine applications.

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Resolving stem and progenitor cells in the adult mouse incisor through gene co-expression analysis

eLife ◽

10.7554/elife.24712 ◽

2017 ◽

Vol 6 ◽

Cited By ~ 19

Author(s):

Kerstin Seidel ◽

Pauline Marangoni ◽

Cynthia Tang ◽

Bahar Houshmand ◽

Wen Du ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Expression Analysis ◽

Adult Mouse ◽

Deep Understanding ◽

Lineage Tracing ◽

Differentiated Cells ◽

Mouse Incisor ◽

Stem Cell Niches

Investigations into stem cell-fueled renewal of an organ benefit from an inventory of cell type-specific markers and a deep understanding of the cellular diversity within stem cell niches. Using the adult mouse incisor as a model for a continuously renewing organ, we performed an unbiased analysis of gene co-expression relationships to identify modules of co-expressed genes that represent differentiated cells, transit-amplifying cells, and residents of stem cell niches. Through in vivo lineage tracing, we demonstrated the power of this approach by showing that co-expression module members Lrig1 and Igfbp5 define populations of incisor epithelial and mesenchymal stem cells. We further discovered that two adjacent mesenchymal tissues, the periodontium and dental pulp, are maintained by distinct pools of stem cells. These findings reveal novel mechanisms of incisor renewal and illustrate how gene co-expression analysis of intact biological systems can provide insights into the transcriptional basis of cellular identity.

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A Novel Mouse Model for Non-Invasive Single Marker Tracking of Mammary Stem Cells In Vivo Reveals Stem Cell Dynamics throughout Pregnancy

PLoS ONE ◽

10.1371/journal.pone.0008035 ◽

2009 ◽

Vol 4 (11) ◽

pp. e8035 ◽

Cited By ~ 14

Author(s):

Benjamin J. Tiede ◽

Leah A. Owens ◽

Feng Li ◽

Christina DeCoste ◽

Yibin Kang

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Mouse Model ◽

Mammary Stem Cells ◽

Cell Dynamics ◽

Marker Tracking ◽

Non Invasive ◽

Mammary Stem ◽

Single Marker

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Transcription factor p63 controls the reserve status but not the stemness of horizontal basal cells in the olfactory epithelium

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1512272112 ◽

2015 ◽

Vol 112 (36) ◽

pp. E5068-E5077 ◽

Cited By ~ 37

Author(s):

Nikolai Schnittke ◽

Daniel B. Herrick ◽

Brian Lin ◽

Jesse Peterson ◽

Julie H. Coleman ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Olfactory Epithelium ◽

Tissue Injury ◽

Cell Types ◽

Stem Cell Population ◽

Basal Cells ◽

Loss Of Function ◽

Down Regulation ◽

Horizontal Basal Cells

Adult tissue stem cells can serve two broad functions: to participate actively in the maintenance and regeneration of a tissue or to wait in reserve and participate only when activated from a dormant state. The adult olfactory epithelium, a site for ongoing, life-long, robust neurogenesis, contains both of these functional stem cell types. Globose basal cells (GBCs) act as the active stem cell population and can give rise to all the differentiated cells found in the normal tissue. Horizontal basal cells (HBCs) act as reserve stem cells and remain dormant unless activated by tissue injury. Here we show that HBC activation following injury by the olfactotoxic gas methyl bromide is coincident with the down-regulation of protein 63 (p63) but anticipates HBC proliferation. Gain- and loss-of-function studies show that this down-regulation of p63 is necessary and sufficient for HBC activation. Moreover, activated HBCs give rise to GBCs that persist for months and continue to act as bona fide stem cells by participating in tissue maintenance and regeneration over the long term. Our analysis provides mechanistic insight into the dynamics between tissue stem cell subtypes and demonstrates that p63 regulates the reserve state but not the stem cell status of HBCs.

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aPKCλ controls epidermal homeostasis and stem cell fate through regulation of division orientation

The Journal of Cell Biology ◽

10.1083/jcb.201307001 ◽

2013 ◽

Vol 202 (6) ◽

pp. 887-900 ◽

Cited By ~ 48

Author(s):

Michaela T. Niessen ◽

Jeanie Scott ◽

Julia G. Zielinski ◽

Susanne Vorhagen ◽

Panagiota A. Sotiropoulou ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cell Fate ◽

Lineage Tracing ◽

Premature Aging ◽

Temporary Increase ◽

Proliferative Potential ◽

Cell Dynamics ◽

Asymmetric Divisions

The atypical protein kinase C (aPKC) is a key regulator of polarity and cell fate in lower organisms. However, whether mammalian aPKCs control stem cells and fate in vivo is not known. Here we show that loss of aPKCλ in a self-renewing epithelium, the epidermis, disturbed tissue homeostasis, differentiation, and stem cell dynamics, causing progressive changes in this tissue. This was accompanied by a gradual loss of quiescent hair follicle bulge stem cells and a temporary increase in proliferating progenitors. Lineage tracing analysis showed that loss of aPKCλ altered the fate of lower bulge/hair germ stem cells. This ultimately led to loss of proliferative potential, stem cell exhaustion, alopecia, and premature aging. Inactivation of aPKCλ produced more asymmetric divisions in different compartments, including the bulge. Thus, aPKCλ is crucial for homeostasis of self-renewing stratifying epithelia, and for the regulation of cell fate, differentiation, and maintenance of epidermal bulge stem cells likely through its role in balancing symmetric and asymmetric division.

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Continual inactivation of genes involved in stem cell functional identity stabilizes progenitor commitment

10.1101/2020.02.03.931972 ◽

2020 ◽

Author(s):

Noemi Rives-Quinto ◽

Hideyuki Komori ◽

Derek H. Janssens ◽

Shu Kondo ◽

Qi Dai ◽

...

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Histone Deacetylation ◽

Type Ii ◽

Functional Identity ◽

Larval Brain ◽

Intermediate Progenitors ◽

Differentiated Cells ◽

Mechanistic Investigation

SummaryExpansion of the pool of stem cells that indirectly generate differentiated cells through intermediate progenitors drives vertebrate brain evolution. Due to a lack of lineage information, mechanistic investigation of the competency of stem cells to generate intermediate progenitors remains impossible. Fly larval brain neuroblasts provide excellent in vivo models for investigating the regulation of stem cell functionality during neurogenesis. Type II neuroblasts undergo indirect neurogenesis by dividing asymmetrically to generate a neuroblast and a progeny that commits to an intermediate progenitor (INP) identity. We identified Tailless (Tll) as the master regulator that maintains type II neuroblast functional identity, including the competency to generate INPs. Successive inactivation during INP commitment inhibits tll activation by Notch, preventing INPs from reacquiring neuroblast functionality. We propose that the continual inactivation of neural stem cell functional identity genes by histone deacetylation allows intermediate progenitors to stably commit to generating diverse differentiated cells during indirect neurogenesis.

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