scholarly journals Quantitative assessment of Fgfr1 expression in neurons and glia

2016 ◽  
Author(s):  
Lisha Choubey ◽  
Jantzen C Collette ◽  
Karen Muller Smith
Keyword(s):  
Glial Cells ◽  
Synapse Formation ◽  
Fluorescent Protein ◽  
Neuropsychiatric Disorders ◽  
Cell Types ◽  
Lipid Binding ◽  
Bergmann Glia ◽  
Radial Glial Cells ◽  
Postnatal Maturation ◽  
Green Fluorescent

Fibroblast growth factors (FGFs) and their receptors (FGFRs) have numerous functions in the developing and adult CNS. For example, the FGFR1 receptor is important for proliferation of radial glial cells in the cortex and hippocampus, oligodendrocyte proliferation and regeneration, midline glia morphology and soma translocation, Bergmann glia morphology, and cerebellar morphogenesis. In addition, FGFR1 signaling in astrocytes is required for postnatal maturation of interneurons expressing parvalbumin (PV). FGFR1 is implicated in synapse formation in the hippocampus, and alterations in the expression of Fgfr1 and its ligand, Fgf2 accompany major depression. Understanding which cell types express Fgfr1 during development may elucidate its roles in normal development of the brain as well as illuminate possible causes of certain neuropsychiatric disorders. Here, we used a BAC transgenic reporter line to trace Fgfr1 expression in the developing murine CNS. The specific transgenic line employed was created by the GENSAT project, tgFGFR1-EGFPGP338Gsat, and includes a gene encoding enhanced green fluorescent protein (EGFP) under the regulation of the Fgfr1 promoter, to trace Fgfr1 expression in the developing CNS. This model reveals that Fgfr1 is primarily expressed in glial cells, in both astrocytes and oligodendrocytes, along with some neurons. Dual labeling experiments indicate that the proportion of GFP+ (Fgfr1+) cells that are also GFAP+ increases from postnatal day 7 (P7) to 1 month, illuminating dynamic changes in Fgfr1 expression during postnatal development of the cortex. In postnatal neurogenic areas, GFP expression was also observed in SOX2, doublecortin (DCX), and brain lipid-binding protein (BLBP) expressing cells. Fgfr1 is also highly expressed in DCX positive cells of the dentate gyrus, but not in the rostral migratory stream. Fgfr1 driven GFP was also observed in tanycytes and GFAP+ cells of the hypothalamus, as well as in Bergmann glia and astrocytes of the cerebellum. Understanding which cell types express Fgfr1 may elucidate its role in neuropsychiatric disorders and brain development.

scholarly journals Quantitative assessment of Fgfr1 expression in neurons and glia

2016 ◽  
Author(s):  
Lisha Choubey ◽  
Jantzen C Collette ◽  
Karen Muller Smith
Keyword(s):  
Glial Cells ◽  
Synapse Formation ◽  
Fluorescent Protein ◽  
Neuropsychiatric Disorders ◽  
Cell Types ◽  
Lipid Binding ◽  
Bergmann Glia ◽  
Radial Glial Cells ◽  
Postnatal Maturation ◽  
Green Fluorescent

Fibroblast growth factors (FGFs) and their receptors (FGFRs) have numerous functions in the developing and adult CNS. For example, the FGFR1 receptor is important for proliferation of radial glial cells in the cortex and hippocampus, oligodendrocyte proliferation and regeneration, midline glia morphology and soma translocation, Bergmann glia morphology, and cerebellar morphogenesis. In addition, FGFR1 signaling in astrocytes is required for postnatal maturation of interneurons expressing parvalbumin (PV). FGFR1 is implicated in synapse formation in the hippocampus, and alterations in the expression of Fgfr1 and its ligand, Fgf2 accompany major depression. Understanding which cell types express Fgfr1 during development may elucidate its roles in normal development of the brain as well as illuminate possible causes of certain neuropsychiatric disorders. Here, we used a BAC transgenic reporter line to trace Fgfr1 expression in the developing murine CNS. The specific transgenic line employed was created by the GENSAT project, tgFGFR1-EGFPGP338Gsat, and includes a gene encoding enhanced green fluorescent protein (EGFP) under the regulation of the Fgfr1 promoter, to trace Fgfr1 expression in the developing CNS. This model reveals that Fgfr1 is primarily expressed in glial cells, in both astrocytes and oligodendrocytes, along with some neurons. Dual labeling experiments indicate that the proportion of GFP+ (Fgfr1+) cells that are also GFAP+ increases from postnatal day 7 (P7) to 1 month, illuminating dynamic changes in Fgfr1 expression during postnatal development of the cortex. In postnatal neurogenic areas, GFP expression was also observed in SOX2, doublecortin (DCX), and brain lipid-binding protein (BLBP) expressing cells. Fgfr1 is also highly expressed in DCX positive cells of the dentate gyrus, but not in the rostral migratory stream. Fgfr1 driven GFP was also observed in tanycytes and GFAP+ cells of the hypothalamus, as well as in Bergmann glia and astrocytes of the cerebellum. Understanding which cell types express Fgfr1 may elucidate its role in neuropsychiatric disorders and brain development.

scholarly journals Quantitative assessment of fibroblast growth factor receptor 1 expression in neurons and glia

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3173 ◽  
Author(s):  
Lisha Choubey ◽  
Jantzen C. Collette ◽  
Karen Müller Smith
Keyword(s):  
Glial Cells ◽  
Synapse Formation ◽  
Fluorescent Protein ◽  
Growth Factor Receptor ◽  
Neuropsychiatric Disorders ◽  
Cell Types ◽  
Lipid Binding ◽  
Bergmann Glia ◽  
Fibroblast Growth ◽  
Postnatal Maturation

BackgroundFibroblast growth factors (FGFs) and their receptors (FGFRs) have numerous functions in the developing and adult central nervous system (CNS). For example, the FGFR1 receptor is important for proliferation and fate specification of radial glial cells in the cortex and hippocampus, oligodendrocyte proliferation and regeneration, midline glia morphology and soma translocation, Bergmann glia morphology, and cerebellar morphogenesis. In addition, FGFR1 signaling in astrocytes is required for postnatal maturation of interneurons expressing parvalbumin (PV). FGFR1 is implicated in synapse formation in the hippocampus, and alterations in the expression ofFgfr1and its ligand,Fgf2accompany major depression. Understanding which cell types expressFgfr1during development may elucidate its roles in normal development of the brain as well as illuminate possible causes of certain neuropsychiatric disorders.MethodsHere, we used a BAC transgenic reporter line to traceFgfr1expression in the developing postnatal murine CNS. The specific transgenic line employed was created by the GENSAT project,tgFGFR1-EGFPGP338Gsat, and includes a gene encoding enhanced green fluorescent protein (EGFP) under the regulation of theFgfr1promoter, to traceFgfr1expression in the developing CNS. Unbiased stereological counts were performed for several cell types in the cortex and hippocampus.ResultsThis model reveals thatFgfr1is primarily expressed in glial cells, in both astrocytes and oligodendrocytes, along with some neurons. Dual labeling experiments indicate that the proportion of GFP+ (Fgfr1+) cells that are also GFAP+ increases from postnatal day 7 (P7) to 1 month, illuminating dynamic changes inFgfr1expression during postnatal development of the cortex. In postnatal neurogenic areas, GFP expression was also observed in SOX2, doublecortin (DCX), and brain lipid-binding protein (BLBP) expressing cells.Fgfr1is also highly expressed in DCX positive cells of the dentate gyrus (DG), but not in the rostral migratory stream.Fgfr1driven GFP was also observed in tanycytes and GFAP+ cells of the hypothalamus, as well as in Bergmann glia and astrocytes of the cerebellum.ConclusionsThetgFGFR1-EGFPGP338Gsatmouse model expresses GFP that is congruent with known functions of FGFR1, including hippocampal development, glial cell development, and stem cell proliferation. Understanding which cell types expressFgfr1may elucidate its role in neuropsychiatric disorders and brain development.

scholarly journals Relaxed Repression of Herpes Simplex Virus Type 1 Genomes in Murine Trigeminal Neurons

2007 ◽  
Vol 81 (22) ◽  
pp. 12394-12405 ◽  
Author(s):  
Tracy Terry-Allison ◽  
Colton A. Smith ◽  
Neal A. DeLuca
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Fluorescent Protein ◽  
Cultured Cells ◽  
Trichostatin A ◽  
Cell Types ◽  
Immediate Early ◽  
Green Fluorescent ◽  
Simplex Virus

ABSTRACT The expression of herpes simplex virus (HSV) genomes in the absence of viral regulatory proteins in sensory neurons is poorly understood. Previously, our group reported an HSV immediate early (IE) mutant (d109) unable to express any of the five IE genes and encoding a model human cytomegalovirus immediate early promoter-green fluorescent protein (GFP) transgene. In cultured cells, GFP expressed from this mutant was observed in only a subset of infected cells. The subset exhibited cell type dependence, as the fractions of GFP-expressing cells varied widely among the cell types examined. Herein, we characterize this mutant in murine embryonic trigeminal ganglion (TG) cultures. We found that d109 was nontoxic to neural cultures and persisted in the cultures throughout their life spans. Unlike with some of the cultured cell lines and strains, expression of the GFP transgene was observed in a surprisingly large subset of neurons. However, very few nonneuronal cells expressed GFP. The abilities of ICP0 and an inhibitor of histone deacetylase, trichostatin A (TSA), to activate GFP expression from nonexpressing cells were also compared. The provision of ICP0 by infection with d105 reactivated quiescent genomes in nearly every cell, whereas reactivation by TSA was much more limited and restricted to the previously nonexpressing neurons. Moreover, we found that d109, which does not express ICP0, consistently reactivated HSV type 1 (KOS) in latently infected adult TG cultures. These results suggest that the state of persisting HSV genomes in some TG neurons may be more dynamic and more easily activated than has been observed with nonneuronal cells.

scholarly journals A PDZ-containing Scaffold Related to the Dystrophin Complex at the Basolateral Membrane of Epithelial Cells

1999 ◽  
Vol 145 (2) ◽  
pp. 391-402 ◽  
Author(s):  
Amy M. Kachinsky ◽  
Stanley C. Froehner ◽  
Sharon L. Milgram
Keyword(s):  
Skeletal Muscle ◽  
Fluorescent Protein ◽  
Protein Complexes ◽  
Basolateral Membrane ◽  
Pdz Domain ◽  
Cell Types ◽  
Protein Domain ◽  
Ph Domain ◽  
Green Fluorescent ◽  
Dystrophin Complex

Membrane scaffolding complexes are key features of many cell types, serving as specialized links between the extracellular matrix and the actin cytoskeleton. An important scaffold in skeletal muscle is the dystrophin-associated protein complex. One of the proteins bound directly to dystrophin is syntrophin, a modular protein comprised entirely of interaction motifs, including PDZ (protein domain named for PSD-95, discs large, ZO-1) and pleckstrin homology (PH) domains. In skeletal muscle, the syntrophin PDZ domain recruits sodium channels and signaling molecules, such as neuronal nitric oxide synthase, to the dystrophin complex. In epithelia, we identified a variation of the dystrophin complex, in which syntrophin, and the dystrophin homologues, utrophin and dystrobrevin, are restricted to the basolateral membrane. We used exogenously expressed green fluorescent protein (GFP)-tagged fusion proteins to determine which domains of syntrophin are responsible for its polarized localization. GFP-tagged full-length syntrophin targeted to the basolateral membrane, but individual domains remained in the cytoplasm. In contrast, the second PH domain tandemly linked to a highly conserved, COOH-terminal region was sufficient for basolateral membrane targeting and association with utrophin. The results suggest an interaction between syntrophin and utrophin that leaves the PDZ domain of syntrophin available to recruit additional proteins to the epithelial basolateral membrane. The assembly of multiprotein signaling complexes at sites of membrane specialization may be a widespread function of dystrophin-related protein complexes.

scholarly journals Nucleocytoplasmic transport in human astrocytes: decreased nuclear uptake of the HIV Rev shuttle protein

2001 ◽  
Vol 114 (9) ◽  
pp. 1717-1729
Author(s):  
M. Neumann ◽  
E. Afonina ◽  
F. Ceccherini-Silberstein ◽  
S. Schlicht ◽  
V. Erfle ◽  
...  
Keyword(s):  
Fluorescent Protein ◽  
Virus Production ◽  
Time Lapse ◽  
Cell Types ◽  
Nucleocytoplasmic Transport ◽  
Nucleocytoplasmic Trafficking ◽  
Nuclear Uptake ◽  
Strong Shift ◽  
Human Astrocytes ◽  
Green Fluorescent

Astrocytes are cellular targets for the human immunodeficiency virus (HIV) that limit virus production, owing, at least in part, to the diminished functionality of the viral post-transcriptional stimulatory factor Rev. To understand the trafficking process in astrocytes, we compared nucleocytoplasmic transport of Rev and various proteins with well-characterized nucleocytoplasmic transport features in human astrocytes and control cells (HeLa). Localization and trafficking characteristics of several cellular and viral proteins, as well as nuclear trafficking of classical peptide signals upon microinjection were similar in both cell types, indicating maintenance of general features of nucleocytoplasmic transport in astrocytes. Quantification of fluorescence in living cells expressing Rev fused to green fluorescent protein (GFP) indicated a strong shift in intracellular distribution of Rev in astrocytes, with 50–70% of Rev in the cytoplasm, whereas the cytoplasmic proportion of Rev in HeLa cells is around 10%. The dynamics of nucleocytoplasmic trafficking of Rev were compared in astrocytes and Rev-permissive cells by monitoring migration of Rev-GFP in cell fusions using highly sensitive time-lapse imaging. Nuclear uptake of Rev was dramatically retarded in homo-polykaryons of astrocytes compared with control cells. Diminished nuclear uptake of Rev was also observed in hetero-polykaryons of Rev-permissive cells and astrocytes. These results indicate that astrocytes contain a cytoplasmic activity that interferes with nuclear uptake of Rev. Our studies suggest a model in which Rev is prevented from functioning efficiently in astrocytes by specific alterations of its nucleocytoplasmic trafficking properties. http://www.biologists.com/JCS/movies/jcs1709.html

Partial Reprogramming As An Emerging Strategy for Safe Induced Cell Generation and Rejuvenation

2019 ◽  
Vol 19 (4) ◽  
pp. 248-254
Author(s):  
Marianne Lehmann ◽  
Martina Canatelli-Mallat ◽  
Priscila Chiavellini ◽  
Gloria M. Cónsole ◽  
Maria D. Gallardo ◽  
...  
Keyword(s):  
Stem Cells ◽  
Somatic Cell ◽  
Fluorescent Protein ◽  
Somatic Cells ◽  
Cell Types ◽  
Cell Reprogramming ◽  
Green Fluorescent ◽  
Original Cell

Background: Conventional cell reprogramming involves converting a somatic cell line into induced pluripotent stem cells (iPSC), which subsequently can be re-differentiated to specific somatic cell types. Alternatively, partial cell reprogramming converts somatic cells into other somatic cell types by transient expression of pluripotency genes thus generating intermediates that retain their original cell identity, but are responsive to appropriate cocktails of specific differentiation factors. Additionally, biological rejuvenation by partial cell reprogramming is an emerging avenue of research. Objective: Here, we will briefly review the emerging information pointing to partial reprogramming as a suitable strategy to achieve cell reprogramming and rejuvenation, bypassing cell dedifferentiation. Methods: In this context, regulatable pluripotency gene expression systems are the most widely used at present to implement partial cell reprogramming. For instance, we have constructed a regulatable bidirectional adenovector expressing Green Fluorescent Protein and oct4, sox2, klf4 and c-myc genes (known as the Yamanaka genes or OSKM). Results: Partial cell reprogramming has been used to reprogram fibroblasts to cardiomyocytes, neural progenitors and neural stem cells. Rejuvenation by cyclic partial reprogramming has been achieved both in vivo and in cell culture using transgenic mice and cells expressing the OSKM genes, respectively, controlled by a regulatable promoter. Conclusion: Partial reprogramming emerges as a powerful tool for the genesis of iPSC-free induced somatic cells of therapeutic value and for the implementation of in vitro and in vivo rejuvenation keeping cell type identity unchanged.

Cell-sorting in aggregates of Dictyostelium discoideum

1999 ◽  
Vol 112 (22) ◽  
pp. 3923-3929 ◽  
Author(s):  
A. Nicol ◽  
W. Rappel ◽  
H. Levine ◽  
W.F. Loomis
Keyword(s):  
Cell Sorting ◽  
Fluorescent Protein ◽  
Cell Mass ◽  
Three Dimensional ◽  
Cell Movement ◽  
Time Lapse ◽  
Cell Types ◽  
Cell Type ◽  
Prespore Cell ◽  
Green Fluorescent

When Dictyostelium cells are induced to develop between a coverslip and a layer of agarose, they aggregate normally into groups containing up to a thousand cells but are then constrained to form disks only a few cells thick that appear to be equivalent to the three-dimensional mounds formed on top of agarose. Such vertically restricted aggregates frequently develop into elongated motile structures, the flattened equivalent of three-dimensional slugs. The advantage of using this system is that the restricted z-dimension enables direct microscopic visualization of most of the cells in the developing structure. We have used time lapse digital fluorescence microscopy of Dictyostelium strains expressing green fluorescent protein (GFP) under the control of either prestalk or prespore specific promoters to follow cell sorting in these flattened mounds. We find that prestalk and prespore cells expressing GFP arise randomly in early aggregates and then rotate rapidly around the disk mixed with the other cell type. After a few hours, the cell types sort out by a process which involves striking changes in relative cell movement. Once sorted, the cell types move independently of each other showing very little heterotypic adhesion. When a group of prestalk cells reaches the edge of the disk, it moves out and is followed by the prespore cell mass. We suggest that sorting may result from cell type specific changes in adhesion and the consequent disruption of movement in the files of cells that are held together by end-to-end adhesion.

Transcriptional activation by the human Hsp70-associating protein Hap50

2001 ◽  
Vol 114 (10) ◽  
pp. 1839-1845
Author(s):  
Y. Niyaz ◽  
M. Zeiner ◽  
U. Gehring
Keyword(s):  
Transcriptional Activation ◽  
Fluorescent Protein ◽  
Biological Effects ◽  
Cell Types ◽  
Mrna Stabilization ◽  
Activation Of Transcription ◽  
Enhanced Expression ◽  
Endogenous Genes ◽  
Green Fluorescent ◽  
Different Cell Types

We investigated human Hap50, the large isoform of the previously characterized Hsp70/Hsc70-associating protein Hap46, also called BAG-1, for effects on transcriptional activities. Overproduction by transient transfection led to enhanced expression of reporter gene constructs in various cell types using different promoters, suggesting independence of promoter type. Similarly, overexpression of Hap50 resulted in increased levels of poly(A)(+)mRNAs in HeLa, COS-7, 3T3 and HTC cells. Concomitantly, the expression of some selected endogenous genes, such as those coding for c-Jun and the glucocorticoid receptor, was enhanced significantly relative to actin. Nuclear runoff transcription assays using HeLa cells showed that the effect is caused by increased transcription rates rather than mRNA stabilization. Activation of transcription by Hap50 occurred at 37 degrees C and did not require prior thermal stress, as is the case for Hap46. In accordance with these biological effects, Hap50 is localized exclusively in the nuclear compartment of different cell types, whereas Hap46 is mostly cytoplasmic in unstressed cells, as revealed by use of fusion constructs with green fluorescent protein. High cellular levels of Hap50 were found to make cells less susceptible to adverse environmental effects such as heat stress. Our data suggest that Hap50 is a nuclear protein that acts in cells to increase the transcription of various genes.

scholarly journals Regulation of Apoptosis by the Ft1 Protein, a New Modulator of Protein Kinase B/Akt

2004 ◽  
Vol 24 (4) ◽  
pp. 1493-1504 ◽  
Author(s):  
Ingrid Remy ◽  
Stephen W. Michnick
Keyword(s):  
Protein Kinase ◽  
Fas Ligand ◽  
Fluorescent Protein ◽  
Protein Kinase B ◽  
Protein A ◽  
Cell Types ◽  
Wide Distribution ◽  
Upstream Kinase ◽  
Protein Fragment Complementation Assay ◽  
Green Fluorescent

ABSTRACT The serine/threonine kinase protein kinase B (PKB)/Akt plays a central role in many cellular processes, including cell growth, glucose metabolism, and apoptosis. However, the identification and validation of novel regulators or effectors is key to future advances in understanding the multiple functions of PKB. Here we report the identification of a novel PKB binding protein, called Ft1, from a cDNA library screen using a green fluorescent protein-based protein-fragment complementation assay. We show that the Ft1 protein interacts directly with PKB, enhancing the phosphorylation of both of its regulatory sites by promoting its interaction with the upstream kinase PDK1. Further, the modulation of PKB activity by Ft1 has a strong effect on the apoptosis susceptibility of T lymphocytes treated with glucocorticoids. We demonstrate that this phenomenon occurs via a PDK1/PKB/GSK3/NF-ATc signaling cascade that controls the production of the proapoptotic hormone Fas ligand. The wide distribution of Ft1 in adult tissues suggests that it could be a general regulator of PKB activity in the control of differentiation, proliferation, and apoptosis in many cell types.

scholarly journals A multipotent precursor in the thymus maps to the branching point of the T versus B lineage decision

2005 ◽  
Vol 202 (1) ◽  
pp. 21-31 ◽  
Author(s):  
Claudia Benz ◽  
Conrad C. Bleul
Keyword(s):  
Dendritic Cells ◽  
Fluorescent Protein ◽  
Single Cells ◽  
Cell Types ◽  
Differentiation Potential ◽  
Lineage Decision ◽  
Branching Point ◽  
B Lineage ◽  
Green Fluorescent

Hematopoietic precursors continuously colonize the thymus where they give rise mainly to T cells, but also to B and dendritic cells. The lineage relationship between these three cell types is unclear, and it remains to be determined if precursors in the thymus are multipotent, oligopotent, or lineage restricted. Resolution of this question necessitates the determination of the clonal differentiation potential of the most immature precursors in the thymus. Using a CC chemokine receptor 9–enhanced green fluorescent protein knock-in allele like a surface marker of unknown function, we identify a multipotent precursor present in bone marrow, blood, and thymus. Single cells of this precursor give rise to T, B, and dendritic cells. A more differentiated stage of this multipotent precursor in the thymus has lost the capacity to generate B but not T, dendritic, and myeloid cells. Thus, the newly identified precursor maps to the branching point of the T versus B lineage decision in the hematopoietic lineage hierarchy.

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