scholarly journals Epidermal development requires ninein for spindle orientation and cortical microtubule organization

2019 ◽  
Vol 2 (2) ◽  
pp. e201900373 ◽  
Author(s):  
Nicolas Lecland ◽  
Chiung-Yueh Hsu ◽  
Cécile Chemin ◽  
Andreas Merdes ◽  
Christiane Bierkamp
Keyword(s):  
Progenitor Cells ◽  
Cortical Microtubule ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Microtubule Organization ◽  
Differentiation Markers ◽  
Epidermal Barrier ◽  
Skin Development ◽  
Protein Levels ◽  
Dependent Transport

In mammalian skin, ninein localizes to the centrosomes of progenitor cells and relocates to the cell cortex upon differentiation of keratinocytes, where cortical arrays of microtubules are formed. To examine the function of ninein in skin development, we use epidermis-specific and constitutive ninein-knockout mice to demonstrate that ninein is necessary for maintaining regular protein levels of the differentiation markers filaggrin and involucrin, for the formation of desmosomes, for the secretion of lamellar bodies, and for the formation of the epidermal barrier. Ninein-deficient mice are viable but develop a thinner skin with partly impaired epidermal barrier. We propose two underlying mechanisms: first, ninein contributes to spindle orientation during the division of progenitor cells, whereas its absence leads to misoriented cell divisions, altering the pool of progenitor cells. Second, ninein is required for the cortical organization of microtubules in differentiating keratinocytes, and for the cortical re-localization of microtubule-organizing proteins, and may thus affect any mechanisms that depend on localized microtubule-dependent transport.

scholarly journals Lis1 is essential for cortical microtubule organization and desmosome stability in the epidermis

2011 ◽  
Vol 194 (4) ◽  
pp. 631-642 ◽  
Author(s):  
Kaelyn D. Sumigray ◽  
Hsin Chen ◽  
Terry Lechler
Keyword(s):  
Cortical Microtubule ◽  
Cell Types ◽  
Epidermal Differentiation ◽  
Cell Cortex ◽  
Microtubule Organization ◽  
Specific Subset ◽  
Keratin Filaments ◽  
Perinatal Lethality ◽  
Cytoskeletal Networks ◽  
Centrosomal Proteins

Desmosomes are cell–cell adhesion structures that integrate cytoskeletal networks. In addition to binding intermediate filaments, the desmosomal protein desmoplakin (DP) regulates microtubule reorganization in the epidermis. In this paper, we identify a specific subset of centrosomal proteins that are recruited to the cell cortex by DP upon epidermal differentiation. These include Lis1 and Ndel1, which are centrosomal proteins that regulate microtubule organization and anchoring in other cell types. This recruitment was mediated by a region of DP specific to a single isoform, DPI. Furthermore, we demonstrate that the epidermal-specific loss of Lis1 results in dramatic defects in microtubule reorganization. Lis1 ablation also causes desmosomal defects, characterized by decreased levels of desmosomal components, decreased attachment of keratin filaments, and increased turnover of desmosomal proteins at the cell cortex. This contributes to loss of epidermal barrier activity, resulting in completely penetrant perinatal lethality. This work reveals essential desmosome-associated components that control cortical microtubule organization and unexpected roles for centrosomal proteins in epidermal function.

scholarly journals Control of Mitotic Spindle Position by the Saccharomyces cerevisiae Formin Bni1p

1999 ◽  
Vol 144 (5) ◽  
pp. 947-961 ◽  
Author(s):  
Laifong Lee ◽  
Saskia K. Klee ◽  
Marie Evangelista ◽  
Charles Boone ◽  
David Pellman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Spindle ◽  
Cortical Microtubule ◽  
Spindle Pole Body ◽  
Spindle Pole ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Cortical Actin ◽  
Bud Site ◽  
Spindle Position

Alignment of the mitotic spindle with the axis of cell division is an essential process in Saccharomyces cerevisiae that is mediated by interactions between cytoplasmic microtubules and the cell cortex. We found that a cortical protein, the yeast formin Bni1p, was required for spindle orientation. Two striking abnormalities were observed in bni1Δ cells. First, the initial movement of the spindle pole body (SPB) toward the emerging bud was defective. This phenotype is similar to that previously observed in cells lacking the kinesin Kip3p and, in fact, BNI1 and KIP3 were found to be in the same genetic pathway. Second, abnormal pulling interactions between microtubules and the cortex appeared to cause preanaphase spindles in bni1Δ cells to transit back and forth between the mother and the bud. We therefore propose that Bni1p may localize or alter the function of cortical microtubule-binding sites in the bud. Additionally, we present evidence that other bipolar bud site determinants together with cortical actin are also required for spindle orientation.

scholarly journals Sas4 links basal bodies to cell division via Hippo signaling

2020 ◽  
Vol 219 (8) ◽  
Author(s):  
Marisa D. Ruehle ◽  
Alexander J. Stemm-Wolf ◽  
Chad G. Pearson
Keyword(s):  
Cell Division ◽  
Signaling Pathway ◽  
Cortical Microtubule ◽  
Cell Cortex ◽  
Basal Bodies ◽  
Hippo Signaling ◽  
Microtubule Organization ◽  
Hippo Signaling Pathway ◽  
Daughter Cells ◽  
Striated Fiber

Basal bodies (BBs) are macromolecular complexes required for the formation and cortical positioning of cilia. Both BB assembly and DNA replication are tightly coordinated with the cell cycle to ensure their accurate segregation and propagation to daughter cells, but the mechanisms ensuring coordination are unclear. The Tetrahymena Sas4/CPAP protein is enriched at assembling BBs, localizing to the core BB structure and to the base of BB-appendage microtubules and striated fiber. Sas4 is necessary for BB assembly and cortical microtubule organization, and Sas4 loss disrupts cell division furrow positioning and DNA segregation. The Hippo signaling pathway is known to regulate cell division furrow position, and Hippo molecules localize to BBs and BB-appendages. We find that Sas4 loss disrupts localization of the Hippo activator, Mob1, suggesting that Sas4 mediates Hippo activity by promoting scaffolds for Mob1 localization to the cell cortex. Thus, Sas4 links BBs with an ancient signaling pathway known to promote the accurate and symmetric segregation of the genome.

scholarly journals Linking cortical microtubule attachment and exocytosis

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 469 ◽  
Author(s):  
Ivar Noordstra ◽  
Anna Akhmanova
Keyword(s):  
Targeted Delivery ◽  
Mammalian Cells ◽  
Spatial Organization ◽  
Protein Complexes ◽  
Cortical Microtubule ◽  
Actin Binding ◽  
Cell Cortex ◽  
Microtubule Organization ◽  
Vesicle Docking ◽  
Microtubule Attachment

Exocytosis is a fundamental cellular process whereby secreted molecules are packaged into vesicles that move along cytoskeletal filaments and fuse with the plasma membrane. To function optimally, cells are strongly dependent on precisely controlled delivery of exocytotic cargo. In mammalian cells, microtubules serve as major tracks for vesicle transport by motor proteins, and thus microtubule organization is important for targeted delivery of secretory carriers. Over the years, multiple microtubule-associated and cortical proteins have been discovered that facilitate the interaction between the microtubule plus ends and the cell cortex. In this review, we focus on mammalian protein complexes that have been shown to participate in both cortical microtubule capture and exocytosis, thereby regulating the spatial organization of secretion. These complexes include microtubule plus-end tracking proteins, scaffolding factors, actin-binding proteins, and components of vesicle docking machinery, which together allow efficient coordination of cargo transport and release.

Abstract 1839: MicroRNA-499 Promotes the Differentiation of Cardiac Progenitor Cells into Myocytes

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Toru Hosoda ◽  
Konrad Urbanek ◽  
Adriana Bastos Carvalho ◽  
Claudia Bearzi ◽  
Silvana Bardelli ◽  
...  
Keyword(s):  
Gap Junctions ◽  
Progenitor Cells ◽  
Target Genes ◽  
Brdu Incorporation ◽  
Cardiac Progenitor Cells ◽  
Partial Recovery ◽  
Rt Pcr ◽  
Protein Levels ◽  
Cell Proliferation And Differentiation ◽  
Reporter Assays

Myocardial regeneration mediated by cardiac progenitor cells (CPCs) results in the partial recovery of the infarcted heart but the newly formed myocytes within the necrotic tissue have fetal-neonatal characteristics. In contrast, CPC activation in the remote viable myocardium results in the formation of mature myocytes, suggesting that CPC differentiation is conditioned by the surrounding cells. Thus, the hypothesis is raised that microRNAs (miRs) that are highly expressed in myocytes and are absent in CPCs, may translocate through gap junctions to adjacent CPCs promoting their differentiation. By employing miR array and Q-RT-PCR, miR-499 was found to be ~500-fold more expressed in myocytes than CPCs. Additionally, we demonstrated that miR-499 translocates from neighboring cells to CPCs through the formation of gap junctions. The translocated miR-499 was functional and repressed the expression of target genes. Among 200 putative targets of miR-499, we have elected to study Sox6 and Rod1. The validation of these putative miR-499-targets was obtained by reporter assays; cells transfected with miR-499 together with plasmids carrying luciferase and the 3′-UTR region of Sox6 or Rod1 show the expected decrease in luciferase activity. Transcripts of Sox6 and Rod1 were measured by Q-RT-PCR in myocytes and CPCs; Sox6 mRNA was 2-fold higher and Rod1 mRNA was 98% lower in myocytes than CPCs. However, the protein levels of Sox6 and Rod1 were significantly lower in myocytes than CPCs suggesting that miR-499 promotes degradation and/or inhibition of translation of these target genes. To document miR-499 function, CPCs were transfected with a miR-499-expression vector and cell proliferation and differentiation were evaluated 3 days later. BrdU incorporation decreased 60% and the cells displayed a marked upregulation of the myocyte-specific transcription factors Nkx2.5 and MEF2C. Similar results were obtained when Sox6 and Rod1 were selectively blocked with siRNA. In both cases, the number of Nkx2.5- and MEF2C-positive cells increased 2–3-fold. Thus, our data indicate that miR-499 translocates via gap junction from myocytes to CPCs where miR-499 is a crucial modulator of the differentiation of CPCs into cardiomyocytes through the repression of Sox6 and Rod1.

scholarly journals Specific Responses in Rat Small Intestinal Epithelial mRNA Expression and Protein Levels During Chemotherapeutic Damage and Regeneration

2002 ◽  
Vol 50 (11) ◽  
pp. 1525-1536 ◽  
Author(s):  
Melissa Verburg ◽  
Ingrid B. Renes ◽  
Danielle J.P.M. Van Nispen ◽  
Sacha Ferdinandusse ◽  
Marieke Jorritsma ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Goblet Cell ◽  
Expression Patterns ◽  
Paneth Cell ◽  
Cytostatic Drug ◽  
Differentiation Markers ◽  
Protein Levels ◽  
Cell Functions ◽  
Enhanced Expression ◽  
Small Intestinal

The rapidly dividing small intestinal epithelium is very sensitive to the cytostatic drug methotrexate. We investigated the regulation of epithelial gene expression in rat jejunum during methotrexate-induced damage and regeneration. Ten differentiation markers were localized on tissue sections and quantified at mRNA and protein levels relative to control levels. We analyzed correlations in temporal expression patterns between markers. mRNA expression of enterocyte and goblet cell markers decreased significantly during damage for a specific period. Of these, sucrase-isomaltase (-62%) and CPS (-82%) were correlated. Correlations were also found between lactase (−76%) and SGLT1 (−77%) and between I-FABP (−52%) and L-FABP (-45%). Decreases in GLUT5 (−53%), MUC2 (-43%), and TFF3 (−54%) mRNAs occurred independently of any of the other markers. In contrast, lysozyme mRNA present in Paneth cells increased (+76%). At the protein level, qualitative and quantitative changes were in agreement with mRNA expression, except for Muc2 (+115%) and TFF3 (+81%), which increased significantly during damage, following independent patterns. During regeneration, expression of each marker returned to control levels. The enhanced expression of cytoprotective molecules (Muc2, TFF3, lysozyme) during damage represents maintenance of goblet cell and Paneth cell functions, most likely to protect the epithelium. Decreased expression of enterocyte-specific markers represents decreased enterocyte function, of which fatty acid transporters were least affected.

scholarly journals Sequential generations of hematopoietic colonies derived from single nonlineage-committed CD34+CD38- progenitor cells

Blood ◽  
1991 ◽  
Vol 77 (6) ◽  
pp. 1218-1227 ◽  
Author(s):  
LW Terstappen ◽  
S Huang ◽  
M Safford ◽  
PM Lansdorp ◽  
MR Loken
Keyword(s):  
Progenitor Cells ◽  
Lineage Commitment ◽  
Multiparameter Flow Cytometry ◽  
Differentiation Markers ◽  
Single Class ◽  
Hematopoietic Stem ◽  
Cd34 Antigen ◽  
Cd34 Cells ◽  
Cell Fractions ◽  
Cd38 Antigen

Abstract Multiparameter flow cytometry was applied on normal human bone marrow (BM) cells to study the lineage commitment of progenitor cells ie, CD34+ cells. Lineage commitment of the CD34+ cells into the erythroid lineage was assessed by the coexpression of high levels of the CD71 antigen, the myeloid lineage by coexpression of the CD33 antigen and the B-lymphoid lineage by the CD10 antigen. Three color immunofluorescence experiments showed that all CD34+ BM cells that expressed the CD71, CD33, and CD10 antigens, concurrently stained brightly with anti-CD38 monoclonal antibodies (MoAbs). In addition, the CD38 antigen was brightly expressed on early T lymphocytes in human thymus, characterized by CD34, CD5, and CD7 expression. Only 1% of the CD34+ cells, 0.01% of nucleated cells in normal BM, did not express the CD38 antigen. The CD34+, CD38- cell population lacked differentiation markers and were homogeneous primitive blast cells by morphology. In contrast the CD34+, CD38 bright cell populations were heterogeneous in morphology and contained myeloblasts and erythroblasts, as well as lymphoblasts. These features are in agreement with properties expected from putative pluripotent hematopoietic stem cells; indeed, the CD34 antigen density decreased concurrently with increasing CD38 antigen density suggesting an upregulation of the CD38 antigen on differentiation of the CD34+ cells. Further evidence for a strong enrichment of early hematopoietic precursors in the CD34+, CD38- cell fraction was obtained from culture experiments in which CD34+ cell fractions with increasing density of the CD38 antigen were sorted singularly and assayed for blast colony formation. On day 14 of incubation, interleukin-3 (IL-3), IL-6, and GM-CSF, G-CSF, and erythropoietin (Epo) were added in each well. Twenty-five percent of the single sorted cells that expressed CD34 but lacked CD38 antigen gave rise to primitive colonies 28 to 34 days after cell sorting. The ability to form primitive colonies decreased rapidly with increasing density of the CD38 antigen. During 120 days of culture, up to five sequential generations of colonies were obtained after replating of the first-generation primitive colonies. This study provides direct evidence for the existence of a single class of progenitors with extensive proliferative capacity in human BM and provides an experimental approach for their purification, manipulation, and further characterization.

scholarly journals LGN regulates mitotic spindle orientation during epithelial morphogenesis

2010 ◽  
Vol 189 (2) ◽  
pp. 275-288 ◽  
Author(s):  
Zhen Zheng ◽  
Huabin Zhu ◽  
Qingwen Wan ◽  
Jing Liu ◽  
Zhuoni Xiao ◽  
...  
Keyword(s):  
Mitotic Spindle ◽  
Mdck Cells ◽  
Apical Cell ◽  
Cell Polarization ◽  
Epithelial Morphogenesis ◽  
Spindle Orientation ◽  
Cell Cortex ◽  
Dividing Cells ◽  
Cortical Localization ◽  
Lateral Cell

Coordinated cell polarization and mitotic spindle orientation are thought to be important for epithelial morphogenesis. Whether spindle orientation is indeed linked to epithelial morphogenesis and how it is controlled at the molecular level is still unknown. Here, we show that the NuMA- and Gα-binding protein LGN is required for directing spindle orientation during cystogenesis of MDCK cells. LGN localizes to the lateral cell cortex, and is excluded from the apical cell cortex of dividing cells. Depleting LGN, preventing its cortical localization, or disrupting its interaction with endogenous NuMA or Gα proteins all lead to spindle misorientation and abnormal cystogenesis. Moreover, artificial mistargeting of endogenous LGN to the apical membrane results in a near 90° rotation of the spindle axis and profound cystogenesis defects that are dependent on cell division. The normal apical exclusion of LGN during mitosis appears to be mediated by atypical PKC. Thus, cell polarization–mediated spatial restriction of spindle orientation determinants is critical for epithelial morphogenesis.

scholarly journals Human Cytomegalovirus Infection Dysregulates the Localization and Stability of NICD1 and Jag1 in Neural Progenitor Cells

2015 ◽  
Vol 89 (13) ◽  
pp. 6792-6804 ◽  
Author(s):  
Xiao-Jun Li ◽  
Xi-Juan Liu ◽  
Bo Yang ◽  
Ya-Ru Fu ◽  
Fei Zhao ◽  
...  
Keyword(s):  
Stem Cell ◽  
Notch Signaling ◽  
Progenitor Cells ◽  
Human Cytomegalovirus ◽  
Neural Progenitor Cells ◽  
Hcmv Infection ◽  
Fetal Brain ◽  
Neural Progenitor ◽  
Notch Signaling Pathway ◽  
Protein Levels

ABSTRACTHuman cytomegalovirus (HCMV) infection of the developing fetus frequently results in major neural developmental damage. In previous studies, HCMV was shown to downregulate neural progenitor/stem cell (NPC) markers and induce abnormal differentiation. As Notch signaling plays a vital role in the maintenance of stem cell status and is a switch that governs NPC differentiation, the effect of HCMV infection on the Notch signaling pathway in NPCs was investigated. HCMV downregulated mRNA levels of Notch1 and its ligand, Jag1, and reduced protein levels and altered the intracellular localization of Jag1 and the intracellular effector form of Notch1, NICD1. These effects required HCMV gene expression and appeared to be mediated through enhanced proteasomal degradation. Transient expression of the viral tegument proteins of pp71 and UL26 reduced NICD1 and Jag1 protein levels endogenously and exogenously. Given the critical role of Notch signaling in NPC growth and differentiation, these findings reveal important mechanisms by which HCMV disturbs neural cell developmentin vitro. Similar eventsin vivomay be associated with HCMV-mediated neuropathogenesis during congenital infection in the fetal brain.IMPORTANCECongenital human cytomegalovirus (HCMV) infection is the leading cause of birth defects that primarily manifest as neurological disabilities. Neural progenitor cells (NPCs), key players in fetal brain development, are the most susceptible cell type for HCMV infection in the fetal brain. Studies have shown that NPCs are fully permissive for HCMV infection, which causes neural cell loss and premature differentiation, thereby perturbing NPC fate. Elucidation of virus-host interactions that govern NPC proliferation and differentiation is critical to understanding neuropathogenesis. The Notch signaling pathway is critical for maintaining stem cell status and functions as a switch for differentiation of NPCs. Our investigation into the impact of HCMV infection on this pathway revealed that HCMV dysregulates Notch signaling by altering expression of the Notch ligand Jag1, Notch1, and its active effector in NPCs. These results suggest a mechanism for the neuropathogenesis induced by HCMV infection that includes altered NPC differentiation and proliferation.

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