Lineage commitment of embryonic cells involves MEK1-dependent clearance of pluripotency regulator Ventx2

During early embryogenesis, cells must exit pluripotency and commit to multiple lineages in all germ-layers. How this transition is operated in vivo is poorly understood. Here, we report that MEK1 and the Nanog-related transcription factor Ventx2 coordinate this transition. MEK1 was required to make Xenopus pluripotent cells competent to respond to all cell fate inducers tested. Importantly, MEK1 activity was necessary to clear the pluripotency protein Ventx2 at the onset of gastrulation. Thus, concomitant MEK1 and Ventx2 knockdown restored the competence of embryonic cells to differentiate. Strikingly, MEK1 appeared to control the asymmetric inheritance of Ventx2 protein following cell division. Consistently, when Ventx2 lacked a functional PEST-destruction motif, it was stabilized, displayed symmetric distribution during cell division and could efficiently maintain pluripotency gene expression over time. We suggest that asymmetric clearance of pluripotency regulators may represent an important mechanism to ensure the progressive assembly of primitive embryonic tissues.

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Dynamic changes in cell size and corresponding cell fate after optic nerve injury

Scientific Reports ◽

10.1038/s41598-020-77760-1 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Benjamin M. Davis ◽

Li Guo ◽

Nivedita Ravindran ◽

Ehtesham Shamsher ◽

Veerle Baekelandt ◽

...

Keyword(s):

Optic Nerve ◽

Cell Size ◽

Nerve Injury ◽

Cell Fate ◽

Adaptive Optics ◽

Retinal Cell ◽

Multiple Time ◽

Optic Nerve Injury ◽

Over Time

AbstractIdentifying disease-specific patterns of retinal cell loss in pathological conditions has been highlighted by the emergence of techniques such as Detection of Apoptotic Retinal Cells and Adaptive Optics confocal Scanning Laser Ophthalmoscopy which have enabled single-cell visualisation in vivo. Cell size has previously been used to stratify Retinal Ganglion Cell (RGC) populations in histological samples of optic neuropathies, and early work in this field suggested that larger RGCs are more susceptible to early loss than smaller RGCs. More recently, however, it has been proposed that RGC soma and axon size may be dynamic and change in response to injury. To address this unresolved controversy, we applied recent advances in maximising information extraction from RGC populations in retinal whole mounts to evaluate the changes in RGC size distribution over time, using three well-established rodent models of optic nerve injury. In contrast to previous studies based on sampling approaches, we examined the whole Brn3a-positive RGC population at multiple time points over the natural history of these models. The morphology of over 4 million RGCs was thus assessed to glean novel insights from this dataset. RGC subpopulations were found to both increase and decrease in size over time, supporting the notion that RGC cell size is dynamic in response to injury. However, this study presents compelling evidence that smaller RGCs are lost more rapidly than larger RGCs despite the dynamism. Finally, using a bootstrap approach, the data strongly suggests that disease-associated changes in RGC spatial distribution and morphology could have potential as novel diagnostic indicators.

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Unequal distribution of 16S mtrRNA at the 2-cell stage regulates cell lineage allocations in mouse embryos

Reproduction ◽

10.1530/rep-15-0301 ◽

2016 ◽

Vol 151 (4) ◽

pp. 351-367 ◽

Cited By ~ 9

Author(s):

Zhuxia Zheng ◽

Hongmei Li ◽

Qinfen Zhang ◽

Lele Yang ◽

Huayu Qi

Keyword(s):

Cell Fate ◽

Rna Binding ◽

Rna Binding Proteins ◽

Cell Lineage ◽

Early Embryogenesis ◽

Embryonic Cells ◽

Cell Stage ◽

Unequal Distribution ◽

Fate Determinants ◽

Cell Fate Determinants

Cell lineage determination during early embryogenesis has profound effects on adult animal development. Pre-patterning of embryos, such as that of Drosophila and Caenorhabditis elegans, is driven by asymmetrically localized maternal or zygotic factors, including mRNA species and RNA binding proteins. However, it is not clear how mammalian early embryogenesis is regulated and what the early cell fate determinants are. Here we show that, in mouse, mitochondrial ribosomal RNAs (mtrRNAs) are differentially distributed between 2-cell sister blastomeres. This distribution pattern is not related to the overall quantity or activity of mitochondria which appears equal between 2-cell sister blastomeres. Like in lower species, 16S mtrRNA is found to localize in the cytoplasm outside of mitochondria in mouse 2-cell embryos. Alterations of 16S mtrRNA levels in one of the 2-cell sister blastomere via microinjection of either sense or anti-sense RNAs drive its progeny into different cell lineages in blastocyst. These results indicate that mtrRNAs are differentially distributed among embryonic cells at the beginning of embryogenesis in mouse and they are functionally involved in the regulation of cell lineage allocations in blastocyst, suggesting an underlying molecular mechanism that regulates pre-implantation embryogenesis in mouse.

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Reticulocytes II: Reexamination of the in vivo survival of stress reticulocytes

Blood ◽

10.1182/blood.v75.9.1877.1877 ◽

1990 ◽

Vol 75 (9) ◽

pp. 1877-1882 ◽

Cited By ~ 23

Author(s):

NA Noble ◽

QP Xu ◽

LL Hoge

Keyword(s):

Cell Division ◽

Terminal Cell ◽

Intrinsic Properties ◽

Over Time

Abstract Very young reticulocytes are released into the circulation in response to the stress of anemia. These stress reticulocytes have shortened in vivo survival when transfused into normal recipients, and are generally considered to be abnormal because they have skipped a terminal cell division. We reevaluated one aspect of their abnormality: that of in vivo survival. Using methodology that accounted for all cells transfused, in vivo survival of both normal and stress reticulocytes was investigated in both normal and anemic recipients. The experiments demonstrate that: (1) survival of reticulocytes is normal only when normal reticulocytes are injected into nonanemic animals; (2) intrinsic properties of stress reticulocytes lead to their immediate removal from the circulation by normal recipients to a significantly greater extent than by anemic recipients; and (3) both stress and normal reticulocytes are removed at an accelerated rate over time by anemic recipients. Taken together, the data indicate that in the course of becoming anemic, an adaptation occurs that allows cells produced during anemia to circulate considerably longer in anemic animals than they could in normal nonanemic animals. Other studies disclosed that increased reticulocyte survival in anemic animals could not be attributed to reticuloendothelial overload, but is induced by adaptation of the spleen, decreasing its removal of stress reticulocytes.

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Regulation of E2F1-Dependent Gene Transcription and Apoptosis by the ETS-Related Transcription Factor GABPγ1

Molecular and Cellular Biology ◽

10.1128/mcb.22.7.2147-2158.2002 ◽

2002 ◽

Vol 22 (7) ◽

pp. 2147-2158 ◽

Cited By ~ 18

Author(s):

Ludger Hauck ◽

Rudolf G. Kaba ◽

Martin Lipp ◽

Rainer Dietz ◽

Rüdiger von Harsdorf

Keyword(s):

Transcription Factor ◽

Transcription Factors ◽

Cell Fate ◽

Binding Domain ◽

Pocket Proteins ◽

Related Transcription Factor ◽

Terminal Amino ◽

Fate Decision

ABSTRACT The E2F family of transcription factors comprises six related members which are involved in the control of the coordinated progression through the G1/S-phase transition of cell cycle or in cell fate decision. Their activity is regulated by pocket proteins, including pRb, p107, and p130. Here we show that E2F1 directly interacts with the ETS-related transcription factor GABPγ1 in vitro and in vivo. The binding domain interacting with GABPγ1 was mapped to the C-terminal amino acids 310 to 437 of E2F1, which include its transactivation and pRb binding domain. Among the E2F family of transcription factors, the interaction with GABPγ1 is restricted to E2F1. DNA-binding E2F1 complexes containing GABPγ1 are characterized by enhanced E2F1-dependent transcriptional activity. Moreover, GABPγ1 suppresses E2F1-dependent apoptosis by mechanisms other than the inhibition of the transactivation capacity of E2F1. In summary, our results provide evidence for a novel pRb-independent mechanism regulating E2F1-dependent transcription and apoptosis.

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Numb-Associated Kinase Interacts with the Phosphotyrosine Binding Domain of Numb and Antagonizes the Function of Numb In Vivo

Molecular and Cellular Biology ◽

10.1128/mcb.18.1.598 ◽

1998 ◽

Vol 18 (1) ◽

pp. 598-607 ◽

Cited By ~ 56

Author(s):

Cheng-ting Chien ◽

Shuwen Wang ◽

Michael Rothenberg ◽

Lily Y. Jan ◽

Yuh Nung Jan

Keyword(s):

Cell Division ◽

Cell Fate ◽

Asymmetric Cell Division ◽

Gene Dosage ◽

Specific Protein ◽

Physical Interaction ◽

Protein Protein Interaction ◽

Phosphotyrosine Binding Domain ◽

Ptb Domain

ABSTRACT During asymmetric cell division, the membrane-associated Numb protein localizes to a crescent in the mitotic progenitor and is segregated predominantly to one of the two daughter cells. We have identified a putative serine/threonine kinase, Numb-associated kinase (Nak), which interacts physically with the phosphotyrosine binding (PTB) domain of Numb. The PTB domains of Shc and insulin receptor substrate bind to an NPXY motif which is not present in the region of Nak that interacts with Numb PTB domain. We found that the Numb PTB domain but not the Shc PTB domain interacts with Nak through a peptide of 11 amino acids, implicating a novel and specific protein-protein interaction. Overexpression of Nak in the sensory organs causes both daughters of a normally asymmetric cell division to adopt the same cell fate, a transformation similar to the loss of numb function phenotype and opposite the cell fate transformation caused by overexpression of Numb. The frequency of cell fate transformation is sensitive to the numb gene dosage, as expected from the physical interaction between Nak and Numb. These findings indicate that Nak may play a role in cell fate determination during asymmetric cell divisions.

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In Vivo Inhibition of Antitumor Immunity by Embryonic Antigens

Tumori Journal ◽

10.1177/030089167606200211 ◽

1976 ◽

Vol 62 (2) ◽

pp. 211-218 ◽

Cited By ~ 1

Author(s):

Rosario Lembo ◽

Giorgio Parmiani

Keyword(s):

Tumor Cells ◽

Antitumor Immunity ◽

Normal Adult ◽

Embryonic Cells ◽

Diffusion Chambers ◽

Embryonic Tissues ◽

Embryonic Antigens ◽

Transplantation Antigens ◽

Adult Cells

Adult female BALB/c mice were immunized against the 3-methylcholanthrene–induced fibrosarcoma ST2 by growth and excision, and then injected with either neoplastic or embryonic mitomycin-C-blocked syngeneic cells before receiving a subcutaneous challenge of 105 ST2 cells. Other groups of similarly immunized females were intraperitoneally implanted with cell-impermeable diffusion chambers containing neoplastic or embryonic tissues, or mated to syngeneic males, before being challenged with ST2 cells. The mice immunized and injected with blocked ST2 but not with embryonic or antigenically unrelated neoplastic cells showed a resistance to the growth of ST2 significantly lower than that of immune mice given normal adult cells. This was particularly evident when ST2 blocked cells were given parenterally but was also detectable when ST2 blocked cells were cultured within diffusion chambers kept in tumor–excised mice. The syngeneic pregnancy had no effect on the antitumor immunity. In a subsequent study, BALB/c females were challenged with the same number of ST2 tumor cells as above, but the challenge was performed at days +4, +2, 0, –-2, –-4 from excision, and the blocked neoplastic embryonic or normal adult cells were given starting at the day of challenge. It was found that when the challenge was done 4 and 2 days after the excision, the administration of either antigenically-related tumor cells or embryonic cells could reduce the antitumor immune protection, while no such an effect was detectable in the other experimental groups. Thus, in addition to tumor-associated transplantation antigens, embryonic antigens also seem to be able to impair the developing antitumor immunity provided they are given within four days from the excision of the immunizing growth.

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The Relations between Cell Division and Cell Type Specification in Floral and Vegetative Meristems of Tomato and Arabidopsis

10.32747/1996.7613032.bard ◽

1996 ◽

Author(s):

Eliezer Lifschitz ◽

Elliot Meyerowitz

Keyword(s):

Cell Proliferation ◽

Cell Division ◽

Cell Fate ◽

Regulatory Gene ◽

Mads Box ◽

Mads Box Genes ◽

Threonine Deaminase ◽

Division Cell ◽

Floral Meristems

Meristems were the central issue of our project. Genes that are required for cell division, cell elongation, cell proliferation and cell fate were studied in the tomato system. The analysis of the dUTPase and threonine deaminase genes, along with the dissection of their regulatory regions is completed, while that of the RNR2 and PPO genes is at an advanced stage. All these genes were isolated in our laboratory. In addition, 8 different MADS box genes were studied in transgenic plants and their genetic relevances discovered. We have also shown that a given MADS box gene can modify the polarity of cell division without affecting the fate of the organ. In vivo interaction between two MADS box genes was demonstrated and the functional dependency of the tomato agamous gene on the TM5 gene product established. We have exploited the Knotted1 meristematic gene in conjunction with tomato leaf meristematic genes to show that simple and compound leaves and, for that matter, sepals and compound leaves, are formed by two different developmental programs. In this context we have also isolated and characterized the tomato Knotted1 gene (TKnl) and studied its expression pattern. A new program in which eight different meristematic genes in tomato will be studied emerged as a result of these studies. In essence, we have shown that it is possible to study and manipulate plant developmental systems using reverse genetic techniques and have provided a wealth of new molecular tools to interested colleagues working with tomato. Similarly, genes responsible for cell division, cell proliferation and cell fate were studied in Arabidopsis floral meristems. Among these genes are the TSO1, TSO2, HANABA TARANU and UNUSUAL FLORAL ORGANS genes, each affecting in its own way the number of pattern of cell divisions, and cell fate, in developing Arabodopsis flowers. In addition, new methods have been established for the assessment of the function of regulatory gene action in the different clonal layers of developing floral meristems.

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Probing tissue-scale deformation by in vivo force application reveals a fast tissue softening during early embryogenesis

10.1101/167155 ◽

2017 ◽

Cited By ~ 2

Author(s):

Arturo D’Angelo ◽

Kai Dierkes ◽

Carlo Carolis ◽

Guillaume Salbreux ◽

Jérôme Solon

Keyword(s):

Material Properties ◽

Rapid Change ◽

Early Embryogenesis ◽

Tissue Response ◽

Mechanical Parameters ◽

Major Contributor ◽

Pharmacological Treatments ◽

Tissue Material ◽

Over Time

AbstractDuring development, cell-generated forces induce tissue-scale deformations to shape the organism. Here, we present a method that allows to quantitatively relate such tissue-scale deformations to spatially localized forces and measure mechanical properties of epithelia in vivo. Our approach is based on the application of controlled forces on microparticles embedded in individual cells of an embryo. Combining measurements of the bead displacement with the analysis of induced deformation fields in a continuum mechanics framework, we can quantify tissue material properties and follow their change over time. In particular, we uncover a rapid change in tissue response occurring during Drosophila cellularization, resulting from a softening of the blastoderm and an increase of external friction. Pharmacological treatments reveal that in addition to actomyosin, the microtubule cytoskeleton is a major contributor to epithelial mechanics at that stage. Overall, our method allows for measuring essential mechanical parameters governing tissue-scale deformations and flows occurring during morphogenesis.

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An RNA Interference Screen Reveals Fate Determinants Implicated in Asymmetric Cell Division as Potential Regulators of Hematopoietic Stem Cell Self-Renewal

Blood ◽

10.1182/blood.v112.11.2462.2462 ◽

2008 ◽

Vol 112 (11) ◽

pp. 2462-2462

Author(s):

Kristin J Hope ◽

Sonia Cellot ◽

Stephen Ting ◽

Guy Sauvageau

Keyword(s):

Cell Death ◽

Stem Cell ◽

Cell Division ◽

Rna Interference ◽

Cell Fate ◽

Self Renewal ◽

Hematopoietic Stem ◽

Fate Determinants ◽

Cell Fate Determinants

Abstract During periods of extensive regeneration of the hematopoietic system, hematopoietic stem cells (HSC) undergo largely symmetrical self-renewal divisions, necessary to rapidly replenish the stem cell pool. Under homeostasis, however, it is likely that HSC rely more on asymmetric self-renewal divisions to retain an appropriate number of HSC while still enabling sufficient production of mature blood cells. The unequal partitioning of intrinsic fate determinants underlies the process of asymmetric stem cell division in lower organisms including Drosophila and C. elegans. The tumor suppressive function of specific determinants has been demonstrated in studies where mutation of fate determinants shown to be inhibitory to the self-renewal of one of the two daughter cells generated upon Drosophila neuroblast division, drives exclusive symmetrical stem cell divisions ultimately leading to the formation of larval brain tumors. As HSCs can not yet be definitively prospectively identified, it has been difficult to determine whether a similar segregation of such cell fate determinants underlies the asymmetric/symmetric self-renewal of these cells or whether deregulation of these determinants could also generate hematopoietic malignancies by inducing constitutive symmetric self-renewal divisions. We addressed these questions through a functional genetics approach taking advantage of systematic RNA interference to interrogate the function of polarity factors and cell fate determinants representing candidate HSC self-renewal regulators. From a list of 72 of such factors identified in the literature, 32 murine homologs were selected based on their differentially high level of expression in HSC-enriched populations. For each candidate we generated 3 unique short hairpin RNA (shRNA) encoding retroviral constructs also carrying EGFP for the purposes of following transduced cells. In a primary screen equal numbers of HSC-enriched Lin-CD150+CD48− cells were infected with the library in an arrayed 96-well format yielding an average gene transfer of 60.0 ± 3.2%. The in vivo reconstituting potential was then evaluated in a CRU assay such that identical proportions of each well were transplanted in duplicate. An average of 37.6 ± 5.1% long-term donor reconstitution was demonstrated by luciferase shRNA transduced controls. Directly following infection, the EGFP+ fraction of a portion of each well was separated by FACS to facilitate qRT-PCR determination of knockdown efficiency. Immunophenotypes, cell viability and morphology of well contents cultured an additional 7 days were also assessed. The percent of EGFP− and EGFP+ donor cell contribution was determined by flow cytometric evaluation of peripheral blood samples taken every 4 weeks for a period of 16 weeks. Genes for which shRNA vectors altered late transplant EGFP levels below or above defined thresholds were considered as hits. At present we have identified 4 genes for which shRNA-mediated depletion negatively affects repopulation but does not induce indiscriminate cell death in culture and 1 gene that may act as a self-renewal inhibitor. In one example, two shRNAs directed against the candidate EB3 showed a dramatic loss of EGFP+ cells in vivo. EB3, a member of the microtubule plus-end binding protein family, has previously described roles in the search-and-capture mechanism of spindle positioning. Interestingly, EB1, a closely related family member is also critical in directing the symmetrical as opposed to asymmetrical divisions of primitive neuroepithelial cells in Drosophila. Validation of all identified hits as well as further evaluation of their function through cell cycle, cell death and homing studies is ongoing.

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Reticulocytes II: Reexamination of the in vivo survival of stress reticulocytes

Blood ◽

10.1182/blood.v75.9.1877.bloodjournal7591877 ◽

1990 ◽

Vol 75 (9) ◽

pp. 1877-1882 ◽

Cited By ~ 1

Author(s):

NA Noble ◽

QP Xu ◽

LL Hoge

Keyword(s):

Cell Division ◽

Terminal Cell ◽

Intrinsic Properties ◽

Over Time

Very young reticulocytes are released into the circulation in response to the stress of anemia. These stress reticulocytes have shortened in vivo survival when transfused into normal recipients, and are generally considered to be abnormal because they have skipped a terminal cell division. We reevaluated one aspect of their abnormality: that of in vivo survival. Using methodology that accounted for all cells transfused, in vivo survival of both normal and stress reticulocytes was investigated in both normal and anemic recipients. The experiments demonstrate that: (1) survival of reticulocytes is normal only when normal reticulocytes are injected into nonanemic animals; (2) intrinsic properties of stress reticulocytes lead to their immediate removal from the circulation by normal recipients to a significantly greater extent than by anemic recipients; and (3) both stress and normal reticulocytes are removed at an accelerated rate over time by anemic recipients. Taken together, the data indicate that in the course of becoming anemic, an adaptation occurs that allows cells produced during anemia to circulate considerably longer in anemic animals than they could in normal nonanemic animals. Other studies disclosed that increased reticulocyte survival in anemic animals could not be attributed to reticuloendothelial overload, but is induced by adaptation of the spleen, decreasing its removal of stress reticulocytes.

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