Distribution Of Cell Fate Determinants Research Articles

The spindle assembly checkpoint and the spatial activation of Polo kinase determine the duration of cell division and prevent tumor formation.

The maintenance of a restricted pool of asymmetrically dividing stem cells is essential for tissue homeostasis. This process requires the control of mitotic progression that ensures the accurate chromosome segregation. In addition, this event is coupled to the asymmetric distribution of cell fate determinants in order to prevent stem cell amplification. How this coupling is regulated remains poorly described. Here, using asymmetrically dividing Drosophila neural stem cells (NSCs), we show that Polo kinase activity levels determine timely Cyclin B degradation and mitotic progression independent of the spindle assembly checkpoint (SAC). This event is mediated by the direct phosphorylation of Polo kinase by Aurora A at spindle poles and Aurora B kinases at centromeres. Furthermore, we show that Aurora A-dependent activation of Polo is the major event that promotes NSC polarization and together with the SAC prevents brain tumor growth. Altogether, our results show that an Aurora/Polo kinase module couples NSC mitotic progression and polarization for tissue homeostasis.

Multifunctional protein 4.1R regulates the asymmetric segregation of Numb during terminal erythroid maturation

The asymmetric cell division of stem or progenitor cells generates daughter cells with distinct fates that balance proliferation and differentiation. Asymmetric segregation of Notch signaling regulatory protein Numb plays a crucial role in cell diversification. However, the molecular mechanism remains unclear. Here, we examined the unequal distribution of Numb in the daughter cells of murine erythroleukemia cells (MELCs) that undergo DMSO-induced erythroid differentiation. In contrast to the cytoplasmic localization of Numb during uninduced cell division, Numb is concentrated at the cell boundary in interphase, near the one-spindle pole in metaphase, and is unequally distributed to one daughter cell in anaphase in induced cells. The inheritance of Numb guides this daughter cell toward erythroid differentiation while the other cell remains a progenitor cell. Mitotic spindle orientation, critical for distribution of cell fate determinants, requires complex communication between the spindle microtubules and the cell cortex mediated by the NuMA-LGN-dynein/dynactin complex. Depletion of each individual member of the complex randomizes the position of Numb relative to the mitotic spindle. Gene replacement confirms that multifunctional erythrocyte protein 4.1R (4.1R) functions as a member of the NuMA-LGN-dynein/dynactin complex and is necessary for regulating spindle orientation, in which interaction between 4.1R and NuMA plays an important role. These results suggest that mispositioning of Numb is the result of spindle misorientation. Finally, disruption of the 4.1R-NuMA-LGN complex increases Notch signaling and decreases the erythroblast population. Together, our results identify a critical role for 4.1R in regulating the asymmetric segregation of Numb to mediate erythropoiesis.

The stem cell division theory of cancer.

All cancer registries constantly show striking differences in cancer incidence by age and among tissues. For example, lung cancer is diagnosed hundreds of times more often at age 70 than at age 20, and lung cancer in nonsmokers occurs thousands of times more frequently than heart cancer in smokers. An analysis of these differences using basic concepts in cell biology indicates that cancer is the end-result of the accumulation of cell divisions in stem cells. In other words, the main determinant of carcinogenesis is the number of cell divisions that the DNA of a stem cell has accumulated in any type of cell from the zygote. Cell division, process by which a cell copies and separates its cellular components to finally split into two cells, is necessary to produce the large number of cells required for living. However, cell division can lead to a variety of cancer-promoting errors, such as mutations and epigenetic mistakes occurring during DNA replication, chromosome aberrations arising during mitosis, errors in the distribution of cell-fate determinants between the daughter cells, and failures to restore physical interactions with other tissue components. Some of these errors are spontaneous, others are promoted by endogenous DNA damage occurring during quiescence, and others are influenced by pathological and environmental factors. The cell divisions required for carcinogenesis are primarily caused by multiple local and systemic physiological signals rather than by errors in the DNA of the cells. As carcinogenesis progresses, the accumulation of DNA errors promotes cell division and eventually triggers cell division under permissive extracellular environments. The accumulation of cell divisions in stem cells drives not only the accumulation of the DNA alterations required for carcinogenesis, but also the formation and growth of the abnormal cell populations that characterize the disease. This model of carcinogenesis provides a new framework for understanding the disease and has important implications for cancer prevention and therapy.

Asymmetric Localization and Distribution of Factors Determining Cell Fate During Early Development of Xenopus laevis.

Asymmetric division is a property of eukaryotic cells that is fundamental to the formation of higher life forms. Despite its importance, the mechanism behind it remains elusive. Asymmetry in the cell is induced by polarization of cell fate determinants that become unevenly distributed among progeny cells. So far dozens of determinants have been identified. Xenopus laevis is an ideal system to study asymmetric cell division during early development, because of the huge size of its oocytes and early-stage blastomeres. Here, we present the current knowledge about localization and distribution of cell fate determinants along the three body axes: animal-vegetal, dorsal-ventral, and left-right. Uneven distribution of cell fate determinants during early development specifies the formation of the embryonic body plan.

Asymmetries in Cell Division, Cell Size, and Furrowing in the Xenopus laevis Embryo.

Asymmetric cell divisions produce two daughter cells with distinct fate. During embryogenesis, this mechanism is fundamental to build tissues and organs because it generates cell diversity. In adults, it remains crucial to maintain stem cells. The enthusiasm for asymmetric cell division is not only motivated by the beauty of the mechanism and the fundamental questions it raises, but has also very pragmatic reasons. Indeed, misregulation of asymmetric cell divisions is believed to have dramatic consequences potentially leading to pathogenesis such as cancers. In diverse model organisms, asymmetric cell divisions result in two daughter cells, which differ not only by their fate but also in size. This is the case for the early Xenopus laevis embryo, in which the two first embryonic divisions are perpendicular to each other and generate two pairs of blastomeres, which usually differ in size: one pair of blastomeres is smaller than the other. Small blastomeres will produce embryonic dorsal structures, whereas the larger pair will evolve into ventral structures. Here, we present a speculative model on the origin of the asymmetry of this cell division in the Xenopus embryo. We also discuss the apparently coincident asymmetric distribution of cell fate determinants and cell-size asymmetry of the 4-cell stage embryo. Finally, we discuss the asymmetric furrowing during epithelial cell cytokinesis occurring later during Xenopus laevis embryo development.

Bällchen participates in proliferation control and prevents the differentiation of Drosophila melanogaster neuronal stem cells.

ABSTRACTStem cells continuously generate differentiating daughter cells and are essential for tissue homeostasis and development. Their capacity to self-renew as undifferentiated and actively dividing cells is controlled by either external signals from a cellular environment, the stem cell niche, or asymmetric distribution of cell fate determinants during cell division. Here we report that the protein kinase Bällchen (BALL) is required to prevent differentiation as well as to maintain normal proliferation of neuronal stem cells of Drosophila melanogaster, called neuroblasts. Our results show that the brains of ball mutant larvae are severely reduced in size, which is caused by a reduced proliferation rate of the neuroblasts. Moreover, ball mutant neuroblasts gradually lose the expression of the neuroblast determinants Miranda and aPKC, suggesting their premature differentiation. Our results indicate that BALL represents a novel cell intrinsic factor with a dual function regulating the proliferative capacity and the differentiation status of neuronal stem cells during development.

Planar Cell Polarity Goes Perpendicular

The planar cell polarity pathway influences the apicobasal distribution of a cell fate determinant.

Abstract 2298: Cell-specific effects of BRafV600E expression disrupt gliogenesis and lead to astrocytoma formation.

Abstract Within the adult mammalian brain the asymmetric distribution of cell-fate determinants during mitosis maintains a balance between differentiated and self-renewing cells. We have previously shown that disruption of asymmetric divisions in oligodendrocyte progenitor cells (OPC) causes cells to aberrantly self-renew and fail to differentiate, giving rise to oligodendroglioma (Sugiarto et al. Cancer Cell; 2011; 20; 320-40). Here we proposed to address whether asymmetric divisions are also disrupted in precursors of malignant astrocytoma, a highly aggressive and therapy resistant type of glioma. The constitutively active BRaf mutant BRafV600E is mutated in a subset of malignant astrocytoma alongside deletion of Cdkn2a, and can transform NSCs into astrocytoma precursors (Huillard et al. PNAS; 2012; 109; 8710-5). We used BRafCA mice (Dankort et al. Genes Dev. 2007; 21; 379-84) with Cre recombinase-activated expression of BRafV600E and deletion of Ink4a/Arf to generate an in situ model of NSC-derived malignant astrocytoma. In vivo studies to assess asymmetric division and related defects were complemented by studies of BRafV600E expressing, Ink4a/Arf-deleted neurosphere cells in cell-based assays of asymmetric division, self-renewal and differentiation. Our results provide unprecedented evidence that BRafV600E, although uniformly expressed in NSC and their progeny, disrupts asymmetric divisions in a cell-type specific manner. NSCs continue to divide asymmetrically but their progeny, including OPC, divide more symmetrically and exhibit defects in differentiation and proliferation. These defects are reversible with BRafV600E-specific inhibitor PLX4720, and are associated with increased expression of the mitotic kinase polo-like kinase 1, increased pERK and decreased pAKT. These data describe a mechanism by which NSC-targeted expression of BRafV600E may generate numerous subpopulations of tumor cells with distinct characteristics. This heterogeneity may mirror that seen in malignant astrocytoma, which has often been associated with high incidence of therapy resistance (Burger et al. Cancer. 1985; 56; 1106-11). Citation Format: Robin G. Lerner, Yuichiro Ihara, Brian Reichholf, Dian Qu, Sista Sugiarto, Amelie Griveau, David James, Martin McMahon, Claudia Petritsch. Cell-specific effects of BRafV600E expression disrupt gliogenesis and lead to astrocytoma formation. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2298. doi:10.1158/1538-7445.AM2013-2298

Microtubule-dependent path to the cell cortex for cytoplasmic dynein in mitotic spindle orientation

During animal development, microtubules (MTs) play a major role in directing cellular and subcellular patterning, impacting cell polarization and subcellular organization, thereby affecting cell fate determination and tissue architecture. In particular, when progenitor cells divide asymmetrically along an anterior-posterior or apical-basal axis, MTs must coordinate the position of the mitotic spindle with the site of cell division to ensure normal distribution of cell fate determinants and equal sequestration of genetic material into the two daughter cells. Emerging data from diverse model systems have led to the prevailing view that, during mitotic spindle positioning, polarity cues at the cell cortex signal for the recruitment of NuMA and the minus-end directed MT motor cytoplasmic dynein.1 The NuMA/dynein complex is believed to connect, in turn, to the mitotic spindle via astral MTs, thus aligning and tethering the spindle, but how this connection is achieved faithfully is unclear. Do astral MTs need to search for and then capture cortical NuMA/dynein? How does dynein capture the astral MTs emanating from the correct spindle pole? Recently, using the classical model of asymmetric cell division—budding yeast S. cerevisiae—we successfully demonstrated that astral MTs assume an active role in cortical dynein targeting, in that astral MTs utilize their distal plus ends to deliver dynein to the daughter cell cortex, the site where dynein activity is needed to perform its spindle alignment function. This observation introduced the novel idea that, during mitotic spindle orientation processes, polarity cues at the cell cortex may actually signal to prime the cortical receptors for MT-dependent dynein delivery. This model is consistent with the observation that dynein/dynactin accumulate prominently at the astral MT plus ends during metaphase in a wide range of cultured mammalian cells.

Transcriptome asymmetry within mouse zygotes but not between early embryonic sister blastomeres

Transcriptome regionalization is an essential polarity determinant among metazoans, directing embryonic axis formation during normal development. Although conservation of this principle in mammals is assumed, recent evidence is conflicting and it is not known whether transcriptome asymmetries exist within unfertilized mammalian eggs or between the respective cleavage products of early embryonic divisions. We here address this by comparing transcriptome profiles of paired single cells and sub-cellular structures obtained microsurgically from mouse oocytes and totipotent embryos. Paired microsurgical spindle and remnant samples from unfertilized metaphase II oocytes possessed distinguishable profiles. Fertilization produces a totipotent 1-cell embryo (zygote) and associated spindle-enriched second polar body whose paired profiles also differed, reflecting spindle transcript enrichment. However, there was no programmed transcriptome asymmetry between sister cells within 2- or 3-cell embryos. Accordingly, there is transcriptome asymmetry within mouse oocytes, but not between the sister blastomeres of early embryos. This work places constraints on pre-patterning in mammals and provides documentation correlating potency changes and transcriptome partitioning at the single-cell level.

MAP Kinase Signaling Antagonizes PAR-1 Function During Polarization of the Early Caenorhabditis elegans Embryo

PAR proteins (partitioning defective) are major regulators of cell polarity and asymmetric cell division. One of the par genes, par-1, encodes a Ser/Thr kinase that is conserved from yeast to mammals. In Caenorhabditis elegans, par-1 governs asymmetric cell division by ensuring the polar distribution of cell fate determinants. However the precise mechanisms by which PAR-1 regulates asymmetric cell division in C. elegans remain to be elucidated. We performed a genomewide RNAi screen and identified six genes that specifically suppress the embryonic lethal phenotype associated with mutations in par-1. One of these suppressors is mpk-1, the C. elegans homolog of the conserved mitogen activated protein (MAP) kinase ERK. Loss of function of mpk-1 restored embryonic viability, asynchronous cell divisions, the asymmetric distribution of cell fate specification markers, and the distribution of PAR-1 protein in par-1 mutant embryos, indicating that this genetic interaction is functionally relevant for embryonic development. Furthermore, disrupting the function of other components of the MAPK signaling pathway resulted in suppression of par-1 embryonic lethality. Our data therefore indicates that MAP kinase signaling antagonizes PAR-1 signaling during early C. elegans embryonic polarization.

Why cells move messages: The biological functions of mRNA localization

RNA localization is a widespread mechanism that allows cells to spatially control protein function by determining their sites of synthesis. In embryos, localized mRNAs are involved in morphogen gradient formation or the asymmetric distribution of cell fate determinants. In somatic cell types, mRNA localization contributes to local assembly of protein complexes or facilitates protein targeting to organelles. Long-distance transport of specific mRNAs in plants allows coordination of developmental processes between different plant organs. In this review, we will discuss the biological significance of different patterns of mRNA localization.

Taking the Time to Determine Cell Fate

Calegari and Huttner exposed mouse embryos to an inhibitor of cyclin-dependent kinases (CDKs) to investigate the hypothesis that an increase in the length of the cell cycle might be a cause (rather than a consequence) of neuronal differentiation. During the development of multicellular organisms, cell division can yield daughter cells with different fates. This is associated with unequal distribution of cell fate determinants to daughter cells; however, such an unequal distribution does not always lead to distinct cell fates. Neuroepithelial (NE) cells, which give rise to all of the neurons in the mammalian central nervous system, switch from a proliferative to a neuron-generating mode in a distinct spatiotemporal pattern, a switch associated with an increase in the G 1 phase of the cell cycle. Calegari and Huttner grew whole mouse embryos in vitro in the presence of the CDK inhibitor olomoucine or an inactive isomer. Moderate concentrations of olomoucine slowed NE cell cycle length, assessed by cumulative labeling with bromodeoxyuridine, without increasing apoptosis, affecting cell size, or blocking the cell cycle. This was associated with a premature switch of NE cells from proliferative to neuron-generating mode and with the premature appearance of neurons, both assessed by immunofluorescence analysis of specific markers (TIS21 and MAP2, respectively). The authors proposed a model in which the interaction between the amount of particular cell fate determinants and the length of time over which such determinants act during the cell cycle determines cell fate and suggested that TIS21, which appears in neuron-generating cells and prolongs G 1 , itself promotes neurogenesis. F. Calegari, W. B. Huttner, An inhibition of cyclin-dependent kinases that lengthens, but does not arrest, neuroepithelial cell cycle induces premature neurogenesis. J. Cell Sci. 116 , 4947-4955 (2003). [Abstract] [Full Text]

Rapsynoid/Partner of Inscuteable Controls Asymmetric Division of Larval Neuroblasts inDrosophila

Asymmetric cell division generates daughter cells with different developmental fates. In Drosophila neuroblasts, asymmetric divisions are characterized by (1) a difference in size between the two daughter cells and (2) an asymmetric distribution of cell fate determinants, including Prospero and Numb, between the two daughter cells. In embryonic neuroblasts, the asymmetric localization of cell fate determinants is under the control of the protein Inscuteable (Insc), which is itself localized asymmetrically as an apical crescent. Here, we describe a new Drosophila protein, Rapsynoid (Raps), which interacts in a two-hybrid assay with the signal transduction protein Galpha(i). We show that Raps is localized asymmetrically in dividing larval neuroblasts and colocalizes with Insc. Moreover, in raps mutants, the asymmetric divisions of neuroblasts are altered: (1) Insc is no longer asymmetrically localized in the dividing neuroblast; and (2) the neuroblast division produces two daughter cells of similar sizes. However, the morphologically symmetrical divisions of raps neuroblasts still lead to daughter cells with different fates, as shown by differences in gene expression. Our data show that Raps is a novel protein involved in the control of asymmetric divisions of neuroblasts.

Controlling cell cycle and cell fate: common strategies in prokaryotes and eukaryotes.

Generating cells of different fate within a group of equipotent cells is the underlying principle of the development of complex multicellular organisms. Cell–cell signaling or environmental differences may cause two equipotent daughter cells to assume different fates. Alternatively, asymmetric distribution of cell fate determinants during cell division can generate two cells with different fates. In either case, developmental cues have to be interpreted correctly by the machinery that controls cell cycle progression and by the programs that induce cell specification. Despite great advances in identifying the proteins that govern cell cycle progression, little is known about how their activity is modulated by developmental signals. In this issue of the Proceedings , Quon et al. (1) provide new insights into understanding this fundamental problem. They identified an inhibitor of DNA replication whose activity is temporally and spatially regulated, thereby restricting its activity during development. However, these findings were not made, as one might have expected, in a eukaryotic organism, but in the eubacterium Caulobacter crescentus . The similarities in the regulation of this bacterial cell cycle regulator and cell fate determinant and that of eukaryotic cell cycle regulators and cell fate determinants will be discussed in this commentary. Cell division in Caulobacter is asymmetric (2). A sessile stalked cell divides to give rise to two daughter cells, a stalked cell and a motile swarmer cell (Fig. 1 A ). The stalked cell is capable of immediately replicating its chromosome and subsequently dividing, whereas the swarmer cell must differentiate into a stalked cell before it can initiate DNA replication and cell division. The generation of a stalked cell and a swarmer cell during cell division and the initiation of DNA replication after the transition of a swarmer cell into a stalked cell are ultimately controlled by the CtrA protein (for cell cycle …

Asymmetry and cell fate in the Drosophila embryonic CNS.

Drosophila CNS precursors, neuroblasts, repeatedly divide to produce a large neuroblast and a smaller GMC. This division is asymmetric with regard to sibling cell size, mitotic potential and gene expression. Recent work has identified a number of molecules that show a polarized distribution during neuroblast mitosis: prospero RNA and Inscuteable, Miranda, Prospero, Staufen, and Numb proteins. The process of asymmetric localization of proteins and RNAs is cell cycle dependent, microfilament dependent and coordinated with the positioning of the mitotic spindle, which results in the unequal distribution of cell fate determinants to a specific daughter cell at cytokinesis.