Asymmetric cell division: fly neuroblast meets worm zygote

Both Drosophila neuroblasts and Caenorhabditis elegans zygotes use a conserved protein complex to establish cell polarity and regulate spindle orientation. Mammalian epithelia also use this complex to regulate apical/basal polarity. Recent results have allowed us to compare the mechanisms regulating asymmetric cell division in Drosophila neuroblasts and the C. elegans zygote.     

Photocontrol of the orientation of cell division inAdiantum

The rate of transition from one- to two-dimensional growth of fernAdiantum gametophytes under white light depends on the age of gametophyte cultured under red light. When gametophytes were cultured for longer period under red light, the rate of transition decreased and the number of abnormal gametophytes increased. Although the first step of the transition was the first longitudinal cell division following the two transverse ones (Wada and Furuya, 1970), the time-lapse-video study revealed that the apical cell of protonemata became flattened in the plane perpendicular to the incident ray of white light before the first longitudinal cell division. Analytical study of growing part of the apical cell with grains of activated charcoal as markers revealed that the apical cell flattening occurred evenly throughout the equatorial circumference of the cell even in the shaded side of the protonemata as well as in the side irradiated with white light.     

Photocontrol of the orientation of cell division in Adiantum

Summary Two-dimensional prothallia of Adiantum capillus-veneris always expanded in a plane which was at a right angle to any given direction of irradiation with continuous white light. The expansion began with a longitudinal division of the apical cell, in the filamentous protonema, and the orientation of the mitotic cell plate of this first longitudinal division as well as the subsequent divisions was always parallel to the direction of the incident light. When three irradiations with white light, interrupted by periods of darkness, were given, two transverse and one subsequent longitudinal division were induced. When the last two irradiations were given from the same direction, the cell plate of the first longitudinal division in most protonemata was oriented parallel to the direction of light. However, when the direction of light during the third irradiation was at right angle to that during the second, the frequency of the longitudinal division greatly decreased but that of the third transverse division increased. Thus, the orientation of the first longitudinal division appeared to be controlled in some way not only by the irradiation which actually induced the third division but also by that inducing the preceding transverse division, while the direction of light for the first transverse division had little effect on the orientation of the third division.     

Dikaryotic cell division of the fission yeast Schizosaccharomyces pombe

Dikaryons, cells with two haploid nuclei contributed by the members of a mating pair, are part of the life cycle of many filamentous fungi, but the molecular mechanisms underlying the division of dikaryons are largely unknown. We found that the fission yeast Schizosaccharomyces pombe has a latent ability to divide as a dikaryon. Cells capable of restarting the mitotic cycle with two nuclei were prepared by transient inactivation of the septation initiation network. Close pairing of the two nuclei before mitosis was dependent on minus-end-directed kinesin Klp2p and was essential for propagation as a dikaryon. The two spindles extended in opposite directions, keeping their old spindle pole bodies at the prospective site of cell division until the mid-anaphase. The spindles then overlapped, exchanging the inner nuclei. Finally, twin mitosis was followed by a single cytokinesis, producing two daughter dikaryons carrying copies of the original pair of nuclei.     

有丝分裂方向的影响因素

有丝分裂方向不仅关系到细胞增殖过程中子细胞的排列方式,而且对细胞分化、组织器官形态结构具有决定性作用。从细胞内环境和所处的微环境2个方面,讨论有丝分裂方向的影响因素。     

Cell division orientation and planar cell polarity pathways

The orientation of cell division has a crucial role in early embryo body plan specification, axis determination and cell fate diversity generation, as well as in the morphogenesis of tissues and organs. In many instances, cell division orientation is regulated by the planar cell polarity (PCP) pathways: the Wnt/Frizzled non-canonical pathway or the Fat/Dachsous/Four-jointed pathway. Firstly, using asymmetric cell division in both Drosophila and C. elegans, we describe the central role of the Wnt/Frizzled pathway in the regulation of asymmetric cell division orientation, focusing on its cooperation with either the Src kinase pathway or the heterotrimeric G protein pathway. Secondly, we describe our present understanding of the mechanisms by which the planar cell polarity pathways drive tissue morphogenesis by regulating the orientation of symmetric cell division within a field of cells. Finally, we will discuss the important avenues that need to be explored in the future to better understand how planar cell polarity pathways control embryo body plan determination, cell fate specification or tissue morphogenesis by mitotic spindle orientation.     

The extracellular matrix guides the orientation of the cell division axis

The cell division axis determines the future positions of daughter cells and is therefore critical for cell fate. The positioning of the division axis has been mostly studied in systems such as embryos or yeasts, in which cell shape is well defined. In these cases, cell shape anisotropy and cell polarity affect spindle orientation. It remains unclear whether cell geometry or cortical cues are determinants for spindle orientation in mammalian cultured cells. The cell environment is composed of an extracellular matrix (ECM), which is connected to the intracellular actin cytoskeleton via transmembrane proteins. We used micro-contact printing to control the spatial distribution of the ECM on the substrate and demonstrated that it has a role in determining the orientation of the division axis of HeLa cells. On the basis of our analysis of the average distributions of actin-binding proteins in interphase and mitosis, we propose that the ECM controls the location of actin dynamics at the membrane, and thus the segregation of cortical components in interphase. This segregation is further maintained on the cortex of mitotic cells and used for spindle orientation.     

The SIDECAR POLLEN gene encodes a microspore-specific LOB/AS2 domain protein required for the correct timing and orientation of asymmetric cell division

Control of ion loading into the xylem has been repeatedly named as a crucial factor determining plant salt tolerance. In this study we further investigate this issue by applying a range of biophysical [the microelectrode ion flux measurement (MIFE) technique for non‐invasive ion flux measurements, the patch clamp technique, membrane potential measurements] and physiological (xylem sap and tissue nutrient analysis, photosynthetic characteristics, stomatal conductance) techniques to barley varieties contrasting in their salt tolerance. We report that restricting Na⁺ loading into the xylem is not essential for conferring salinity tolerance in barley, with tolerant varieties showing xylem Na⁺ concentrations at least as high as those of sensitive ones. At the same time, tolerant genotypes are capable of maintaining higher xylem K⁺/Na⁺ ratios and efficiently sequester the accumulated Na⁺ in leaves. The former is achieved by more efficient loading of K⁺ into the xylem. We argue that the observed increases in xylem K⁺ and Na⁺ concentrations in tolerant genotypes are required for efficient osmotic adjustment, needed to support leaf expansion growth. We also provide evidence that K⁺‐permeable voltage‐sensitive channels are involved in xylem loading and operate in a feedback manner to maintain a constant K⁺/Na⁺ ratio in the xylem sap.     

An essential function of sphingolipids in yeast cell division

Ceramides are bioactive lipids and precursors to sphingolipids. They have been shown to take part in a wide variety of different physiological processes in eukaryotic organisms and are thought to be toxic at high concentrations. Ceramide is synthesized by condensation of the sphingoid base sphinganine and a fatty acyl CoA by ceramide synthases, a family of enzymes that differ in their specificity for the length of the acyl CoA substrate. We have engineered a yeast strain where the endogenous ceramide synthase has been replaced by one of the putative enzymes from cotton. As a result, the yeast strain produces C18 rather than C26 ceramides showing that the cotton protein is a bona fide ceramide synthase with specificity towards C18 acyl CoA. Strikingly, the accumulation of C18 ceramide is not toxic in Saccharomyces cerevisiae. This allows survival of the yeast after deletion of the normally essential AUR1 (inositol phosphorylceramide synthase) gene permitting us to address the essential roles of sphingolipids. Deletion of AUR1 allows cell growth, but leads to a defect in cytokinesis, which takes twice as long as in wild-type strains. Nuclear division and recruitment of septins is apparently not affected, but cytokinesis is delayed and cell separation is incomplete.     

The role of cAMP in controlling yeast cell division

The studies on the cAMP-requiring mutants and their suppressors in the yeast, Saccharomyces cerevisiae, revealed that cAMP-dependent protein phosphorylation is involved in the G1 phase of the cell cycle, in conjugation, and in the post-meiotic stage of sporulation, and that inhibition of cAMP-dependent protein phosphorylation is required to induce meiotic division.     

Mutagenesis of yeast by hydrazine: dependence upon post-treatment cell division.

Hydrazine was found to be mutagenic for yeast (Saccharomyces cerevisiae) at exposures (concentration × time) ranging over nearly three orders of magnitude. Little or no forward mutation from CAN1 to can1 was detectable upon immediate plating following treatment in neutral buffer suspension. Post-treatment cell division in yeast extract peptone dextrose complex growth medium was required for expression of induced mutation to canavanine resistance. Frequencies of induced mutation rose to levels approximately 10-fold higher than spontaneous levels for exposures between 0.1 and 12.0 min mol/l. Survival remained at 100%. For exposures greater than 80 min mol/l viability and mutation frequency began to decrease sharply. By contrast, single treatments of ethyl methanesulfonate, methyl methanesulfonate, N-methyl-N′-nitro-N-nitro-soguanidine, nitrous acid, hydroxylamine, and ultraviolet light were able to increase mutation frequency with this system upon immediate assay. Further growth-dependent increases in mutation frequency were not observed with HA and UV.Expression of HZ-induced mutation was detectable after treated cells had undergone less than one population doubling in YEPD. Such mutation expression could be blocked by the inhibitors cycloheximide and hydroxyurea, which block protein synthesis and DNA synthesis respectively. Results were similar to those obtained previously with Haemophilus influenzae and similarly suggest that, in this eukaryote, HZ-induced lesions lead to mutation by causing base mispairing at DNA replication rather than by means of an error-prone repair mechanism.     

Symmetric cell division in pseudohyphae of the yeast Saccharomyces cerevisiae.

Laboratory strains of Saccharomyces cerevisiae are dimorphic; in response to nitrogen starvation they switch from a yeast form (YF) to a filamentous pseudohyphal (PH) form. Time-lapse video microscopy of dividing cells reveals that YF and PH cells differ in their cell cycles and budding polarity. The YF cell cycle is controlled at the G1/S transition by the cell-size checkpoint Start. YF cells divide asymmetrically, producing small daughters from full-sized mothers. As a result, mothers and daughters bud asynchronously. Mothers bud immediately but daughters grow in G1 until they achieve a critical cell size. By contrast, PH cells divide symmetrically, restricting mitosis until the bud grows to the size of the mother. Thus, mother and daughter bud synchronously in the next cycle, without a G1 delay before Start. YF and PH cells also exhibit distinct bud-site selection patterns. YF cells are bipolar, producing their second and subsequent buds at either pole. PH cells are unipolar, producing their second and subsequent buds only from the end opposite the junction with their mother. We propose that in PH cells a G2 cell-size checkpoint delays mitosis until bud size reaches that of the mother cell. We conclude that yeast and PH forms are distinct cell types each with a unique cell cycle, budding pattern, and cell shape.     

Automated spectrophotometric assay for cell division regulation in yeast

A spectrophotometric assay is presented for monitoring the regulation of cell division by the polypeptide alpha-factor in cultures of living cells of Saccharomyces cerevisiae yeast. This assay is simple, automated, and may have wider application in the study of other eucaryotic cells that do not require anchorage for cell growth. The kinetics of absorbance change were monitored continuously over time in yeast cell cultures that were mixed and aerated in cuvettes fitted with top-loading propeller stirrers. The absorbance doubling time. TD(Abs), was identical to the cell number doubling time in the absence of cell division arrest by alpha-factor. alpha-Factor lengthened the TD(Abs) during division arrest. At pH 5.8, 10(5) 381G cells/ml, the Khalf-maximal was 250 +/- 50 nM alpha-factor for the TD(Abs) increase during arrest, with a maximum increase of five-fold. After a period of time the TD(Abs) abruptly shortened. This is defined as the spectrophotometric recovery time (RTspec) and was compared to the time of recovery that is due to the reinitiation of cell division monitored by bud emergence (RTBE). RTBE occurred 40 +/- 5 min prior to RTspec when recovery was spontaneous or was artificially induced by the removal of alpha-factor (pH 5.8, 381G). The difference between RTBE and RTspec was independent of alpha-factor concentration between 0.05 and 1 microM and cell concentration between 1 and greater than or equal to 25 x 10(5) cells/ml (pH 5.8, 381G) but was both pH and cell strain dependent. At pH 5.8 and 2.7 the recovery from arrest occurred by inactivation of alpha-factor. The TD(Abs) increase during arrest appears to be due to an alpha-factor-induced inhibition of net cell mass increase, an effect that has not been reported previously. Evidence is presented that this process is also correlated with the formation of cell projections.     

Real-time monitoring of yeast cell division by dielectric spectroscopy.

To assay cell cycle progression in synchronized culture of yeast we have applied dielectric spectroscopy to its real-time monitoring. The dielectric monitoring is based on the electromagnetic induction method, regarded as a nonelectrode method, which has resolved the problems encountered in measurements with metal electrodes, namely electrode polarization and bubble formation on electrodes. In the synchronized culture with temperature-sensitive cell division cycle mutants, the permittivity of the culture broth showed cyclic changes at frequencies below 300 kHz. The increase and decrease in the cyclic changes of the relative permittivity correspond to the increase in cell length and bud size and to the septum formation between mother and daughter cells, respectively.     

A survey of essential gene function in the yeast cell division cycle

Mutations impacting specific stages of cell growth and division have provided a foundation for dissecting mechanisms that underlie cell cycle progression. We have undertaken an objective examination of the yeast cell cycle through flow cytometric analysis of DNA content in TetO(7) promoter mutant strains representing 75% of all essential yeast genes. More than 65% of the strains displayed specific alterations in DNA content, suggesting that reduced function of an essential gene in most cases impairs progression through a specific stage of the cell cycle. Because of the large number of essential genes required for protein biosynthesis, G1 accumulation was the most common phenotype observed in our analysis. In contrast, relatively few mutants displayed S-phase delay, and most of these were defective in genes required for DNA replication or nucleotide metabolism. G2 accumulation appeared to arise from a variety of defects. In addition to providing a global view of the diversity of essential cellular processes that influence cell cycle progression, these data also provided predictions regarding the functions of individual genes: we identified four new genes involved in protein trafficking (NUS1, PHS1, PGA2, PGA3), and we found that CSE1 and SMC4 are important for DNA replication.     

The orientation of cell division influences cell-fate choice in the developing mammalian retina

Asymmetric segregation of cell-fate determinants during cell division plays an important part in generating cell diversity in invertebrates. We showed previously that cells in the neonatal rat retina divide at various orientations and that some dividing cells asymmetrically distribute the cell-fate determinant Numb to the two daughter cells. Here, we test the possibility that such asymmetric divisions contribute to retinal cell diversification. We have used long-term videomicroscopy of green-fluorescent-protein (GFP)-labeled retinal explants from neonatal rats to visualize the plane of cell division and follow the differentiation of the daughter cells. We found that cells that divided with a horizontal mitotic spindle, where both daughter cells should inherit Numb, tended to produce daughters that became the same cell type, whereas cells that divided with a vertical mitotic spindle, where only one daughter cell should inherit Numb, tended to produce daughters that became different. Moreover, overexpression of Numb in the dividing cells promoted the development of photoreceptor cells at the expense of interneurons and Müller glial cells. These findings indicate that the plane of cell division influences cell-fate choice in the neonatal rat retina and support the hypothesis that the asymmetric segregation of Numb normally influences some of these choices.