Genetic Manipulation of Glycogen Allocation Affects Replicative Lifespan in E. coli

In bacteria, replicative aging manifests as a difference in growth or survival between the two cells emerging from division. One cell can be regarded as an aging mother with a decreased potential for future survival and division, the other as a rejuvenated daughter. Here, we aimed at investigating some of the processes involved in aging in the bacterium Escherichia coli, where the two types of cells can be distinguished by the age of their cell poles. We found that certain changes in the regulation of the carbohydrate metabolism can affect aging. A mutation in the carbon storage regulator gene, csrA, leads to a dramatically shorter replicative lifespan; csrA mutants stop dividing once their pole exceeds an age of about five divisions. These old-pole cells accumulate glycogen at their old cell poles; after their last division, they do not contain a chromosome, presumably because of spatial exclusion by the glycogen aggregates. The new-pole daughters produced by these aging mothers are born young; they only express the deleterious phenotype once their pole is old. These results demonstrate how manipulations of nutrient allocation can lead to the exclusion of the chromosome and limit replicative lifespan in E. coli, and illustrate how mutations can have phenotypic effects that are specific for cells with old poles. This raises the question how bacteria can avoid the accumulation of such mutations in their genomes over evolutionary times, and how they can achieve the long replicative lifespans that have recently been reported.     

Preferential retrotransposition in aging yeast mother cells is correlated with increased genome instability

Retrotransposon expression or mobility is increased with age in multiple species and could promote genome instability or altered gene expression during aging. However, it is unclear whether activation of retrotransposons during aging is an indirect result of global changes in chromatin and gene regulation or a result of retrotransposon-specific mechanisms. Retromobility of a marked chromosomal Ty1 retrotransposon in Saccharomyces cerevisiae was elevated in mother cells relative to their daughter cells, as determined by magnetic cell sorting of mothers and daughters. Retromobility frequencies in aging mother cells were significantly higher than those predicted by cell age and the rate of mobility in young populations, beginning when mother cells were only several generations old. New Ty1 insertions in aging mothers were more strongly correlated with gross chromosome rearrangements than in young cells and were more often at non-preferred target sites. Mother cells were more likely to have high concentrations and bright foci of Ty1 Gag–GFP than their daughter cells. Levels of extrachromosomal Ty1 cDNA were also significantly higher in aged mother cell populations than their daughter cell populations. These observations are consistent with a retrotransposon-specific mechanism that causes retrotransposition to occur preferentially in yeast mother cells as they begin to age, as opposed to activation by phenotypic changes associated with very old age. These findings will likely be relevant for understanding retrotransposons and aging in many organisms, based on similarities in regulation and consequences of retrotransposition in diverse species.     

A mutation in the ATP2 gene abrogates the age asymmetry between mother and daughter cells of the yeast Saccharomyces cerevisiae

The yeast Saccharomyces cerevisiae reproduces by asymmetric cell division, or budding. In each cell division, the daughter cell is usually smaller and younger than the mother cell, as defined by the number of divisions it can potentially complete before it dies. Although individual yeast cells have a limited life span, this age asymmetry between mother and daughter ensures that the yeast strain remains immortal. To understand the mechanisms underlying age asymmetry, we have isolated temperature-sensitive mutants that have limited growth capacity. One of these clonal-senescence mutants was in ATP2, the gene encoding the beta-subunit of mitochondrial F(1), F(0)-ATPase. A point mutation in this gene caused a valine-to-isoleucine substitution at the ninetieth amino acid of the mature polypeptide. This mutation did not affect the growth rate on a nonfermentable carbon source. Life-span determinations following temperature shift-down showed that the clonal-senescence phenotype results from a loss of age asymmetry at 36 degrees, such that daughters are born old. It was characterized by a loss of mitochondrial membrane potential followed by the lack of proper segregation of active mitochondria to daughter cells. This was associated with a change in mitochondrial morphology and distribution in the mother cell and ultimately resulted in the generation of cells totally lacking mitochondria. The results indicate that segregation of active mitochondria to daughter cells is important for maintenance of age asymmetry and raise the possibility that mitochondrial dysfunction may be a normal cause of aging. The finding that dysfunctional mitochondria accumulated in yeasts as they aged and the propensity for old mother cells to produce daughters depleted of active mitochondria lend support to this notion. We propose, more generally, that age asymmetry depends on partition of active and undamaged cellular components to the progeny and that this "filter" breaks down with age.     

Asymmetric Numb distribution is critical for asymmetric cell division of mouse cerebral cortical stem cells and neuroblasts

Stem cells and neuroblasts derived from mouse embryos undergo repeated asymmetric cell divisions, generating neural lineage trees similar to those of invertebrates. In Drosophila, unequal distribution of Numb protein during mitosis produces asymmetric cell divisions and consequently diverse neural cell fates. We investigated whether a mouse homologue m-numb had a similar role during mouse cortical development. Progenitor cells isolated from the embryonic mouse cortex were followed as they underwent their next cell division in vitro. Numb distribution was predominantly asymmetric during asymmetric cell divisions yielding a beta-tubulin III(-) progenitor and a beta-tubulin III(+) neuronal cell (P/N divisions) and predominantly symmetric during divisions producing two neurons (N/N divisions). Cells from the numb knockout mouse underwent significantly fewer asymmetric P/N divisions compared to wild type, indicating a causal role for Numb. When progenitor cells derived from early (E10) cortex undergo P/N divisions, both daughters express the progenitor marker Nestin, indicating their immature state, and Numb segregates into the P or N daughter with similar frequency. In contrast, when progenitor cells derived from later E13 cortex (during active neurogenesis in vivo) undergo P/N divisions they produce a Nestin(+) progenitor and a Nestin(-) neuronal daughter, and Numb segregates preferentially into the neuronal daughter. Thus during mouse cortical neurogenesis, as in Drosophila neurogenesis, asymmetric segregation of Numb could inhibit Notch activity in one daughter to induce neuronal differentiation. At terminal divisions generating two neurons, Numb was symmetrically distributed in approximately 80% of pairs and asymmetrically in 20%. We found a significant association between Numb distribution and morphology: most sisters of neuron pairs with symmetric Numb were similar and most with asymmetric Numb were different. Developing cortical neurons with Numb had longer processes than those without. Numb is expressed by neuroblasts and stem cells and can be asymmetrically segregated by both. These data indicate Numb has an important role in generating asymmetric cell divisions and diverse cell fates during mouse cortical development.     

Asymmetric production of surface-dividing and non-surface-dividing cortical progenitor cells

Mature neocortical layers all derive from the cortical plate (CP), a transient zone in the dorsal telencephalon into which young neurons are continuously delivered. To understand cytogenetic and histogenetic events that trigger the emergence of the CP, we have used a slice culture technique. Most divisions at the ventricular surface generated paired cycling daughters (P/P divisions) and the majority of the P/P divisions were asymmetric in daughter cell behavior; they frequently sent one daughter cell to a non-surface (NS) position, the subventricular zone (SVZ), within a single cell-cycle length while keeping the other mitotic daughter for division at the surface. The NS-dividing cells were mostly Hu+ and their daughters were also Hu+, suggesting their commitment to the neuronal lineage and supply of early neurons at a position much closer to their destiny than from the ventricular surface. The release of a cycling daughter cell to SVZ was achieved by collapse of the ventricular process of the cell, followed by its NS division. Neurogenin2 (Ngn2) was immunohistochemically detected in a certain cycling population during G1 phase and was further restricted during G2-M phases to the SVZ-directed population. Its retroviral introduction converted surface divisions to NS divisions. The asymmetric P/P division may therefore contribute to efficient neuron/progenitor segregation required for CP initiation through cell cycle-dependent and lineage-restricted expression of Ngn2.     

Non‐equivalence in old‐ and new‐flagellum daughter cells of a proliferative division in Trypanosoma brucei

Differentiation of Trypanosoma brucei, a flagellated protozoan parasite, between life cycle stages typically occurs through an asymmetric cell division process, producing two morphologically distinct daughter cells. Conversely, proliferative cell divisions produce two daughter cells, which look similar but are not identical. To examine in detail differences between the daughter cells of a proliferative division of procyclic T. brucei we used the recently identified constituents of the flagella connector. These segregate asymmetrically during cytokinesis allowing the new‐flagellum and the old‐flagellum daughters to be distinguished. We discovered that there are distinct morphological differences between the two daughters, with the new‐flagellum daughter in particular re‐modelling rapidly and extensively in early G1. This re‐modelling process involves an increase in cell body, flagellum and flagellum attachment zone length and is accompanied by architectural changes to the anterior cell end. The old‐flagellum daughter undergoes a different G1 re‐modelling, however, despite this there was no difference in G1 duration of their respective cell cycles. This work demonstrates that the two daughters of a proliferative division of T. brucei are non‐equivalent and enables more refined morphological analysis of mutant phenotypes. We suggest all proliferative divisions in T. brucei and related organisms will involve non‐equivalence.     

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.     

Sir2-dependent daughter-to-mother transport of the damaged proteins in yeast is required to prevent high stress sensitivity of the daughters

The budding yeast Saccharomyces cerevisiae actively transports adverse factors (e.g. oxidized proteins) from the daughter to mother cells. The transport is believed to ensure that the daughters are born "young", thus preventing clonal senescence. Is this the only reason for the existence of such transport? We subjected yeast cells to various stress conditions and compared survival of mother and daughter cells. It was found that replicative age-dependent mortality under our experimental stress conditions was U-shaped: the resistance of both virgin daughters and old mother cells (more than three bud scars) was lower compared to the young mothers. SIR2 mutants were shown to fail to maintain the mother-daughter asymmetry. We showed that sir2 knockout affects the relative stress resistance in favor of the mothers. Thus, daughter cells are more vulnerable to a variety of stresses than the young mothers, and Sir2-dependent transport of the adverse factors acts to equalize the resistance.     

Yeast mother cell-specific ageing, genetic (in)stability, and the somatic mutation theory of ageing

Yeast mother cell-specific ageing is characterized by a limited capacity to produce daughter cells. The replicative lifespan is determined by the number of cell cycles a mother cell has undergone, not by calendar time, and in a population of cells its distribution follows the Gompertz law. Daughter cells reset their clock to zero and enjoy the full lifespan characteristic for the strain. This kind of replicative ageing of a cell population based on asymmetric cell divisions is investigated as a model for the ageing of a stem cell population in higher organisms. The simple fact that the daughter cells can reset their clock to zero precludes the accumulation of chromosomal mutations as the cause of ageing, because semiconservative replication would lead to the same mutations in the daughters. However, nature is more complicated than that because, (i) the very last daughters of old mothers do not reset the clock; and (ii) mutations in mitochondrial DNA could play a role in ageing due to the large copy number in the cell and a possible asymmetric distribution of damaged mitochondrial DNA between mother and daughter cell. Investigation of the loss of heterozygosity in diploid cells at the end of their mother cell-specific lifespan has shown that genomic rearrangements do occur in old mother cells. However, it is not clear if this kind of genomic instability is causative for the ageing process. Damaged material other than DNA, for instance misfolded, oxidized or otherwise damaged proteins, seem to play a major role in ageing, depending on the balance between production and removal through various repair processes, for instance several kinds of proteolysis and autophagy. We are reviewing here the evidence for genetic change and its causality in the mother cell-specific ageing process of yeast.     

Cell cycle phases in the unequal mother/daughter cell cycles of Saccharomyces cerevisiae.

During cell division in the yeast Saccharomyces cerevisiae mother cells produce buds (daughter cells) which are smaller and have longer cell cycles. We performed experiments to compare the lengths of cell cycle phases in mothers and daughters. As anticipated from earlier indirect observations, the longer cell cycle time of daughter cells is accounted for by a longer G1 interval. The S-phase and the G2-phase are of the same duration in mother and daughter cells. An analysis of five isogenic strains shows that cell cycle phase lengths are independent of cell ploidy and mating type.     

Evidence for the involvement of a cytoplasmic factor in the aging of the yeast Saccharomyces cerevisiae.

The life spans of individual Saccharomyces cerevisiae cells were determined microscopically by counting the number of buds produced by each cell to provide a measure of the number of cell generations (age) before death. As the cells aged, their generation times increased five- to sixfold. The generation times of daughter cells were virtually identical to those of their mothers throughout the life spans of the mothers. However, within two to three cell divisions after the daughters were detached from their mothers by micromanipulation, their generation times reverted to that characteristic of their own age. Recovery from the mother cell effect was also observed when the daughters were left attached to their mothers. The results suggest that senescence, as manifested by the increase in generation time, is a phenotypically dominant feature in yeast cells and that it is determined by a diffusible cytoplasmic factor(s) that undergoes turnover. This factor(s) appeared to be transmitted by a cell not only to its daughter, but also indirectly to its granddaughter. In separate studies, it was determined that the induced deposition of chitin, the major component of the bud scar, in the yeast cell wall had no appreciable effect on life span. We raise the possibility that the cytoplasmic factor(s) that appears to mediate the "senescent phenotype" is a major determinant of yeast life span. This factor(s) may be the product of age-specific gene expression.     

Ephrin-Eph signalling drives the asymmetric division of notochord/neural precursors in Ciona embryos

Asymmetric cell divisions produce two sibling cells with distinct fates, providing an important means of generating cell diversity in developing embryos. Many examples of such cell divisions have been described, but so far only a limited number of the underlying mechanisms have been elucidated. Here, we have uncovered a novel mechanism controlling an asymmetric cell division in the ascidian embryo. This division produces one notochord and one neural precursor. Differential activation of extracellular-signal-regulated kinase (ERK) between the sibling cells determines their distinct fates, with ERK activation promoting notochord fate. We first demonstrate that the segregation of notochord and neural fates is an autonomous property of the mother cell and that the mother cell acquires this functional polarity via interactions with neighbouring ectoderm precursors. We show that these cellular interactions are mediated by the ephrin-Eph signalling system, previously implicated in controlling cell movement and adhesion. Disruption of contacts with the signalling cells or inhibition of the ephrin-Eph signal results in the symmetric division of the mother cell, generating two notochord precursors. Finally, we demonstrate that the ephrin-Eph signal acts via attenuation of ERK activation in the neural-fated daughter cell. We propose a model whereby directional ephrin-Eph signals functionally polarise the notochord/neural mother cell, leading to asymmetric modulation of the FGF-Ras-ERK pathway between the daughter cells and, thus, to their differential fate specification.     

Apical complex genes control mitotic spindle geometry and relative size of daughter cells in Drosophila neuroblast and pI asymmetric divisions

Drosophila neuroblast asymmetric divisions generate two daughters of unequal size and fate. A complex of apically localized molecules mediates basal localization of cell fate determinants and apicobasal orientation of the mitotic spindle, but how daughter cell size is controlled remains unclear. Here we show that mitotic spindle geometry and unequal daughter cell size are controlled by two parallel pathways (Bazooka/DaPKC and Pins/G alpha i) within the apical complex. While the localized activity of either pathway alone is sufficient to mediate the generation of an asymmetric mitotic spindle and unequal size neuroblast daughters, loss of both pathways results in symmetric divisions. In sensory organ precursors, Bazooka/DaPKC and Pins/G alpha i localize to opposite sides of the cortex and function in opposition to generate a symmetric spindle.     

Asymmetric cell divisions in flowering plants - one mother, "two-many" daughters

Plant development shows a fascinating range of asymmetric cell divisions. Over the years, however, cellular differentiation has been interpreted mostly in terms of a mother cell dividing mitotically to produce two daughter cells of different fates. This popular view has masked the significance of an entirely different cell fate specification pathway, where the mother cell first becomes a coenocyte and then cellularizes to simultaneously produce more than two specialized daughter cells. The "one mother - two different daughters" pathways rely on spindle-assisted mechanisms, such as translocation of the nucleus/spindle to a specific cellular site and orientation of the spindle, which are coordinated with cell-specific allocation of cell fate determinants and cytokinesis. By contrast, during "coenocyte-cellularization" pathways, the spindle-assisted mechanisms are irrelevant since cell fate specification emerges only after the nuclear divisions are complete, and the number of specialized daughter cells produced depends on the developmental context. The key events, such as the formation of a coenocyte and migration of the nuclei to specific cellular locations, are coordinated with cellularization by unique types of cell wall formation. Both one mother - two different daughters and the coenocyte-cellularization pathways are used by higher plants in precise spatial and time windows during development. In both the pathways, epigenetic regulation of gene expression is crucial not only for cell fate specification but also for its maintenance through cell lineage. In this review, the focus is on the coenocyte-cellularization pathways in the context of our current understanding of the asymmetric cell divisions. Instances where cell differentiation does not involve an asymmetric division are also discussed to provide a comprehensive account of cell differentiation.     

C. elegans HAM-1 positions the cleavage plane and regulates apoptosis in asymmetric neuroblast divisions

Asymmetric cell division occurs when a mother cell divides to generate two distinct daughter cells, a process that promotes the generation of cellular diversity in metazoans. During Caenorhabditis elegans development, the asymmetric divisions of neural progenitors generate neurons, neural support cells and apoptotic cells. C. elegans HAM-1 is an asymmetrically distributed cortical protein that regulates several of these asymmetric neuroblast divisions. Here, we show that HAM-1 is a novel protein and define residues important for HAM-1 function and distribution to the cell cortex. Our phenotypic analysis of ham-1 mutant embryos suggests that HAM-1 controls only neuroblast divisions that produce apoptotic cells. Moreover, ham-1 mutant embryos contain many unusually large cell-death corpses. An investigation of this corpse phenotype revealed that it results from a reversal of neuroblast polarity. A misplacement of the neuroblast cleavage plane generates daughter cells of abnormal size, with the apoptotic daughters larger than normal. Thus, HAM-1 regulates the position of the cleavage plane, apoptosis and mitotic potential in C. elegans asymmetric cell divisions.     

Differences of asymmetrical division between the pseudomycelial and yeast forms of Candida albicans and their effect on multiplication

Unlike asymmetrical division of budding yeast, the daughter cell in the pseudomycelial form of Candida albicans at division was nearly equal in size to the mother cell, it had a larger amount of protein, RNA and active protoplasm (cell size minus vacuolar volume) than the mother cell, and it budded earlier than the mother cell. Results presented here suggest that the cell size control over bud initiation found in budding yeasts is also applicable to the pseudomycelial cells of C. albicans if vacuolar volume is omitted from cell size.     

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.     

Gender Differences in Associations of Eating Pathology between Mothers and Their Adolescent Offspring

Objective: To study the association of eating pathology between mothers and their adolescent offspring in a population sample. Research Methods and Procedures: The participants were 481 women (mean age, 47 ± SD 5 years; BMI, 25 ± 4 kg/m²) and their 481 adolescent children 16 to 17 years old (BMI, 21 ± 3 kg/m²) of the Stockholm Weight Development Study. Assessment methods were the Three‐Factor Eating Questionnaire Revised 18 and the Eating Disorder Inventory 2. Results: A higher body weight was most related to cognitive restraint for adolescents and to emotional eating for adult women. A mother‐daughter link could be identified for eating pathology, with the strongest link found for emotional eating. No mother‐son link could be identified. Age subgroup analyses revealed a stronger mother‐daughter link for body attitudes in younger mothers and for cognitive restraint in older mothers. Discussion: Gender differences revealed that eating pathology was shared by mothers and daughters but not by mothers and sons. A psychological strategy such as eating as a response to negative emotions was most interrelated between mothers and daughters. Younger mothers shared more attitudes toward the body with their daughters, whereas older mothers shared more restrictive eating behaviors with their daughters. The mother‐daughter links found may be due to gender‐specific genetic and psychological family transmission and gender‐specific environmental influences. The sons’ eating behaviors seem to be more independent and would be formed by other factors than for the girls.     

Cortical domains and the mechanisms of asymmetric cell division

Asymmetric cell divisions are central to the generation of cell-fate diversity because factors that are present in a mother cell and distributed unequally at cell division can generate distinct daughters. The process o f asymmetric cell division can be described as consisting of three steps: setting up an asymmetric cue in the mother cell, localizing factors with respect to this cue, and positioning the plane o f cell division so that localized factors are partitioned asymmetrically between daughters. This review describes how specialized cortical domains play a key role in each of these steps and discusses our current understanding of the molecular nature o f cortical domains and the mechanisms by which they may orchestrate asymmetric cell divisions.