Mitotic disturbance associated with mosaic aneuploidies

Summary The association of various unsystematic aneuploidies with premature centromere division (PCD) was observed in a patient with conspicuous clinical features and combined immunodeficiency. Trisomies and monosomies of almost all autosomes and gonosomal aberrations were found separately or in combination in a majority of the proband's lymphocytes and fibroblasts. The chromosome number varied from 44 to 50. A high proportion of the metaphases showed PCD or had the appearance of C-anaphases. These findings probably represent a new mutant affecting mitosis and causing mosaic aneuploidies.     

A mechanism of x chromosome aneuploidy in lymphocytes of aging women.

One and sometimes both X chromosomes in cultured lyphocytes of women donors showed division of the centromere when the centromeres of other chromosomes were entire. This premature centromere division (PCD) was associated with evidence of non-disjunction of the X chromosome. On average, 2% of metaphases from 32 women donors showed PCD, but the incidence was 4 times greater in women over 59 years of age than in women under 40 years. Increased X chromosome aneuploidy was associated with the higher frequency of PCD in cultured lymphocytes from older women. PCD of the X chromosome is considered to be the mechanism of non-disjunction causing the previously described aneuploidy in cultured lymphocytes of aging women.     

C-anaphases: a mitotic variant.

In order to assess the frequency of C-anaphases in colchicine-arrested lymphocyte cultures, the authors studied 100 patients classified in four groups: spontaneous abortion (n = 17), subfertility (n = 12), aneuploidy (n = 18) and miscellaneous (n = 53). At least 300 mitotic G-banded cells were scored by individual. In 12 individuals no C-anaphases were observed; in 87 individuals the range was 1-7 with a mode of 2 and a mean of 2.14; the remaining individual had 19 C-anaphases in 330 cells (5.7%). The statistical analysis did not show significant differences between the groups (p > 0.05). These data along with previous studies indicate that normally most individuals have < or = 3% of C-anaphases in habitual lymphocyte cultures. Moreover, there exists an autosomal dominant form in which individuals with the trait have > 5% of the cells with such mitotic configurations. We conclude that both the low frequency common and the high frequency familiar forms are mitotic variants without pathological significance.     

Premature centromere division of the X chromosome in neurons in Alzheimer's disease

Premature centromere division (PCD) represents a loss of control over the sequential separation and segregation of chromosome centromeres. Although first described in aging women, PCD on the X chromosome (PCD,X) is markedly elevated in peripheral blood lymphocytes of individuals suffering from Alzheimer disease (AD). The present study evaluated PCD,X, using a fluorescent in situ hybridization method, in interphase nuclei of frontal cerebral cortex neurons from sporadic AD patients and age-matched controls. The average frequency of PCD,X in AD patients (8.60 ± 1.20%) was almost three times higher ( p  < 0.01) than in the control group (2.96 ± 1.20). However, consistent with previous studies, no mitotic cells were found in neurons in either AD or control brain, suggesting an intrinsic inability of post-mitotic neurons to divide. In view of the fact that it has been well-documented that neurons in AD can re-enter into the cell division cycle, the findings presented here of increased PCD advance the hypothesis that deregulation of the cell cycle may contribute to neuronal degeneration and subsequent cognitive deficits in AD.     

The frequency of induced premature centromere division in human populations occupationally exposed to genotoxic chemicals

Premature (early) centromere division (PCD, i.e., the separation of centromeres during the prometaphase/metaphase of the mitotic cycle) seems to be a possible manifestation of chromosome instability in human chromosome-breakage syndromes. Chromosome instability also frequently occurs in the peripheral blood lymphocytes (PBL) of humans occupationally exposed to clastogenic agents, and is considered an etiologic factor of neoplastic diseases. In order to investigate the importance of PCD in cancer risk assessment, we studied the frequency of PCDs in PBL of 400 Hungarian subjects. The various groups comprised 188 control donors and 212 subjects occupationally exposed to different genotoxic chemicals, such as acrylonitrile (ACN) and/or dimethylformamide (DMF), benzene, cytostatic drugs, ethylene oxide (ETO), mixed exposure in the rubber industry, mixed organic solvents including CCl4, hot oil-mist, bitumen, and polychlorinated biphenyls (PCB). Data were compared with chromosomal aberration frequencies determined in the same samples. PCD yields are significantly higher in populations exposed to mixed chemicals, crude oil and cytostatic drugs, compared with controls. PCDs involving more than three chromosomes are also more frequent in ETO- and oil mist-exposed groups than in the others. The results indicate that the induction of PCDs is neither incidental nor artificial. As a consequence, we suggest that PCD can be developed into a new, exposure-related cytogenetic biomarker for a more adequate occupational cancer risk assessment. A further, follow-up epidemiological and cytogenetic investigation of PCD is in progress.     

Colchicine resistance in human cell lines. Pleiotropic phenotype and decreased membrane permeability

Summary Colchicine-resistant human cells were initially observed in patients exhibiting C-anaphases or tetraploidy in lymphocyte cultures. Cell lines established from these patients displayed cross-resistances to daunomycin, emetine, vinblastine, and vincristine and collateral sensitivity to Xylocaine, showing a pleiotropic phenotype similar to that described in permeability mutants in CHO cells. ³H colchicine uptake and binding assays confirmed a decreased permeability to the drug.     

玉米（Zea mays L.）过氧化物酶编码基因px和冷调控蛋白编码基因cld的物理定位

过氧化物酶是植物抗病过程中的关键酶,也参与植物抗低温胁迫以及一些正常的植物生长发育生理过程。冷调控蛋白是植物抗低、高温的重要蛋白。近年的研究表明植物抗病、某些正常生长发育过程。冷调控蛋白是植物细胞程序性死亡有关。以玉米自效系黄早四为材料,采用生物素标记,利用原位杂效技术绎玉米中过氧化物酶和冷调控蛋白编码基因ｐｘ与ｃｌｄ进行了原位杂效定位,ＤＡＢ和荧光检测得到了一致的结果。在２号和７号染色体长臂同时     

Classes of programmed cell death in plants, compared to those in animals

Relatively little is known about programmed cell death (PCD) in plants. It is nonetheless suggested here that tonoplast rupture and the subsequent rapid destruction of the cytoplasm can distinguish two large PCD classes. One class, which is here called 'autolytic', shows this feature, whilst the second class (called 'non-autolytic') can include tonoplast rupture but does not show the rapid cytoplasm clearance. Examples of the 'autolytic' PCD class mainly occur during normal plant development and after mild abiotic stress. The 'non-autolytic' PCD class is mainly found during PCD that is due to plant-pathogen interactions. Three categories of PCD are currently recognized in animals: apoptosis, autophagy, and necrosis. An attempt is made to reconcile the recognized plant PCD classes with these groups. Apoptosis is apparently absent in plants. Autophagic PCD in animals is defined as being accompanied by an increase in the number of autophagosomes, autolysosomes, and small lytic vacuoles produced by autolysosomes. When very strictly adhering to this definition, there is no (proof for) autophagic PCD in plants. Upon a slightly more lenient definition, however, the 'autolytic' class of plant PCD can be merged with the autophagic PCD type in animal cells. The 'non-autolytic' class of plant PCD, as defined here, can be merged with necrotic PCD in animals.     

Sequential changes of programmed cell death in developing fetal mouse limbs and its possible roles in limb morphogenesis

Apoptotic cell death in the developing limb of mouse fetuses was examined sequentially on days 11–15 of gestation by means of Nile blue (NB) sulfate staining with special reference to its relation to limb morphogenesis. With some exceptions, programmed cell death (PCD) in the hand and foot was observed in the mesenchyme but not in the surface ectoderm. We found that during digital formation PCD begins at the proximal portion of the interdigital mesenchyme and subsequently expands distally. Therefore, the initial PCD that occurs in the interdigital zones may determine the proximal ends of digital separation and also contribute to the demarcation between the palm (sole) and digits (toes). During digital separation, the areas of PCD in the interdigital zones were found to become larger and expand distally on day 13, which may be necessary for the separation of digits and for determining the interdigital area to disappear. PCD in presumptive phalangeal joints was also found to proceed from proximal to more distal joints. The PCD in presumptive joints may be required for the separation of phalanges and metacarpal (metatarsal) bones and for the formation of joint cavities. In addition, intense PCD was observed in the radial (tibial) and ulnar (fibular) margins of the hand and foot plates for 4–5 days. Such PCD at marginal areas seems to prevent the formation of supernumerary digits (preaxial and postaxial polydactyly) and other digital malformations. Therefore, the timing when PCD commences and ends, the sites where PCD occurs, and the intensity, duration, and proximo-distal progress of PCD appear to be genetically determined, and the elimination of unnecessary cells by PCD may be essential for normal limb morphogenesis. The present findings also suggest that the normal progress of PCD in the hand and foot plates of rodent fetuses may prevent the formation of some limb malformations such as webbing fusion of digits, polydactyly, or cleft hand/foot. © 1996 Wiley-Liss, Inc.     

Programmed cell death of larval tissues induced by juvenile hormone in the bamboo borer, Omphisa fuscidentalis

Programmed cell death (PCD) plays a critical role during animal development through the destruction of unneeded cells and tissues. In some insects, the prothoracic glands (PGs) and anterior silk glands (ASGs) are larval-specific tissues that are normally eliminated by PCD after pupation. Previous studies report that juvenile hormone analog (JHA) terminates the larval diapause of Omphisa fuscidentalis by increasing the hemolymph ecdysteroids that trigger PCD. Because JHA may indirectly induce the PCD of the PGs and ASGs of Omphisa diapausing larvae, the effects of JHA on the induction of PCD were determined. The application of 1μg JHA induced PCD in the PGs and ASGs of larvae identified as stage G0 (prior to pupation). The injection of 1μg 20E triggered the PCD of the ASGs when the larvae expressed a G0-G1 morphology, whereas PCD occurred in the PGs on day 1 post-injection. Histological studies revealed similar patterns of morphological changes during the PG and ASG PCD in the JHA- and 20E-treated larvae. Furthermore, to confirm that PCD was induced by a high ecdysteroid level that increases after JHA application, the expression profiles of EcR-A and EcR-B1 in the PGs and ASGs from the JHA-treated larvae were examined, and the results showed that the expression levels of EcR-A and EcR-B1 mRNA increased during the G0 stage. These results suggest that JHA may be involved in PCD by increasing the ecdysteroid titer, leading to termination of the larval diapause period in Omphisa fuscidentalis.     

The murine TRAIL receptor signals caspase-independent cell death through ceramide

Death receptors such as the 55 kDa tumor necrosis factor (TNF) receptor (TNF-R55) or Fas can initiate both apoptotic (caspase-dependent) and caspase-independent routes to programmed cell death (PCD). Here, we demonstrate for the first time that the single murine receptor for (TNF)-related apoptosis-inducing ligand (mTRAIL-R2) can induce a caspase-independent form of PCD with necrosis-like features in addition to apoptosis. Analysis of morphological and cellular features of caspase-independent PCD in response to TRAIL and TNF suggests that mTRAIL-R2 and TNF-R55 elicit caspase-independent PCD through similar pathways, although without participation of cathepsins. Cells overexpressing acid ceramidase (AC), an enzyme that metabolizes the sphingolipid ceramide, show enhanced survival from TRAIL-induced caspase-independent PCD but not from apoptosis, implicating a function of ceramide as a key mediator in caspase-independent PCD (but not apoptosis) induced by mTRAIL-R2. In concert with the enhanced resistance of AC-overexpressing cells against caspase-independent PCD induced by TNF, our results suggest that ceramide acts as a common mediator of caspase-independent PCD caused by death receptors such as mTRAIL-R2 and TNF-R55.     

Acrocentric centromere organization within the chromocenter of the human sperm nucleus.

It has recently been reported that in human sperm cells, the centromeres are clustered in a chromocenter in the interior region of the nucleus. The aim of the present study was to determine the intra-chromocenter organization of the five centromeres of the acrocentric chromosomes responsible for the biosynthesis of rRNA. The acrocentric centromeres were labeled by fluorescence in situ hybridization (FISH) after mild decondensation of the sperm nuclei to preserve the tail structure. The tail was used as a topographical marker for the orientation of the nucleus. The following results were obtained: (a) the association among the five centromeres was higher than expected from random distribution; (b) all the centromeres observed were randomly located within the chromocenter, occupying about 87% of the total area of the internal nucleus; (c) a major subpopulation of centromeres was located in a preferred area occupying 8.3% of the total nuclear area, with a peak 0.6 microm on the lateral axis and 1.0 microm on the apical side of the longitudinal axis; and (d) The dispersion of the centromeres was not influenced by the degree of the nuclear decondensation. We conclude that in human sperm nuclei, the acrocentric centromeres are organized within a nonlocalized structural element in the chromocenter. The chromocenter can range from an expanded size of 87% of the whole nucleus to a preferred size of 8.3% independent of the degree of nuclear decondensation. These findings have important implications for nuclear function (rRNA) that is not directly related to sperm cell function or early embryo development.     

Formation of primary constriction and heterochromatin in mouse does not require minor satellite DNA.

Whereas the major satellite fraction in mouse extends its domain from the centromere to the distal end of the pericentric heterochromatin, the minor satellite DNA is present specifically in the centromere or primary constriction. We hybridized the biotinylated minor satellite sequence to L929 cells of mouse origin. The sequence hybridized to all chromosomes. Whereas hybridization was detected on all active centromeres, the inactive centromeres in certain dicentrics did not show any signal. This satellite, however, was detected in all inactive centromeres in a heptacentric chromosome. The intensity of fluorescence on the inactive centromeres of the heptacentric was similar to that present on the active centromeres. Several heterochromatin blocks, which were not associated with any centromere, were also found to lack hybridization with the minor satellite. The inactive centromeres, whether carrying the minor satellite DNA fraction or not, generally do not react with the antikinetochore antibodies present in the scleroderma serum. These studies are interpreted to show that (1) the primary constriction in mouse can be formed without the participation of minor satellite, (2) heterochromatin in mouse may constitute without this fraction, (3) the major and minor satellite may not be interspersed but are joined at some defined boundary, and (4) the binding of CENP-B does not depend upon the quantity of minor satellite or the number of CENP boxes present in the inactive centromeres.     

The myelin proteolipid protein gene modulates apoptosis in neural and non-neural tissues

Mutations of the myelin proteolipid protein gene (Plp) are associated with excessive programmed cell death (PCD) of oligodendrocytes. We show for the first time that PLP is a molecule ubiquitously expressed in non-neural tissues during normal development, and that the level of native PLP modulates the level of PCD. We analyze three non-neural tissues, and show that native PLP is expressed in trophoblasts, spermatogonia, and cells of interdigital webbing. The non-neural cells that express high levels of native PLP also undergo PCD. The level of PLP expression modulates the level of PCD because mice that overexpress native PLP have increased PCD and mice deficient in PLP have decreased PCD. We show that overexpression of native PLP causes a dramatic acidification of extracellular fluid that, in turn, causes increased PCD. These studies show that the level of native PLP modulates the amount of PCD during normal development via a pH-dependent mechanism.     

Maize Centromere Structure and Evolution: Sequence Analysis of Centromeres 2 and 5 Reveals Dynamic Loci Shaped Primarily by Retrotransposons

We describe a comprehensive and general approach for mapping centromeres and present a detailed characterization of two maize centromeres. Centromeres are difficult to map and analyze because they consist primarily of repetitive DNA sequences, which in maize are the tandem satellite repeat CentC and interspersed centromeric retrotransposons of maize (CRM). Centromeres are defined epigenetically by the centromeric histone H3 variant, CENH3. Using novel markers derived from centromere repeats, we have mapped all ten centromeres onto the physical and genetic maps of maize. We were able to completely traverse centromeres 2 and 5, confirm physical maps by fluorescence in situ hybridization (FISH), and delineate their functional regions by chromatin immunoprecipitation (ChIP) with anti-CENH3 antibody followed by pyrosequencing. These two centromeres differ substantially in size, apparent CENH3 density, and arrangement of centromeric repeats; and they are larger than the rice centromeres characterized to date. Furthermore, centromere 5 consists of two distinct CENH3 domains that are separated by several megabases. Succession of centromere repeat classes is evidenced by the fact that elements belonging to the recently active recombinant subgroups of CRM1 colonize the present day centromeres, while elements of the ancestral subgroups are also found in the flanking regions. Using abundant CRM and non-CRM retrotransposons that inserted in and near these two centromeres to create a historical record of centromere location, we show that maize centromeres are fluid genomic regions whose borders are heavily influenced by the interplay of retrotransposons and epigenetic marks. Furthermore, we propose that CRMs may be involved in removal of centromeric DNA (specifically CentC), invasion of centromeres by non-CRM retrotransposons, and local repositioning of the CENH3.     

MPK6, sphinganine and the LCB2a gene from serine palmitoyltransferase are required in the signaling pathway that mediates cell death induced by long chain bases in Arabidopsis

Long chain bases (LCBs) are sphingolipid intermediates acting as second messengers in programmed cell death (PCD) in plants. Most of the molecular and cellular features of this signaling function remain unknown. We induced PCD conditions in Arabidopsis thaliana seedlings and analyzed LCB accumulation kinetics, cell ultrastructure and phenotypes in serine palmitoyltransferase (spt), mitogen-activated protein kinase (mpk), mitogen-activated protein phosphatase (mkp1) and lcb-hydroxylase (sbh) mutants. The lcb2a-1 mutant was unable to mount an effective PCD in response to fumonisin B1 (FB1), revealing that the LCB2a gene is essential for the induction of PCD. The accumulation kinetics of LCBs in wild-type (WT) and lcb2a-1 plants and reconstitution experiments with sphinganine indicated that this LCB was primarily responsible for PCD elicitation. The resistance of the null mpk6 mutant to manifest PCD on FB1 and sphinganine addition and the failure to show resistance on pathogen infection and MPK6 activation by FB1 and LCBs indicated that MPK6 mediates PCD downstream of LCBs. This work describes MPK6 as a novel transducer in the pathway leading to LCB-induced PCD in Arabidopsis, and reveals that sphinganine and the LCB2a gene are required in a PCD process that operates as one of the more effective strategies used as defense against pathogens in plants.     

Promoter analysis of the Drosophila melanogaster gene encoding the pterin 4alpha-carbinolamine dehydratase.

Pterin-4alpha-carbinolamine dehydratase (PCD) is a key enzyme in the regeneration pathway of tetrahydrobiopterin. Previously, we isolated and reported the Drosophila melanogaster gene encoding PCD. In the present study, we isolated and characterized the Drosophila virilis gene encoding PCD. The Drosophila virilis PCD gene has two introns and an open reading frame to encode a protein of 101 amino acids. The amino acid sequence of Drosophila virilis PCD shows a 83% homology to that of the Drosophila melanogaster PCD protein. From the alignment of the nucleotide sequence in the 5'-flanking region of the Drosophila melanogaster and Drosophila virilis PCD genes, we found four conserved sequences. Using a transient transfection assay, we showed that one of the conserved sequences (-127 to approximately -115) is critical for expression, also the minimal promoter region between -127 and +51 is necessary for the efficient expression of Drosophila melanogaster PCD.     

Programmed cell death in plants

The modern concepts of programmed cell death (PCD) in plants are reviewed as compared to PCD (apoptosis) in animals. Special attention is focused on considering the potential mechanisms of implementation of this fundamental biological process and its participants. In particular, the proteolytic enzymes involved in PCD in animals (caspases) and plants (phytaspases) are compared. Emphasis is put on elucidation of both common features and substantial differences of PCD implementation in plants and animals.     

Many ways to exit? Cell death categories in plants

Programmed cell death (PCD) is an integral part of plant development and defence. It occurs at all stages of the life cycle, from fertilization of the ovule to death of the whole plant. Without it, tall trees would probably not be possible and plants would more easily succumb to invading microorganisms. Here, we have attempted to categorize plant PCD in relation to three established morphological types of metazoan cell death: apoptosis, autophagy and non-lysosomal PCD. We conclude that (i) no examples of plant PCD conform to the apoptotic type, (ii) many examples of PCD during plant development agree with the autophagic type, and (iii) that other examples are apparently neither apoptotic nor autophagic.