Premeiotic Interphase Research Articles

Sexual Reproduction in the Fungal Parasite, Zygorhizidium planktonicum

The fungal parasite Zygorhizidium planktonicum (Chytridiales) exhibits a haplontic life cycle characterized by an asexual infection phase and the formation of sexual resting spores following gametangial conjugation. Uninucleate gametangia fuse via an epibiotic conjugation tube formed by the smaller, donor thallus. Plasmogamy entails allogamous and autogamous fusions; however, mechanisms for sexual expression remain unresolved. Karyogamy occurs within the recipient gametangium as documented by a single nucleus following deposition of a thick, multi-layered wall. The zygotic resting spore remains undifferentiated throughout dormancy at 4 C. Upon activation, a single germination pore is formed, and a thin-walled, operculate meiosporangium emerges as an extension of the inner spore wall. The nucleus migrates into the meiosporangium during premeiotic interphase or prophase of meiosis I. Meiotic nuclei are evidenced by synaptonemal complex formation 24–48 h after germination. Nuclear division and meiosis II proceed in rapid succession followed by at least one mitotic division prior to zoosporogenesis. Confirmation of zygotic meiosis in Z. planktonicum further substantiates the prevalence of haplontic life cycles in the Chytridiomycetes. The taxonomic implications of sexual reproduction are reevaluated with particular attention to plasmogamous mechanisms and nuclear cycles.

The effect of gibberellic acid on male fertility in bread wheat

The effect of gibberellic acid (GA3) on male fertility was investigated in wheat. Greenhouse and field experiments were carried out. GA3 induced high levels of male sterility both in gibberellin (GA)-sensitive and GA-insensitive genotypes. The optimum concentration was 2000 ppm of GA3 when applied at successive sprays. The critical period for GA3 treatment, in the general sense, extended from glume differentiation to premeiotic interphase in the oldest florets of the spike, though differences were found between GA-sensitive and GA-insensitive genotypes in the extension of this period. The effect of GA3 on plant height was also studied. The potential use of GA3 as a chemical hybridizing agent in wheat breeding is discussed.

Radioactive response in primary mouse spermatocytes revealed by analysis of synaptonemal complexes

This study was aimed at estimating the damage to the synaptonemal complex (SC) of male mice caused by radiation exposure at the ultimate pre-meiotic interphase. Four experimental groups were fromed: group 1 was an untreated control, group 2 received a low dose (50 mGy) of radiations, group 3 a high dose (3 Gy) of radiation, group 4 was first treated with a low dose (50 mGy) and 4 h later with a high dose (4 Gy). Mice were linked 4 days after the treatment. Early pachytene cells were selected for the electron microscopic analysis of SCs. These cells were supposed to be at pre-meiotic interphase at the time of irradiation. Treatment with a low dose of radiation did not produce any substantial increase in the frequency of SC aberrations, while exposure to the high dose resulted in various types of damage. Group 4 demonstrated significantly lower frequencies of axial breaks/fragments and multiaxial configurations than group 3. At the same time, these two groups did not differ in the frequencies of inter and intrachromosomal exchanges such as translocation, inversions, and deletions. We suppose that the number of breaks produced by a high dose of radiation was the same in the both groups. However, a proportion of the breaks that remained unrepaired until pachytene and were expressed as gaps, fragments, and multiaxial configurations was reduced by pretreatment with the low dose of radiation.

Specific developmental stages of gametogenesis for radiosensitivity and mutagenesis in rice

Developmental stages during gametogenesis of rice were histologically examined in the period from differentiation of reproductive organs to anthesis. Plants were exposed to acute X-rays of 20 Gy. Radiosensitivity and mutation frequency were investigated in relation to the developmental stages of reproductive organs. The most radiosensitive stage, as measured by reduction of the M1 pollen-and seed-fertilities, was the last premeiotic interphase. Mutations induced at different developmental stages were scored in M3 strains. Sterility mutants and short-culm mutants were most frequently observed. Grain shape, panicle morphology, heading-date and endosperm character mutants were induced at a relatively low frequency. The overall mutation frequency varied with the developmental stage at the time of irradiation. The highest overall mutation frequency was observed when radiation was applied 10 days before anthesis, the late tetrad stage of microspores. Radiation exposure of florets at the late tetrad stage was found to be a more efficient method of inducing a large number of mutations than radiations applied to seeds and fertilized egg cells.

The role of the ameiotic1 gene in the initiation of meiosis and in subsequent meiotic events in maize.

Understanding the initiation of meiosis and the relationship of this event with other key cytogenetic processes are major goals in studying the genetic control of meiosis in higher plants. Our genetic and structural analysis of two mutant alleles of the ameiotic1 gene (am1 and am1-praI) suggest that this locus plays an essential role in the initiation of meiosis in maize. The product of the ameiotic1 gene affects an earlier stage in the meiotic sequence than any other known gene in maize and is important for the irreversible commitment of cells to meiosis and for crucial events marking the passage from premeiotic interphase into prophase I including chromosome synapsis. It appears that the period of ameiotic1 gene function in meiosis at a minimum covers the interval from some point during premeiotic interphase until the early zygotene stage of meiosis. To study the interaction of genes in the progression of meiosis, several double meiotic mutants were constructed. In these double mutants (i) the ameiotic1 mutant allele was brought together with the meiotic mutation (afd1) responsible for the fixation of centromeres in meiosis; and with the mutant alleles of the three meiotic genes that control homologous chromosome segregation (dv1, ms43 and ms28), which impair microtubule organizing center organization, the orientation of the spindle fiber apparatus, and the depolymerization of spindle filaments after the first meiotic division, respectively; (ii) the afd1 mutation was combined with two mutations (dsy1 and as1) affecting homologous pairing; (iii) the ms43 mutation was combined with the as1, the ms28 and the dv1 mutations; and (iv) the ms28 mutation was combined with the dv1 mutation and the ms4 (polymitotic1) mutations. An analysis of gene interaction in the double mutants led us to conclude that the ameiotic1 gene is epistatic over the afd1, the dv1, the ms43 and the ms28 genes but the significance of this relationship requires further analysis. The afd gene appears to function from premeiotic interphase throughout the first meiotic division, but it is likely that its function begins after the start of the ameiotic1 gene expression. The afd1 gene is epistatic over the two synaptic mutations dsy1 and as1 and also over the dv1 mutation. The new ameiotic*-485 and leptotene arrest*-487 mutations isolated from an active Robertson's Mutator stocks take part in the control of the initiation of meiosis.

Diplosporous embryo-sac formation and the degree of diplospory inAllium tuberosum

To obtain information on the dynamic and quantitative aspects of displospory in a facultative apomict,Allium tuberosum (2n = 32), we cytologically observed the entire course of female meiosis and embryosac development in cv ‘Tender-Pole’ by the clearing-staining-squash method. A time-table of its course was constructed in terms of bud length and days pre-anthesis. Chromosome threads were observed in pairs at the earliest stage of meiosis, after which 32 autobivalents developed. Thus, endoreduplication must have occurred during premeiotic interphase or at the onset of female meiosis. Such endoreduplicational meiosis also occurred on the male side, though the frequency (3.9%) was much lower than that on the female side (80%). The degree of diplospory calculated as the percentage of reduplicated embryo-sac mother cells varied to some extent among six cultivars: ‘Huhehaote’ and ‘Manchuria’ showed the highest degree of diplospory (98%), and ‘Kaohsiung’ the lowest (76%).

Antibodies directed against a meiosis-specific, chromatin-associated protein identify conserved meiotic epitopes

The molecular mechanisms by which meiotic events are regulated are at present unknown. To approach this problem, we have exploited the natural synchrony of Lilium meiocytes to compare the nuclear protein profiles of a variety of stages of meiosis. This approach has facilitated the identification of a number of nuclear proteins that appear and disappear in a stage-specific fashion. Here we report the presence of an abundant nuclear protein that first appears during premeiotic interphase, a period during which the irreversible commitment to meiosis occurs. Antibodies directed against this protein demonstrate its meiosis specificity as well as conservation of the epitope(s) in both mono- and dicotyledonous plant species. Chromatin fractionation studies indicate that this protein, which we have termed meiotin-1, is associated with strings of nucleosomes. Implications for meiotic chromatin packaging and chromosome structure are discussed.

Characterization of the Arrest in Anther Development Associated with Gibberellin Deficiency of the gib-1 Mutant of Tomato

The role of gibberellins in flower bud development was investigated by studying the gib-1 mutant of tomato, Lycopersicon esculentum. This gibberellin-deficient mutant initiates flower buds, but floral development is not completed unless the mutant is treated with gibberellin. Treatment with other plant growth regulators does not induce normal flower development. Development of gib-1 flower buds, as measured by progress toward anthesis, ceases at a bud length of 2.5 millimeters; however, increase in size of the bud continues. Buds between 2.5 and 3.7 millimeters are developmentally arrested but still are capable of developing normally after treatment with gibberellic acid. Anthers of these developmentally arrested buds contain pollen mother cells that are in the G1 phase of premeiotic interphase. Following treatment of developmentally arrested buds with gibberellic acid, premeiotic DNA synthesis and callose accumulation in pollen mother cells are evident by 48 hours posttreatment, and within 66 hours, prophase I of meiosis- and meiosis-related changes in tapetum development are observable.

Spindle membranes and calcium sequestration during meiosis ofDysdercus intermedius (Heteroptera)

The fate of intracellular membranes stained by the osmium ferricyanide (OsFeCN) procedure was followed from premeiotic interphase to interkinesis inDysdercus intermedius. During diakinesis the centrioles forming primary cilia attach temporarily with their proximal ends to the nuclear envelope which is stretched from pole to pole. Breakdown of the nuclear envelope is preceded by deep indentations with microtubules from growing asters. Vesicles of smooth endoplasmic reticulum which accumulate gradually in the course of prophase contribute to the ensheathment of the chromosomes with membranes. When the nuclear envelope breaks down, the polar parts of the formerly perinuclear membranes follow the ingrowth of the spindle microtubules towards the cell equator where the seven bivalents are arranged in a circle with the X1X2 sex chromosomes in the centre. The metaphase I spindle thus contains longitudinally oriented membranes between the poles, membranous envelopes around all chromosomes and radial connections from the autosomes to the sex chromosomes in the centre. At anaphase the homologues leave their common sheath and a microtubular stembody surrounded by membranes appears between the receding dyads. In the interkinetic nucleus the gonosomes are separated from the autosomes by a common membranous sheath which may be instrumental in their joint assignment to only one pole in the second meiotic division. Calcium sequestering sites visualized by oxalate precipitation are the Golgi lamellae and vesicles derived from them that surround the whole spindle body.

Induction of reinitiation of meiosis in amphibian Bufo oocytes by injection of ciliate Paramecium extracts

Germinal vesicle breakdown (GVBD) of amphibian Bufo oocytes can be induced if Paramecium extracts were injected into them. The activity of meiosis-reinitiation-inducing factor (MRIF) appeared in premeiotic G 1 cells, then the activity fluctuates according to the degrees of micronuclear chromatin condensation in meiosis. Proliferating micronucleate and amicronucleate cells also showed the same activity. MRIF differed from MPF (M phase promoting factor), because MRIF appeared not only in M phase cells but also in premeiotic interphase cells and its action on the induction of GVBD was inhibited by cycloheximide. Preliminary experiments showed that MRIF was a heat-labile soluble protein.

Coenocytism, ameiosis, and chromosome diminution in intergeneric hybrids in the perennial Triticeae

Three different pathways of ameiotic microsporogenesis were observed in some intergeneric hybrids of the perennial Triticeae grasses. In one of the hybrids between Pseudoroegneria spicata ssp. inermis and Psathyrostachys juncea, pollen mother cells remained as premeiotic interphase cells when the pollen grain wall started to form. The microspores in such an ameiotic plant are presumably unreduced. Coenocyte formation coupled with ameiosis occurred in two hybrid plants of Psathyrostachys huashanica × Secale montanum. Less than 10% of the pollen mother cells had one nucleus. An average of 4.44 nuclei, ranging from 1 to 25 per pollen mother cell, was observed. The nuclei in coenocytes remained unfused when the pollen grain wall was formed. Nucleus splitting followed by cytoplasmic budding or cleavage, possibly a process of chromosome diminution or elimination, replaced meiotic divisions in most of the pollen mother cells in one plant of Leymus angustus × Hordeum bulbosum and two plants of Thinopyrum elongatum × Psathyrostachys juncea. It is evident that these meiotic abnormalities are under genetic control. Probable locations for these genes controlling these phenomena are suggested.Key words: coenocyte, ameiosis, chromosome diminution, chromosome elimination, microsporogenesis, unreduced gamete, polyploidy, intergeneric hybrid.

Effect of auxins and cytokinins upon the start of meiosis in cultured anthers of rye (Secale cereale L.)

Anthers of rye excised at premeiotic interphase were cultured in vitro on a basic medium and on a series of auxin- and cytokinin-containing media to ascertain the effect of these plant hormones upon the start of meiosis. Only in the hormone-containing media was it possible to find anthers in which pollen mother cells had started the meiotic division. The different hormones and concentrations tested exhibited a differential capacity to induce the start of meiosis, the percentages of anthers with pollen mother cells in meiosis varying from 0% in one medium to more than 60% in two other media. In some media, pollen mother cells were able to complete the whole meiotic division, while in others they were arrested at different stages of meiosis.Key words: Secale cereale, meiosis induction, anther culture, plant hormones.

The association of centromeres of nonhomologous chromosomes at meiotic prophase in Beta vulgaris L.

Centromere association involving two or three nonhomologous bivalents at meiotic prophase of Beta vulgaris and monosomic additions of B. vulgaris × B. procumbens are described. The associations, most conspicuous at late pachytene and persisting until diakinesis, occur abundantly in a small group of genetically related plants. The presence of normal bivalents at metaphase I and the absence of irregularities at later stages show that centromere associations do not disturb meiotic recombination and transmission. A mathematical model proposed to test for randomness with respect to the number of bivalents involved in centromere associations, showed significant difference between observed and expected frequencies. On the basis of our own observations and data from the literature, it is suggested that the associations at pachytene and diplotene are delayed separations of centromere aggregates that arise at premeiotic interphase or during the formation of the synizetic knot. A possible molecular mechanism is discussed.Key words: centromere association, meiotic prophase, pachytene chromosomes, nonhomologous association, sugar beet, Beta vulgaris.

The meiotic process and aneuploidy.

Normal females developing at 25 degrees C produce their first population of oocytes at 132 +/- 2 hr post oviposition. Entrance of the oocytes into premeiotic interphase signals initiation of DNA replication which continues for 30 hr. Coincidentally, extensive SCs appear, averaging 50 microns (132 hr), peaking at 75 microns (144 hr), and continuing into early vitellarial stages. Recombinational response to heat, evidenced by enhancement or induction of exchange, is limited to the S-phase with a peak at 144 hr coinciding with maximal extension of the SC. Coincidence of synapsis and recombination response with S at premeiotic interphase is contrary to their conventional localization at meiotic prophase. The interrelationship between exchange and nondisjunction has been clarified by the Distributive Pairing Model of meiosis. Originally revealed through high frequencies of nonrandom assortment of nonhomologous chromosomes, distributive pairing has been shown to follow and to be noncompetitive with exchange, to be based on size-recognition, not homology, and as a raison d'etre, to provide a segregational mechanism for noncrossover homologs. Rearrangements, recombination mutants, and aneuploids may contribute noncrossover chromosomes to the distributive pool and so promote the nonhomologous associations responsible for nondisjunction of homologs and regular segregation of nonhomologs. Further information concerning early meiotic events and their relation to segregation has been revealed by studies of the ts rec-1(26) mutant. Application of the restrictive temperature (31 degrees C) at sequential times during development shows wild-type activity to be drastically reduced beginning at 126/132 hr, terminating at 162 hr, and so coinciding with S and the heat-sensitive period of the normal genome. EMs of serially sectioned oocyte nuclei maintained at the restrictive or control temperature reveal SCs to be indistinguishable, and implicate recombination rather than synapsis as the target of the mutant. Activity of rec-1(26) in the range 17 degrees to 31 degrees C reveals a sharp decline between 28+ degrees and 31 degrees C, typical of a denaturation curve. If denaturation of the rec protein by the restrictive temperature marks its active phase, it follows that recombination terminates at the end of S when the restrictive temperature becomes ineffective. The notion that synapsis ensures regular segregation (35) is invalidated by rec-1(26) whose normal synapsis at the restrictive temperature is followed by a 200-fold increase in nondisjunction at segregation, as compared to the control.(ABSTRACT TRUNCATED AT 400 WORDS)

Observations on Premeiocytes of Wheat

Electron microscopic observations of wheat premeiocytes in premeiotic mitosis (PMM) and premeiotic interphase (PMI) are reported. In archesporial cells of columns at last PMM there is a significant increase in activity of endoplasmic reticulum. Characteristic states of chromatin of premeiocyte nuclei in the stages of PMI are illustrated. Changes in the primary walls of PMCs at Stage 1, PMI, precede the formation of callose in Stage 2. In general, callose deposition throughout the PMC column follows the pattern described elsewhere in Sorghum and other Gramineae. The behavior of the plasmalemma during callose synthesis resembles that described elsewhere in Cucurbita. A role for the cell endomembrane system in callose synthesis is suggested. Blebs arising from the inner membrane of the nuclear envelope, described elsewhere in endosperm cells of wheat and by us in wheat PMCs in this report, give rise to vesicles which carry nuclear material into the cell cytoplasm. Differences in this transport mechanism as it appears in the two tissue systems are described.

OBSERVATIONS ON PREMEIOCYTES OF WHEAT

Electron microscopic observations of wheat premeiocytes in premeiotic mitosis (PMM) and premeiotic interphase (PMI) are reported. In archesporial cells of columns at last PMM there is a significant increase in activity of endoplasmic reticulum. Characteristic states of chromatin of premeiocyte nuclei in the stages of PMI are illustrated. Changes in the primary walls of PMCs at Stage 1, PMI, precede the formation of callose in Stage 2. In general, callose deposition throughout the PMC column follows the pattern described elsewhere in Sorghum and other Gramineae. The behavior of the plasmalemma during callose synthesis resembles that described elsewhere in Cucurbita. A role for the cell endomembrane system in callose synthesis is suggested. Blebs arising from the inner membrane of the nuclear envelope, described elsewhere in endosperm cells of wheat and by us in wheat PMCs in this report, give rise to vesicles which carry nuclear material into the cell cytoplasm. Differences in this transport mechanism as it appears in the two tissue systems are described.

Syntheses of nucleic acids during spermatogenesis in Nereis diversicolor (annelida polychaeta): a quantitative autoradiographic study.

The development of DNA and RNA synthesis in the germ cell population was studied after a 3H-thymidine or 3H-uridine pulse at each stage of spermatogenesis. The autoradiographic results show that the first sign (after 3 days in vitro) of cellular changes is an increase in RNA synthesis which reaches a maximum at day 5. DNA replication (premeiotic S phase) occurred at day 7, then cells entered meiotic prophase (day 9). Meiotic divisions and spermiogenesis occurred after 11 days. Silver grain counts permit the conclusion that RNA synthesis is clearly higher during premeiotic interphase (days 3-7) than during spermatogonial proliferation (day 0). It appears therefore that male meiotic differentiation in Nereidae is accompanied by increased RNA synthesis.

Microspectrophotometric and autoradiographic study of the timing and duration of pre-meiotic DNA synthesis in Paramecium caudatum.

Timing and duration of pre-meiotic G1, S and G2 phases in Paramecium caudatum have been clarified by microspectrophotometry of DNA content and autoradiography with [3H]thymidine ([3H]dThd) and [3H]deoxyuridinemonophosphate ([3H]dUMP). Microspectrophotometric measurement of DNA content showed that cells in stationary phase have a G1 micronucleus, and that pre-meiotic DNA synthesis begins with the micronuclear swelling and ends immediately before the nucleus enters meiotic prophase. The results indicate that the micronucleus is in G1 during the first 1.5 h after onset of mating reaction at 25 degrees C and in pre-meiotic S phase during the following 1 h, and that G2 phase is very short or lacking. Therefore, in P. caudatum, the transition point from pre-meiotic interphase to meiotic prophase is at or very close to the end of pre-meiotic DNA synthesis. Autoradiographic study showed that although [3H]dThd and [3H]dUMP were well utilized in pre-mitotic DNA synthesis, only [3H]dUMP was well utilized in pre-meiotic DNA synthesis. [3H]dThd was utilized very little in pre-meiotic DNA synthesis even if the precursor was taken into the cell, suggesting that the activity of thymidine kinase is low in these cells.

A temporal study at the ultrastructural level of the developing pro-oocyte of Drosophila melanogaster.

The first pro-oocyte in developing pupal germaria of females grown at 25 ° has been followed at 6 h intervals from its formation at ∼129 h post-ovipostion until Stage 1, to provide an unambiguous temporal order. EM autoradiographs were made of sectioned germaria, scanned at lower magnification for location of the pro-oocyte(s) within the most posterior 16-cell cyst and photographed at higher magnification to show the presence of label indicating DNA replication and synaptonemal complex indicating synapsis in the same pro-oocyte nucleus. Label, detected at 132 h and at all subsequent intervals up to and including 162 h, delimited an S-phase of 30 h and identified this period as premeiotic interphase. Extensive SCs (av. length 50 μm/genome) measured in serial sections at 132 h provide irrefutable evidence that synapsis in Drosophila begins close to both pro-oocyte formation and initiation of premeiotic interphase. Measurements of SCs at 6 h intervals during interphase reveal a sharp increase between 132 and 138 h, a peak length (∼75 μm/genome) at 144 h, a decrease and subsequent plateau (∼ 60 μm/genome) from 150–162 h and a further drop (R~50 μm/genome) at Stage 1. Maximal extension of SC at 144 h coincides with maximal genome response to heat (Grell and Day, 1974) and with midpremeiotic-S. Spherical nodules, detected at ∼ 1/genome between 138 and 150 h would, on the questionable assumption that they are the sites of recombination, provide proof of recombination during early interphase, as genetic evidence strongly implies. Evidence contrary to interpretations of fibrillar material within the nucleus as either precursor of the central region of the SC or sagittal sections of the central element of the SC, is presented. No structure corresponding to the polytene chromocenter was observed.

Commitment of Mitotic Cells to Meiosis during the G<sub>2</sub> Phase of Premeiosis

In the developing anther, archesporial cells that proliferate by mitotic division are converted into meiotic cells during the premeiotic interphase. Experiments with explanted microsporocytes of Lilium and Trillium were made to obtain evidence for the conversion of mitotic to meiotic cells during the premeiotic period. Explanted premeiotic cells were cultured through the division cycle at relatively high division frequencies and showed a variety of division types with respect to chromosomal events. The type of division depended on the premeiotic stage at which the cells were explanted. Cells in the G1, S and early G2 phases underwent mitotic division and formed a diad or binucleate monad. Cells explanted at the late G2 phase were cultured throughout the normal meiotic cycle, which resulted in typical tetrad configuration. In microsporocytes explanted during the main part of the G2 interval, centromere behavior was meiotic, but chromosome pairing and chiasma formation were disturbed. Thus, she G2 interval was shown to be critical for the commitment of mitotic cells to meiotic division. Detailed analysis showed that the intracellular changes that commit the cells to meiosis begin shortly after completion of premeiotic DNA synthesis and that these changes are progressive and cumulative.