Behavior of sperm nuclei in intact and bisected metaphase II mouse oocytes fertilized in the presence of colcemid

Our objective was to examine the developmental fate of sperm nuclei in oocytes fertilized under conditions of meiotic arrest. Therefore zona-free metaphase II oocytes and oocyte fragments (nucleate and anucleate) were fertilized in the presence of colcemid. In anucleate oocyte fragments, normal male pronuclei develop. In contrast, in intact oocytes and nucleate fragments sperm nuclei after initial decondensation undergo secondary condensation. This state is maintained as long as the oocytes are treated with colcemid. When the drug is removed 3 h after insemination, the meiotic spindle(s) is reconstructed, the second polar body(ies) is extruded, and a female pronucleus (or micronuclei) forms. At the same time the sperm nucleus decondenses again and transforms into a male pronucleus. In addition oocytes fertilized in the presence of colcemid could not be refertilized. These observations suggest that oocytes and oocyte fragments fertilized in the presence of colcemid undergo activation despite the failure of pronucleus formation. The inhibitory effect of colcemid on the formation of pronuclei is expressed only in the presence of oocyte chromosomes. We suggest that colcemid stabilizes factors responsible for chromosome condensation that are associated with oocyte chromosomes but not factors (whether the same or different) present in the cytoplasm.     

Pattern formation in fragmented eggs of the short germ insect Schistocerca gregaria

Summary The effect of transverse fragmentation on the segment pattern of the short germ embryo of the locust Schistocerca gregaria has been investigated at two stages subsequent to the formation of the germ anlage. Following fragmentation both anterior and posterior partial embryos were observed, although rarely in a single egg. Anterior partial patterns usually terminated with a segment visible at the time of fragmentation or with the next segment due to appear. Posterior partial patterns began with a wide range of segments depending on the level of fragmentation.Anterior and posterior partial patterns developing in a single egg were usually not complementary and the segments missing sometimes included some segments visible when the embryo was fragmented. Non-complementary patterns resulted following fragmentation in all regions, while complementary patterns only occurred after fragmentation in the visibly-segmented region.The results suggest that following fragmentation isolated posterior portions of the embryo continue to form segments, while isolated anterior regions usually do not. This effect could result from variable damage to an existing pattern of unequally-sized segment primordia, or from the disruption of a process of sequential segmentation in the elongating posterior region of the embryo. The results are broadly compatible with the progress zone model proposed by Summerbell et al. (1973).     

Reaction on alternating GC oligonucleotide templates

Summary The condensation reactions of activated nucleotides, ImpN or 2-MeImpN, with the selfcomplementary ribo-octanucleotide GCGCGCGC or with the partially self-complementary heptanucleotide GCGCGCG were studied. The templatedirected reaction of 2-meImpC with the heptamer yields the 3–5 octamer as the main product. All other reactions yield 2–5-linked octamers and pyrophosphates as major products. Surprisingly, 2-MeImpG facilitates the reaction of 2-MeImpC with the heptamer.Procedures for the analysis by gel electrophoresis of the oligomeric products obtained in reactions of this kind are described.     

草鱼染色体的包装（间期—中期）

以草鱼ＺＣ７９０１细胞株为材料,观察鱼类细胞从间期染色质到中期染色体的包装过程。主要通过（１）分裂期与间期细胞融合,诱导染色体早熟凝集;（２）染色体“伸长”处理;（３）培养细胞的低渗处理;（４）染色质辅展等方法,制作染色体标本,进行扫描和透射电镜观察。观察表明,鱼类染色质的基本结构与哺乳类细胞相同,也是直径约１０ｎｍ的核丝。染色体的色装有两种形式：一种是多级螺旋化形成直径约３００ｎｍ的染色单体,     

Successful synthesis of a glial-specific blood–brain barrier shuttle peptide following a fragment condensation approach on a solid-phase resin

Successful manual synthesis of the TD2.2 peptide acting as a blood–brain barrier shuttle was achieved. TD2.2 was successfully synthesised by sequential condensation of four protected peptide fragments on solid-phase settings, after several unsuccessful attempts using the stepwise approach. These fragments were chosen to minimise the number of demanding amino acids (in terms of coupling, Fmoc removal) in each fragment that are expected to hamper the overall synthetic process. Thus, the hydrophobic amino acids as well as Arg(Pbf) were strategically spread over multiple fragments rather than having them congested in one fragment. This study shows how a peptide that shows big challenges in the synthesis using the common stepwise elongation methodology can be synthesised with an acceptable purity. It also emphasises that choosing the right fragment with certain amino acid constituents is key for a successful synthesis. It is worth highlighting that lower amounts of reagents were required to synthesise the final peptide with an identical purity to that obtained by the automatic synthesiser.     

Chemical probes for investigating protein liquid-liquid phase separation and aggregation

Protein liquid-liquid phase separation drives the dynamic assembly of membraneless organelles for fulfilling different physiological functions. Under diseased condition, protein may undergo liquid-to-solid condensation to form pathological amyloid aggregates closely associated with neurodegenerative diseases. Chemical probe serves as an important chemical tool not only for exploring the basic principle of the dynamic assembly of different protein condensates in vitro and in cell but also for clinical diagnosis and therapeutics of the related diseases. In this review, we first introduce chemical probes to image and regulate protein condensates. Then, we summarized three different categories of chemical probes including general amyloid dye, selective positron emission tomography tracer, and disaggregating binder, which feature distinct interaction pattern and activity upon binding to different pathological amyloid fibrillar aggregates. Next, we discuss the development of chemical probes for tracking protein amorphous aggregates in cells. Finally, we point out future direction in expanding the probes’ chemical space and applications.     

Cold-induced Chromosome Regions and Karyosystematics in Sambucus and Viburnum

The genera Viburnum, Sambucus and Lonicera have been investigated for chromosome number and karyomorphology including Giemsa-C-banding, fluorochrome (DAPI/CMA) banding and cold treatment. Cold-induced undercontracted chromosome regions (CIRs) are found in Viburnum and Sambucus for the first time and are apparently identical with larger hc regions, shown by Giemsa C-banding. Certain narrow C-bands are not cold-sensitive. CIRs frequently react brightly CMA-positive in Viburnum and Sambucus, while DAPI fluorescence is virtually ineffective. The occurrence of CIRs within plants is possibly linked to certain nuclear characters such as large chromosomes and continuous condensation behaviour. Cold-induction has possibly also some influence on euchromatin condensation characteristics in prophasic chromosomes. Several karyological characters point to a closer relationship between Viburnum, Sambucus and Adoxa: Relatively large chromosomes, continuous condensation behaviour, reticulate to semireticulate interphase nuclei and presence of CIRs. These genera appear isolated from Lonicera and the Caprifoliaceae s.str., which differ remarkably in karyomorphology.     

The function of type IV collagen during Drosophila embryogenesis

A collagen gene (Dcg1) was characterized in Drosophila melanogaster and shown to encode a peptide related to vertebrate basement membrane type IV collagen chains. To study the function of type IV collagen during Drosophila development, we transformed flies with a partially truncated Dcg1 gene under the control of a heat-shock promotor. This construct induced synthesis of shortened pro- chains which associated with normal ones and thereby caused degradation of the shortened and normal pro- chains through a process called pro-collagen suicide. A large proportion of embryos expressing the transgene developed a phenotype exhibiting absence or partial retraction of the germ band with defects in nerve cord condensation and dorsal closure. Together these results indicated that, during embryogenesis, type IV collagen was an essential guiding factor for cell-matrix interactions in morphogenetic events.     

Induction of Blocks in Nuclear Divisions and Overcondensation of Meiotic Chromosomes with Cycloheximide During Conjugation of Tetrahymena thermophila

During conjugation, the micronucleus of Tetrahymena thermophila undergoes five consecutive nuclear divisions: meiosis, third prezygotic division (pregamic mitosis) and two postzygotic mitoses of the synkaryon. The four products of the synkaryon differentiate into macronuclear anlagen and new micronuclei and the old macronucleus is resorbed. The protein synthesis inhibitor cycloheximide, applied during conjugation, induced several developmental blocks. Pairs shifted to the drug during early meiotic prophase (stages I–III) were arrested at prophase. Cycloheximide applied to cells at pachytene (stages IV-VI) to metaphase arrested the conjugants at the stage of modified prometaphase/metaphase with overcondensed, swollen bivalents. In contrast to other systems, in the presence of cycloheximide, separation of chromatids, decondensation of chromosomes and exit from metaphase I were inhibited in both diploid and haploid cells. Pairs shifted to the drug after metaphase I were arrested at postmeiotic interphase after completing one nuclear cycle. The same rule applied to the subsequent cycle; then cells were arrested at the stage of pronuclei, and those pairs with functional pronuclei and synkarya were arrested at the stage of two products of the first postzygotic division (pronuclei were not arrested in nuclear transfer and karyogamy). Only pairs with two products of the first postzygotic division were arrested at the same stage after the cycloheximide treatment. Pairs shifted to cycloheximide during the second postzygotic division were arrested in development of macronuclear anlagen and resorption of old macronuclei. The postmeiotic conjugants pulse-treated with cycloheximide (2 h) yielded heterokaryons retaining parental macronuclei (i.e. they exhibited macronuclear retention).     

Rapid reorganization of microtubular cytoskeleton accompanies early changes in nuclear ploidy and chromatin structure in postmitotic cells of barley leaves infected with powdery mildew

Summary Post-mitotic epidermal cells of barley leaves were found to contain, in addition to cortical microtubules (CMTs), distinct arrays of endoplasmic microtubules (EMTs). These encircle nuclei and continuously merge into the CMT arrays that underly the plasmalemma. Detailed three-dimensional reconstruction of both types of MTs during fungal infection showed that profound and very rapid MT rearrangements occurred especially in the case of incompatible (resistant) barley-powdery mildew genotype combination. The most early MT responses, followed by their subsequent complete disintegration, were recorded around nuclei. These events might be relevant for the induction of such nuclear processes as onset of DNA synthesis and nuclear chromatin condensation. Observed pattern of early infection events, as well as less prominent responses in the case of compatible (susceptible) barley-powdery mildew genotype combination, both findings suggest that rapid reorganization of the MT cytoskeleton could be involved in recognition of the fungus by host cells and in the initiation of resistance responses in barley leaves. We hypothesize that the integrity and dynamics of the MT cytoskeleton, especially of its perinuclear part, might participate in control mechanisms involved in activation of resistance genes.Abbreviations CMTs cortical microtubules - EMTs endoplasmic microtubules - MT microtubules - PI propidium iodide - SC sensitive combination - RC resistant combination