Chronology and pattern of replication in the bone marrow chromosomes of Gallus domesticus

The duration of the cell generation, the chronology, and the pattern of chromosome duplication was studied in the bone marrow of Gallus domesticus. The duration of the phases of the cell cycle is: cell generation 17.5 hours, S period 9 hours. G₂ period plus prophase stage 2.5 hours, G₁ period 6 hours. Chromosome replication begins at many sites. During middle S it extends to the whole complement and finally finishes in small, late replicating regions of the macrochromosomes. Interchromosomal asynchrony of duplication at the initiation or at the end of the S period was not observed. Z-chromosomes begin and finish DNA synthesis synchronously with the other macrochromosomes. The W-chromosome in females is the last microchromosome to finish replication. However it ends DNA synthesis at about the same time as the macrochromosomes. Similarities and differences between chromosome replication in Aves and Mammalia are considered.     

Evolution of karyotype,sex chromosomes,and meiosis in mygalomorph spiders (Araneae: Mygalomorphae)

Spider diversity is partitioned into three primary clades, namely Mesothelae, Mygalomorphae, and Araneomorphae. Mygalomorph cytogenetics is largely unknown. Our study revealed a remarkable karyotype diversity of mygalomorphs. Unlike araneomorphs, they show no general trend towards a decrease of 2n, as the chromosome number was reduced in some lineages and increased in others. A biarmed karyotype is a symplesiomorphy of mygalomorphs and araneomorphs. Male meiosis of some mygalomorphs is achiasmatic, or includes the diffuse stage. The sex chromosome system X₁X₂0, which is supposedly ancestral in spiders, is uncommon in mygalomorphs. Many mygalomorphs exhibit more than two (and up to 13) X chromosomes in males. The evolution of X chromosomes proceeded via the duplication of chromosomes, fissions, X–X, and X‐autosome fusions. Spiders also exhibit a homomorphic sex chromosome pair. In the germline of mygalomorph males these chromosomes are often deactivated; their deactivation and pairing is initiated already at spermatogonia. Remarkably, pairing of sex chromosomes in mygalomorph females is also initiated at gonial cells. Some mygalomorphs have two sex chromosome pairs. The second pair presumably arose in early‐diverging mygalomorphs, probably via genome duplication. The unique behaviour of spider sex chromosomes in the germline may promote meiotic pairing of homologous sex chromosomes and structural differentiation of their duplicates, as well as the establishment of polyploid genomes. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 109 , 377–408.     

Genetic and physiological parameters associated with cadmium toxicity in Drosophila melanogaster

Two strains of Drosophila melanogaster represent the extremes in resistance and sensitivity to the lethal effects of CdCl₂. The strain containing the mutations vermilion and brown (v; bw) and the strain Austin had LC₅₀'s of 3.3 and 1.3mm CdCl₂, respectively. The three major chromosomes from these two strains were assorted genetically into the six possible combinations. The measured LC₅₀'s for CdCl₂ for these six genotypes fell into two groups according to the X chromosome; those containing the X chromosome from v; bw had LC₅₀'s 0.5–1.0mm greater than those in which the X chromosome was from Austin. Since the parent strains differed by 2mm, we suggest that the X chromosome is a major, but not the sole, site of genes to produce resistance to CdCl₂. When ¹⁰⁹Cd was in the diet the uptake by v; bw and Austin over 2 days was the same. After 4 days of uptake, the Austin strain excreted the ¹⁰⁹Cd five times faster than v; bw but the six genotypes did not differ appreciably in excretion rate from one another and resembled the sensitive parent Austin more than the resistant one. Thus a second process is indicated that distinguishes resistance to CdCl₂ that apparently is not associated with the X chromosome.This research was sponsored by the Office of Health and Environmental Research, U.S. Department of Energy, under Contract DE-AC05-84OR21400 with Martin Marietta Energy Systems, Inc.     

Partial trisomy of the long arm of human chromosome 1 as demonstrated by in situ hybridization with 5S ribosomal RNA

Summary In a newborn boy with multiple malformations, a tandem duplication was detected at the distal end of the long arm of one human chromosome 1. The Giemsa bands, 1q31 to 1q43–44, were repeated serially. Since 5S rRNA genes are located at 1q42–43, in situ hybridization of ¹²⁵I 5S rRNA with fixed chromosome preparations was used to confirm the chromosomal duplication. The infant exhibited numerous developmental and clinical abnormalities as might be expected with an abnormality of chromosome structure relating to a ribosome component.     

THE PATTERN OF DNA SYNTHESIS IN THE CHROMOSOMES OF HUMAN BLOOD CELLS

The sequence in which various regions of the chromosomes of human blood cells complete DNA synthesis in vitro has been studied through the use of H³-thymidine labeling and autoradiography. Certain of its aspects have been defined, and these may serve as a basis for comparing the pattern of synthesis in cells of other tissues. In general, the long chromosomes continue replication later than the short ones. Variability of the sequence has been prominent. One pair from Group 13–15 and pair No. 17 complete replication early. In certain other chromosomes, replication is very active late in the S period, e.g. one X of the female cell, the Y of the male cell, two of Group 4–5, two of Group 13–15, the Nos. 16, and the Nos. 18. In the normal human female a striking correlation exists between the late replication of one of the X chromosomes, condensation during the intermitotic period, and presumed genetical inactivation. The pattern of replication characterizes certain chromosomes whose structural features alone are non-distinctive, and it may be useful in studies of cells in which a chromosomal aberration occurs.     

Mapping of the α4 subunit gene (GABRA4) to human chromosome 4 defines an α2—α4—β1—γ1 gene cluster: further evidence that modern GABAA receptor gene clusters are derived from an ancestral cluster

We demonstrated previously that an α₁—β₂—γ₂ gene cluster of the γ-aminobutyric acid (GABA_A) receptor is located on human chromosome 5q34–q35 and that an ancestral α—β—γ gene cluster probably spawned clusters on chromosomes 4, 5, and 15. Here, we report that the α₄ gene (GABRA4) maps to human chromosome 4p14–q12, defining a cluster comprising the α₂, α₄, β₁, and γ₁ genes. The existence of an α₂—α₄—β₁—γ₁ cluster on chromosome 4 and an α₁—α₆—β₂—γ₂ cluster on chromosome 5 provides further evidence that the number of ancestral GABA_A receptor subunit genes has been expanded by duplication within an ancestral gene cluster. Moreover, if duplication of the α gene occurred before duplication of the ancestral gene cluster, then a heretofore undiscovered subtype of α subunit should be located on human chromosome 15q11–q13 within an α₅—α_x—β₃—γ₃ gene cluster at the locus for Angelman and Prader—Willi syndromes.     

Replication of X chromosomes in complete moles

Summary DNA replication patterns of X chromosomes in complete hydatidiform moles were studied using cultured fibroblasts from three 46,XX moles resulting from duplication of a haploid sperm, and from a 46,XY mole originating from dispermy. Control cultures included skin fibroblasts from an adult woman and a female fetus as well as PB lymphocytes from an adult woman. Cultures were treated with 5-bromodeoxyuridine for the last 2–4h of the S phase, and the chromosome slides prepared were stained by the Hoechst 33258-Giemsa procedure. Each of the three XX moles studied revealed one early-replicating and one late-replicating X chromosomes, while the XY mole revealed one early-replicating X chromosome. DNA replication patterns of molar X chromosomes were similar to those of adult and fetal fibroblasts, but different from those in adult lymphocytes. These findings indicate that DNA replication kinetics of molar fibroblasts are tissue-specific rather than origin- or developmental-stage specific.     

Reversion of XO to XY sex chromosome mechanism in a phasmid

Summary The sex chromosomes of the male phasmid Isagoras schraderi Rehn comprise an X and a Y, — each with a submedian kinetochore, and one euchromatic and one heterochromatic arm. At meiosis X and Y form an unequal sex bivalent in which the euchromatic arms are terminally associated. Relatively recent reversion from the XO-XX mechanism characteristic of the Phasmidae is indicated by the presence of the euchromatic arm in both X and Y. The diploid number of the male is 34.Unequal autosomal bivalents are found at meiosis in two other species of Isagoras — Isagoras subaquiles Rehn and Isagoras sp. — and in Pseudophasma menius Westwood. The chromosome complements of these species are described.     

Chromosome delineation induced by a combined potassium permanganate-sodium bisulfite treatment

A cytological procedure for in vitro chromosome delineation has been studied using human and mouse (Mus musculus) chromosomes. This method, consisting of slide incubation in KMnO₄ at 0–5° C for 24 h followed by a short exposure to NaHSO₃ (1–3 min) and Giemsa staining, induces extraction of chromatin from human and mouse interphase nuclei and chromosomes. Autoradiography after ³H-ThD incorporation in vitro and cytophotometry confirmed that DNA is removed. Well contour-delineated and non-distorted chromosomes are observed in both species allowing the identification of all human chromosome groups. Contour chromosome delineation and its relationship to chromosome organization is briefly discussed.     

Mykologische Studien

Ohne ZusammenfassungMykol. Studien I–X mit insges. 158 Seiten, 12 Tafeln und 35 Textabb.:I. Ein Experiment mitPhallus, Archiv f. Protistenkunde, Bd. 64, 1928, S. 1–18, 1 Tafel.II.Geaster triplex, ebenda, Bd. 65, 1929, S. 65–77, 8 Textabb.III.Xanthochrous cuticularis, ebenda, Bd. 65, 1929, S. 321–329, 5Textabb.IV. Zur Entwicklungsgeschichte vonMutinus caninus, ebenda, Bd. 72, 1930, S. 214–246, 2 Tafeln.V. ZuXanthochrous cuticularis undXanth. hispidus, ebenda, Bd. 72, 1930, S. 420–432, 4 Tafeln, 3 Textabb.VI.Spongipellis Litschaueri (=Polyporus Schulzeri Fr. sensuBresadola), ebenda, Bd. 75, 1931, S. 297–314, 2 Tafeln, 2 Textabb.VII.Mycenastrum corium, ein für Deutscheuropa neuer Gastromycet, ebenda, Bd. 78, 1932, S. 473–494, 7 Textabb.VIII.Bovista echinella undLycoperdon velatum, Beihefte z. Bot. Centralbl., Bd. 51, 1933, Abt. I, S. 269–286, l Tafel, 4 Textabb.IX. Über die Fruchtkörperentwicklung der Geastraceen, ebenda, Bd. 52, 1934, Abt. A, S. 269–289, 1 Tafel, 6 Textabb.X.Pleurotus calyptratus Fr., Biologia generalis, Bd. 10, 1934, S. 457–368, 1 Tafel.     

Microelectrode characterization of the basolateral membrane of rabbit S3 proximal tubule

Summary The purpose of this study was to characterize the basolateral membrane of the S3 segment of the rabbit proximal tubule using conventional and ion-selective microelectrodes. When compared with results from S1 and S2 segments, S3 cells under control conditions have a more negative basolateral membrane potential (V _bl=–69 mV), a higher relative potassium conductance (t _K=0.6), lower intracellular Na⁺ activity (A _Na=18.4mm), and higher intracellular K⁺ activity (A _K=67.8mm). No evidence for a conductive sodium-dependent or sodium-independent HCO ₃ ^– pathway could be demonstrated. The basolateral Na–K pump is inhibited by 10^–4 m ouabain and bath perfusion with a potassium-free (0-K) solution. 0-K perfusion results inA _Na=64.8mm,A _K=18.5mm, andV _bl=–28 mV. Basolateral potassium channels are blocked by barium and by acidification of the bathing medium. The relative K⁺ conductance, as evaluated by increasing bath K⁺ to 17mm, is dependent upon the restingV _bl in both S2 and S3 cells. In summary, the basolateral membrane of S3 cells contains a pump-leak system with similar properties to S1 and S2 proximal tubule cells. The absence of conductive bicarbonate pathways results in a hyperpolarized cell and larger Na⁺ and K⁺ gradients across the cell borders, which will influence the transport properties and intracellular ion activities in this tubule segment.     

DNA REPLICATION PATTERNS AND CHROMOSOMAL PROTEIN SYNTHESIS IN OPOSSUM LYMPHOCYTES IN VITRO

DNA replication patterns were determined in the autosomes and sex chromosomes of phytohemagglutinin-stimulated lymphocytes from the opossum (Didelphis virginiana) by employing thymidine-³H labeling and high-resolution radioautography. Opossum chromosomes are desirable experimental material due to their large size, low number (2n = 22), and morphologically distinct sex chromosomes. The autosomes in both sexes began DNA synthesis synchronously and terminated replication asynchronously. One female X chromosome synthesized DNA throughout most of the S phase. Its homologue, however, began replication approximately 3.5 hr later. The two X's terminated DNA synthesis synchronously, slightly later than the autosomes. This form of late replication, in which one X chromosome begins DNA synthesis later than its homologue but completes replication at the same time as its homologue, is apparently unique in the opossum. The male X synthesized DNA throughout S while the Y chromosome exhibited late-replicating characteristics. The two sex chromosomes completed synthesis synchronously, slightly later than the autosomes. Grain counts were performed on all chromosomes to analyze trends in labeling intensity at hourly intervals of S. By analyzing the percent of labeled mitotic figures on radioautographs at various intervals after introduction of arginine-³H, chromosomal protein synthesis was found not to be restricted to any portion of interphase but to increase throughout S and into G₂.     

Initial phases of DNA synthesis in Drosophila melanogaster

Replication patterns of the X chromosomes and autosomes in D. melanogaster male and female larvae during the discontinuously labeled initial and end phases of DNA synthesis were compared. In female larvae X and autosomes behaved correspondingly during all the replication stages. In males, however, the X chromosome shows a differential replication behavior from that of the autosomes already during the discontinuously labeled initial stage.—In those nuclei of both sexes, in which the autosomes correspond in their initial replication patterns, significantly more labeled regions are to be found over the male X than over the female X. The complementary behavior during the end phases (Berendes, 1966), i.e. the reverse of that above, leads to an earlier completion of the replication cycle in most of the labeled regions of the male X chromosome. The differential replication revealed in the autoradiograms is interpreted as a consequence of the polytene structure in giant chromosomes.     

Ultrastructural characterization of the sex chromosomes during spermatogenesis of spiders having holocentric chromosomes and a long diffuse stage

An ultrastructural study has been made of spermatogenesis in two species of primitive spiders having holocentric chromosomes (Dysdera crocata, XO and Segestria florentia X₁X₂O). Analysis of the meiotic prophase shows a scarcity or absence of typical leptotene to pachytene stages. Only in D. crocata have synaptonemal complex (SC) remnants been seen, and these occurred in nuclei with an extreme chromatin decondensation. In both species typical early prophase stages have been replaced by nuclei lacking SC and with their chromatin almost completely decondensed, constituting a long and well-defined diffuse stage. Only nucleoli and the condensed sex chromosomes can be identified. — In S. florentina paired non-homologous sex chromosomes lack a junction lamina and thus clearly differ from the sex chromosomes of more evolved spiders with an X₁X₂O male sex determination mechanism. In the same species, sex chromosomes can be recognized during metaphase I due to their special structural details, while in D. crocata the X chromosome is not distinguishable from the autosomes at this stage. — The diffuse stage and particularly the structural characteristics of the sex chromosomes during meiotic prophase are reviewed and discussed in relation to the meiotic process in other arachnid groups.     

Classification of staphylococci based on chemical and biochemical properties

Summary In the present work the chemical cell wall composition and some other biochemical characteristics were studied in staphylococci with the intention of utilizing the data obtained in their classification.According to the cell wall peptidoglycans and teichoic acids, the 130 strains of staphylococci studied are divided into 10 major groups. This division of staphylococci into groups is in good agreement with their present classification only in some cases. All of the 47Staphylococcus aureus strains contain a cell wall peptidoglycan of thel-Lys-Gly_5–6 type and ribitol teichoic acid. Coagulase-negative staphylococci are more heterogeneous and are divided according to their cell wall composition into 9 major groups. 21 strains of them are classified asS. epidermidis sensu stricto. They form a natural group and are distinguished by the occurrence of thel-Lys-Gly_4–5,l-Ser_0.5–1.8 peptidoglycan type, glycerol teichoic acid and anl-lactate dehydrogenase which is activated by fructose-1,6-diphosphate. 8 strains with peptidoglycan of thel-Lys-Gly_4–5,l-Ser_0.5–1.8 type and ribitol teichoic acid are labeled asS. saprophyticus. The remaining groups have not been given species names and require further extensive comparative study.     

Replication variants of the human inactive X chromosome

Replication variants of the inactive X chromosome were investigated in lymphocytes from six donors by means of terminal BrdU or thymidine incorporation. There were interindividual differences in the incidence of particular variants. In endoreduplicated and tetraploid cells both allocyclic X chromosomes showed the same replication sequence. The Xp22 band of the allocyclic X chromosome seemed to replicate later than the homologous material in some cells. Initiation time of DNA synthesis within the inactive X chromosome was found to be stable; termination time, however, varied greatly relative to the other chromosomes. Early completion of replication within the heterochromatic X chromosome could be demonstrated preferentially for the Xq25–27 terminal sequence, but other variants expressed the phenomenon also. A variable replication rate of the inactive X chromosome is believed to be responsible for its asynchronous, independent replication. The biological significance of the phenomenon is discussed with respect to cell differentiation.     

Purification and characterization of two ascorbate peroxidases of rice (<Emphasis Type="Italic">Oryza sativa </Emphasis>L.) expressed in <Emphasis Type="Italic">Escherichia coli</Emphasis>

To clarify the diversity and function of isozymes of ascorbate peroxidase (APX) in plants, a method of producing large quantities of these proteins is needed. Here, we describe an Escherichia coli expression system for the rapid and economic expression of two rice APX genes, APXa and APXb (GeneBank accession Nos. D45423 and AB053297, respectively). The two genes were cloned into the pGEX-6p-3 vector to allow expression of APX as a glutathione-S-transferase (GST) fusion protein. The GST-APXa and GST-APXb fusion proteins were purified by affinity chromatography using a glutathione-Sepharose 4B column, with final yields of 40 and 73 mg g^–1 dry cells, respectively. Specific activities were 15 and 20 mM ascorbate min^–1 mg^–1 protein, respectively. The K_m values for ascorbate were 4 and 1 mM, respectively, and those for H₂O₂ were 0.3 and 0.7 mM, respectively indicating that the two rice isoenzymes have different properties.Revisions requested 27 September 2004; Revisions received 12 November 2004     

Ultrastructure of the nuclei during different phases of the cell cycle in the antheridial filaments ofChara vulgaris L.

Summary Synchronously dividing nuclei of the antheridial filaments ofChara vulgaris at the 32-celled stage have different structure depending on the period of interphase.During S phase which begins as early as at the start of telophase (coincidently with the nuclear envelope formation and chromosome decondensation) one can observe a gradual reduction in the content of condensed chromatin, having the appearance of an indistinct network. During the middle S period the area of condensed chromatin decreases to the lowest level of about 29% of nuclear profile and the nuclear envelope becomes folded. At the end of S phase the condensed chromatin forms a more distinct and thicker reticulum which covers an area of about 52%.During the early G₂ phase, the area occupied by the condensed chromatin was about 41% and it was found to assume the shape of large and iregular clusters localized mainly near the nucleoli. The reticulate form of chromatin, characteristic of the S period, disappears almost completely. During the next period of interphase the condensed chromatin disperses considerably and covers now 24% of the area. At the end of the G₂ phase the condensed chromatin reappears and transforms into chromosomes. Then the condensed chromatin removes from the nuclear envelope.This work was supported by the Polish Academy of Sciences within the project 09.7.3.1.4.     

Dissociation of cellular K+ accumulation from net Na+ transport by toad urinary bladder

Summary A number of published data suggest a variable stoichiometry between the rates of cellular potassium uptake and net sodium transport (J _Na) across the urinary bladder of the toad. This problem was examined by simultaneously studying the intracellular chemical activity of potassium (a _K) with open-tip K⁺-selective microelectrodes and micropipets, and monitoringJ _Na by measuring the short-circuit current (SCC). When bathed in the short-circuited state with solutions containing ana _K of 2.7mm, the mean ±sem values for intracellulara _K were 43±0.6mm.Ouabain, at a concentration of 10^–2 m, reduced intracellulara _K by 56–67% and SCC by 96–100%. At 5×10^–4 m, ouabain reversibly reduced intracellulara _K by 40–55%, and SCC by 63–68%; the inhibition of SCC was only partly reversible during the period of observation.Removal of external potassium reduced intracellulara _K by 69–80% and SCC by 51–76%. Restoration of external potassium entirely returned intracellulara _K to its control value, but only partially reversed the inhibition of SCC during the period of study. Furthermore, recovery ofa _K began 19–43 min before that of SCC; recovery ofa _K was 90–97% complete before any increase in SCC could be measured. Although other interpretations are possible, the simplest interpretation of the data is that the processes responsible for potassium accumulation and transepithelial sodium transport are not identical. We propose the existence of a separate transfer mechanism at the basolateral cell membrane, responsible for accumulating intracellular potassium, and not directly coupled to active sodium transport.     

Chromosome replication in four species of snakes

Chromosome measurements were performed in four species of snakes related at the level of suborder (Boa constrictor amarali, Xenodon merremii, Philodryas patagoniensis, Bothrops jararaca). The data obtained point out that pairs 1–3 were common to the four snakes and probably inherited from the ancestor of the suborder Serpentes. Pairs 5–8-W were characteristic of each snake; hence, it is possible to assume that they followed evolving after the appearing of the suborder Serpentes. Z-chromosomes were metacentric in B. constrictor amarali, X. merremii and B. jararaca and slightly submetacentric in P. patagoniensis. Area of these chromosomes varied from 8.6–10.6% of the haploid set in the four species studied.-The study of chromosome replication at the end of the S period points out that shared chromosomes have similar patterns of labeling. Therefore, it is proposed that the distribution of late replicating regions and heterochromatin in the genome is phylogenetically transmitted and probably genetically determined.—The analysis of the ending-sequence of chromosome replication shows that sex chromosomes finish earlier than macroautosomes. It is concluded that snakes probably have no mechanism of sex chromosome heterochromatinization in either sex. The absence of late replicating Z-chromosome in the males, favours the hypothesis that no mechanism of sex dosage compensation is acting in the suborder Serpentes.This work was partially supported by Public Health Service Research grant No. GM-14577-03 from the National Institute of General Medical Sciences and by the Fundo de Pesquisas do Instituto Butantan.