G1 Chromosomes Research Articles

Mitotic chromosomes are self-entangled and disentangle through a topoisomerase-II-dependent two-stage exit from mitosis

The topological state of chromosomes determines their mechanical properties, dynamics, and function. Recent work indicated that interphase chromosomes are largely free of entanglements. Here, we use Hi-C, polymer simulations, and multi-contact 3C and find that, by contrast, mitotic chromosomes are self-entangled. We explore how a mitotic self-entangled state is converted into an unentangled interphase state during mitotic exit. Most mitotic entanglements are removed during anaphase/telophase, with remaining ones removed during early G1, in a topoisomerase-II-dependent process. Polymer models suggest a two-stage disentanglement pathway: first, decondensation of mitotic chromosomes with remaining condensin loops produces entropic forces that bias topoisomerase II activity toward decatenation. At the second stage, the loops are released, and the formation of new entanglements is prevented by lower topoisomerase II activity, allowing the establishment of unentangled and territorial G1 chromosomes. When mitotic entanglements are not removed in experiments and models, a normal interphase state cannot be acquired.

Cell-cycle regulation of cohesin stability along fission yeast chromosomes

Sister chromatid cohesion is mediated by cohesin, but the process of cohesion establishment during S-phase is still enigmatic. In mammalian cells, cohesin binding to chromatin is dynamic in G1, but becomes stabilized during S-phase. Whether the regulation of cohesin stability is integral to the process of cohesion establishment is unknown. Here, we provide evidence that fission yeast cohesin also displays dynamic behavior. Cohesin association with G1 chromosomes requires continued activity of the cohesin loader Mis4/Ssl3, suggesting that repeated loading cycles maintain cohesin binding. Cohesin instability in G1 depends on wpl1, the fission yeast ortholog of mammalian Wapl, suggestive of a conserved mechanism that controls cohesin stability on chromosomes. wpl1 is nonessential, indicating that a change in wpl1-dependent cohesin dynamics is dispensable for cohesion establishment. Instead, we find that cohesin stability increases at the time of S-phase in a reaction that can be uncoupled from DNA replication. Hence, cohesin stabilization might be a pre-requisite for cohesion establishment rather than its consequence.

A polymer model for the structural organization of chromatin loops and minibands in interphase chromosomes.

A quantitative model of interphase chromosome higher-order structure is presented based on the isochore model of the genome and results obtained in the field of copolymer research. G1 chromosomes are approximated in the model as multiblock copolymers of the 30-nm chromatin fiber, which alternately contain two types of 0.5- to 1-Mbp blocks (R and G minibands) differing in GC content and DNA-bound proteins. A G1 chromosome forms a single-chain string of loop clusters (micelles), with each loop approximately 1-2 Mbp in size. The number of approximately 20 loops per micelle was estimated from the dependence of geometrical versus genomic distances between two points on a G1 chromosome. The greater degree of chromatin extension in R versus G minibands and a difference in the replication time for these minibands (early S phase for R versus late S phase for G) are explained in this model as a result of the location of R minibands at micelle cores and G minibands at loop apices. The estimated number of micelles per nucleus is close to the observed number of replication clusters at the onset of S phase. A relationship between chromosomal and nuclear sizes for several types of higher eukaryotic cells (insects, plants, and mammals) is well described through the micelle structure of interphase chromosomes. For yeast cells, this relationship is described by a linear coil configuration of chromosomes.

The Relationship of DNA and Chromosome Damage to Survival of Synchronized X-Irradiated L5178Y Cells: I. Initial Damage

We investigated the role of initial DNA and chromosome damage in determining the radiosensitivity difference between the variant murine leukemic lymphoblast cell lines L5178Y-S (sensitive) and L5178Y-R (resistant) and the difference in cell cycle-dependent variations in radiosensitivity of L5178Y-S cells. We measured initial DNA damage (by the neutral filter elution method) and chromosome damage (by the premature chromosome condensation method) and compared them with survival (measured by cloning) for both cell lines synchronized in G1 or G2 phase of the cell cycle (by centrifugal elutriation) and irradiated with low doses of X rays (up to 10 Gy). The initial yield of DNA and chromosome damage in G2 L5178Y-S cells was almost twice that in G1 L5178Y-S cells and G1 or G2 L5178Y-R cells. In all cases DNA damage expressed as relative elution corresponded with chromosome damage (breaks in G1 chromosomes, breaks and gaps in G2 chromosomes). Also we found that the initial DNA and chromosome damage did not determine cell age-dependent radiosensitivity variations in L5178Y-S cells, as there was less initial damage in the more sensitive G1 phase than in the G2 phase. L5178Y-R cells showed only small changes in survival or initial yield of DNA and chromosome damage throughout the cell cycle. Because survival and initial damage in sensitive and resistant cells irradiated in G2 phase correlated, the difference in radiosensitivity between L5178Y-S and L5178Y-R cells might be determined by initial damage in G2 phase only.

Condensation-inhibition by 33258-Hoechst of centromeric heterochromatin in prematurely condensed mouse chromosomes

The phenomenon of premature chromosome condensation has been applied to study the kinetics of condensation-inhibition exerted by the fluorochrome 33258-Hoechst (33258-H) on the centromeric heterochromatic regions of mouse chromosomes. Asynchronous mouse A-9 cells in culture were fused with mitotic HeLa cells in the presence of 33258-H. Pronounced condensation-inhibition of the c-heterochromatin was observed in prematurely condensed early G2, S and late G1 chromosomes in the 33258-H-treated cells. It is concluded that the c-heterochromatic regions begin to condense quite early in G2, decondense again late in G1 and remain decondensed in the S phase.

Visualization of interphase chromosomes.

Using a modified Giemsa-banding technique we have observed what appear to be chromosomes during interphase in nuclei from Allium cepa root tips and Chinese hamster cells (CHO line). During telophase through G1 chromosomes progressively uncoil and decondense. During S chromosomes are comparatively decondensed, but some segments have structure similar to chromosomes in G1 and G2. During G2 the chromosomes progressively recondense and coil in apparent preparation for prophase. Although specific structural modifications of chromosomes occur in G1, Sand G2 nuclei, chromosomes appear never to decondense to the point that they lose their 3-dimensional integrity, but remain in distinct domains throughout interphase.

Is chromatid-type damage in ataxia telangiectasia after irradiation at G0 a consequence of defective repair?

THE radiosensitivity of patients with ataxia telangiectasia (AT) has been indicated by in vivo studies1–3 and in vitro studies on skin fibroblasts4. We present data here to show that this radiosensitivity is observed at the chromosomal level in blood lymphocytes X irradiated at different stages of the cell cycle and for the same range of doses as our survival experiments4. Our results are consistent with the hypothesis that there is a defect in some form of DNA repair in these patients. From the observation of the types of chromosome aberrations seen in AT lymphocytes after irradiation at G0/early G1, we propose a mechanism for the origin of these aberrations, based on the premise that the G0/early G1 chromosome contains a single DNA double helix, in which one polynucleotide chain can be damaged independently of the other.

Analysis of Interphase Chromosome Damage by Means of Premature Chromosome Condensation after X- and Ultraviolet-Irradiation

Sendai virus-mediated fusion between mitotic and interphase mammalian cells causes the rapid condensation of the interphase chromosomes into distinct structures, a process termed premature chromosome condensation. This phenomenon has been used to assess the immediate action of x-rays and ultraviolet light on the chromosomes of HeLa cells irradiated in the G1 phase of the life cycle. X-irradiation produces fragmented chromosomes; but even the most finely chopped fragments retain the condensed morphology characteristic of the premature chromosome condensation of unirradiated G1 cells. For doses up to about 1800 rads, the increase in the number of fragments is linearly related to the dose. One mean lethal dose (about 100 rads) yields a net increase of 10-15 fragments per G1 cell, which is considerably larger than previous estimates based on scoring of mitotic chromosomes. Incubation of irradiated cells produces a rapid (within 2 hr) reduction in the number of fragments, indicative of a rejoining process. The decrease in the number of pieces is not accompanied by unscheduled DNA synthesis detectable by radioautography. G1 chromosomes of cells irradiated with UV light in G1 phase are not appreciably fragmented but are elongated and attenuated so that they resemble the premature-chromosome-condensation chromosomes of unirradiated S-phase cells. Both the degree of "S-like" character attained by the G1 chromosomes in a cell and the percentage of the cell population displaying the G1 --> S transition increase with the dose and incubation time after irradiation. Thus, in contrast to the immediate manifestation of damage from x-rays, the maximum induction of the "S-like" state does not occur until about 2 hr after irradiation. The "S-like" chromosomes are capable of unscheduled DNA synthesis. We suggest that the difference in chromosome morphology found after UV- and x-irradiation underlies the reason why the former, but not the latter, induces unscheduled DNA synthesis in G1 cells.

Methylation in the phage carried by Escherichia coli 15T − (DNA methylation in phage ψ)

Mitomycin C (MMC), actinomycins (ACMs), daunomycin (DNM) and adriamycin (ADR) are antineoplastic, antibiotics, with the anthracyclines (DNM and ADR) being more promising that the others. All these chemicals interact with DNA. MMC brings about cross-linking of the DNA strands and acts as mono-, as well as, bifunctional alkylating agent. The other three intercalate with the DNA molecule. ACMs, especially ACM-D, inhibits RNA synthesis preferentially and discriminates between different species of RNA. At high concentrations, DNA, RNA and protein synthesis are all inhibited by any of these antibiotics. The agents effect the cell cycle traverse at various stages; MMC being most unspecific of all. Mitotic cells at post-G2 stages are the least effected by any of these chemicals.All these chemicals induce chromosome aberrations of chromatid type. The anthracyclines, in addition, produce chromosome-type aberrations perhaps by affecting the single stranded G1 chromosome. Rejoining occurs, but MMC creates a high frequency of quadriradial configurations by utilizing the analogous nucleotide sequences in the repetitive DNA. At other points, it mostly creates free fragments. The mechanisms of action appear to be different for induction of chromosome aberrations by MMC, ACT and the anthracyclines. Both in vitro and in vivo aberration induction has been reported. The associated phenomenon of sister chromatid exchanges is strongly potentiated by these agents.MMC has been a known mutagen in both akaryotes and eukaryotes, and it also increases the frequency of meiotic recombination. ACM-D does not appear to strongly effect these phenomena while sufficient data with the anthracyclines are not available, and perhaps, will turn out to be generally negative. MMC and ACM-D also induced somatic recombination and somatic mosaicism; the former is a potent recombinogen. Correlation between genetic effect and the information available on the binding of ACM-D to chromatin is discussed and some space has been allocated to the discussion of resistance and cross resistance induced by the four chemicals.The data point to a strong need for testing of chemicals for genetic toxicology; particular attention must be given to the subject of co-mutagenicity.

The effect of lysogenic induction with mitomycin C on the DNA and DNA polymerase of [formula omitted][formula omitted] K12λ

Mitomycin C (MMC), actinomycins (ACMs), daunomycin (DNM) and adriamycin (ADR) are antineoplastic, antibiotics, with the anthracyclines (DNM and ADR) being more promising that the others. All these chemicals interact with DNA. MMC brings about cross-linking of the DNA strands and acts as mono-, as well as, bifunctional alkylating agent. The other three intercalate with the DNA molecule. ACMs, especially ACM-D, inhibits RNA synthesis preferentially and discriminates between different species of RNA. At high concentrations, DNA, RNA and protein synthesis are all inhibited by any of these antibiotics. The agents effect the cell cycle traverse at various stages; MMC being most unspecific of all. Mitotic cells at post-G2 stages are the least effected by any of these chemicals.All these chemicals induce chromosome aberrations of chromatid type. The anthracyclines, in addition, produce chromosome-type aberrations perhaps by affecting the single stranded G1 chromosome. Rejoining occurs, but MMC creates a high frequency of quadriradial configurations by utilizing the analogous nucleotide sequences in the repetitive DNA. At other points, it mostly creates free fragments. The mechanisms of action appear to be different for induction of chromosome aberrations by MMC, ACT and the anthracyclines. Both in vitro and in vivo aberration induction has been reported. The associated phenomenon of sister chromatid exchanges is strongly potentiated by these agents.MMC has been a known mutagen in both akaryotes and eukaryotes, and it also increases the frequency of meiotic recombination. ACM-D does not appear to strongly effect these phenomena while sufficient data with the anthracyclines are not available, and perhaps, will turn out to be generally negative. MMC and ACM-D also induced somatic recombination and somatic mosaicism; the former is a potent recombinogen. Correlation between genetic effect and the information available on the binding of ACM-D to chromatin is discussed and some space has been allocated to the discussion of resistance and cross resistance induced by the four chemicals.The data point to a strong need for testing of chemicals for genetic toxicology; particular attention must be given to the subject of co-mutagenicity.