Chromosome segregation in Eubacteria

It is now clear that bacterial chromosomes rapidly separate in a manner independent of cell elongation, suggesting the existence of a mitotic apparatus in bacteria. Recent studies of bacterial cells reveal filamentous structures similar to the eukaryotic cytoskeleton, proteins that mediate polar chromosome anchoring during Bacillus subtilis sporulation, and SMC interacting proteins that are involved in chromosome condensation. A picture is thereby developing of how bacterial chromosomes are organized within the cell, how they are separated following duplication, and how these processes are coordinated with the cell cycle.     

Refined radiation hybrid map of mouse Chromosome 17

We have made a radiation hybrid map of mouse Chromosome (Chr) 17 with 75 microsatellite markers, including those from McCarthy et al. (Genome Res 7, 1153–1161, 1997). Seventy-four of the markers are linked at LOD > 9, and all link at LOD > 5. A LOD 3 framework of 18 markers was used to construct a placement map. The order obtained is in good agreement with genetic maps, and distance estimates give an idea of how recombination rates vary across the chromosome. Recombination is remarkably low with respect to RH break frequency in the region from the centromere to the end of H2. This is similar in interspecific and intersubspecific crosses despite the inversion of a substantial part of this region in Mus spretus with respect to Mus musculus. Received: 10 April 1998 / Accepted: 18 June 1998     

Chromosome organization and segregation in bacteria

In the recent years, considerable advances have been made towards understanding the structure and function of the bacterial chromosome. A number of different factors appear to cooperate in condensing DNA into a highly dynamic assembly of supercoiled loops. Despite this variability in the lower levels of chromatin structure, the global arrangement of chromosomal DNA within the cell is surprisingly conserved, with loci being arrayed along the cellular long axis in line with their order on the genomic map. This conserved pattern is propagated during the course of DNA segregation. First, after entry into S-phase, the newly synthesized origin regions are segregated in an active and directed process, involving the bacterial actin homolog MreB. Subsequent DNA segments then follow by different mechanisms. They are separated immediately after release from the replisome and move rapidly to their conserved positions in the incipient daughter cell compartments. Partitioning of the bacterial chromosome thus takes place while DNA replication is in progress.     

Chromosome segregation and genomic stability

The acquisition of genomic instability is a crucial step in the development of human cancer. Genomic instability has multiple causes of which chromosomal instability (CIN) and microsatellite instability (MIN) have received the most attention. Whereas the connection between a MIN phenotype and cancer is now proven, the argument that CIN causes cancer remains circumstantial. Nonetheless, the ubiquity of aneuploidy in human cancers, particularly solid tumors, suggests a fundamental link between errors in chromosome segregation and tumorigenesis. Current research in the field is focused on elucidating the molecular basis of CIN, including the possible roles of defects in the spindle checkpoint and other regulators of mitosis.     

Chromosome cohesion and segregation in mitosis and meiosis

The faithful segregation of the genetic material into daughter cells during cell division is crucial for the production of healthy progeny. Sister chromatid cohesion and separation are fundamental to this process. Progress has been made in our molecular understanding of cohesion and mechanisms for the dissolution of cohesion have been uncovered.     

Detailed chromosomal and radiation hybrid mapping in the proximal part of rat Chromosome 10 and gene order comparison with mouse and human

The rat provides valuable and sometimes unique models of human complex diseases. To fully exploit the rat models in biomedical research, it is important to have access to detailed knowledge of the rat genome organization as well as its relation to the human genome. Rat Chromosome 10 (RNO10) harbors several important cancer-related genes. Deletions in the proximal part of RNO10 were repeatedly found in a rat model for endometrial cancer. To identify functional and positional candidate genes in the affected region, we used radiation hybrid (RH) mapping and single- and dual-color fluorescence in situ hybridization (FISH) techniques to construct a detailed chromosomal map of the proximal part of RNO10. The regional localization of 14 genes, most of them cancer-related (Grin2a, Gspt1, Crebbp, Gfer, Tsc2, Tpsb1, Il9r, Il4, Irf1, Csf2, Sparc, Tp53, Thra1, Gh1), and of five microsatellite markers (D10Mit10, D10Rat42, D10Rat50, D10Rat72, and D10Rat165) was determined on RNO10. For a fifteenth gene, Ppm1b, which had previously been assigned to RNO10, the map position was corrected to RNO6q12-q13.     

Identification and characterization of oxidation and deamidation sites in monoclonal rat/mouse hybrid antibodies

Oxidation of methionine residues and deamidation of asparagine residues are the major causes of chemical degradation of biological pharmaceuticals. The mechanism of these non-enzymatic chemical reactions has been studied in great detail. However, the identification and quantification of oxidation and deamidation sites in a given protein still remains a challenge. In this study, we identified and characterized several oxidation and deamidation sites in a rat/mouse hybrid antibody. We evaluated the effects of the sample preparation on oxidation and deamidation levels and optimized the peptide mapping method to minimize oxidation and deamidation artifacts. Out of a total number of 18 methionine residues, we identified six methionine residues most susceptible to oxidation. We determined the oxidation rate of the six methionine residues using 0.05% H₂O₂ at different temperatures. Methionine residue 256 of the mouse heavy chain showed the fastest rate of oxidation under those conditions with a half life of approximately 200 min at 4 °C and 27 min at 37 °C. We identified five asparagine residues prone to deamidation under accelerated conditions of pH 8.6 at 37 °C. Kinetic characterization of the deamidation sites showed that asparagine residue 218 of the rat heavy chain exhibited the fastest rate of deamidation with a half live of 1.5 days at pH 8.6 and 37 °C. Analysis of antibody isoforms using free flow electrophoresis showed that deamidation is the major cause of the charged variants of this rat/mouse hybrid antibody.     

Chromosome segregation: playing polo in prophase

During mitosis, in most eukaryotes, cohesin is removed from chromosomes in two steps. A paper in the March issue of Molecular Cell identifies Polo-like kinase as a key regulator for the first step that releases much of cohesin during prophase.     

Phenotypic expression of malignancy in hybrid and cybrid mouse cells

This study has been directed toward the effect of cytoplasmic transfer on the expression of marker properties in hybrid cell systems. Conventional hybrids between two nucleated cells were constructed between tumorigenic and nontumorigenic cells. Cytoplasmic hybrids, or cybrids, were constructed between enucleated chloramphenicol resistant (CAP R) donor cells (cytoplasts) and nucleated recipient cells. Clear-cut evidence for the cytoplasmic transmission of CAP resistance was obtained. Although cytoplasmic transfer had no effect on tumorigenicity or growth in soft agar, preliminary evidence was found that saturation density of the recipient cells could be altered by cytoplasmic addition in cybrids.     

Chromosome length controls mitotic chromosome segregation in yeast

We have examined the effect of physical length on the mitotic segregation of artificial chromosomes and fragments of natural yeast chromosomes. Increasing the length of artificial chromosomes decreases the rate at which they are lost during mitosis. We have made fragments of chromosome III by integrating new telomeres at different positions along the length of the chromosome. Chromosome fragments of 42 and 72 kb behave like artificial chromosomes: they are lost in mitosis much more frequently than natural chromosomes. In contrast, a chromosome fragment of 150 kb is as mitotically stable as the full-length chromosome from which it is derived. The structural instability of a short dicentric artificial chromosome demonstrates that, although short artificial chromosomes segregate poorly in mitosis, they do attach to the mitotic spindle. We discuss these results in the context of a model in which chromosome segregation is directed by the intercatenation of the segregating DNA molecules.     

Chromosome segregation in Escherichia coli B/r at various growth rates.

Chromosome segregation was analyzed in three substrains of Escherichia coli B/r growing at various rates. The cultures were pulse labeled with [14C]thymidine and bound to the bottom surface of a nitrocellulose membrane filter, and the radioactivity in newborn cells released from the surface during continuous elution with growth medium was measured. Since there was a fixed orientation in the release of newborn cells, the time course of the change in radioactivity per effluent cell could be used to investigate the orientation of chromosome segregation. If the radioactive deoxyribonucleic acid strands were partitioned at random between the progenies remaining attached to the membrane filter and those released into the effluent, the radioactivity per cell would decrease twofold after each generation of elution. The decrease in radioactivity was less than twofold at C + D min of elution and larger than twofold one generation later, indicating that chromosome segregation was nonrandom.     

Chromosome strand segregation during sporulation in Bacillus subtilis

After the initiation of spore formation in Bacillus subtilis, the products of the final round of DNA replication segregate into two cells, i.e. the prespore and the mother cell. The prespore, which is known to contain a single completed chromosome, develops into a mature endospore which can be readily separated from mother cells and non-sporulating cells on the basis of its resistance properties. We have used a procedure originally developed to label the terminus region of the B. subtilis chromosome to specifically label the newly synthesized strands of DNA during the final round of DNA replication before sporulation. We have purified prespore DNA and used strand-specific probes to measure the radioactivity incorporated. The results show that the sister chromosomes segregate at random into the prespore. This result has implications for the segregation of chromosomes during vegetative growth and for the generation of cellular asymmetry during sporulation.     

Chromosome segregation: keeping kinetochores in the loop

The Ndc80 complex is a key component of the kinetochore-microtubule interface. Two studies now demonstrate that a conserved loop region within the extended coiled-coil of Ndc80 plays an unexpected role in recruiting proteins to the kinetochore.     

Two distinct types of mitochondrial DNA segregation in mouse-rat hybrid cells. Stochastic segregation and chromosome-dependent segregation

Two distinct patterns of mitochondrial DNA (mtDNA) segregation were found in different mouse-rat hybrid cell lines. On mouse-rat hybrid cell line, H2, retained complete sets of chromosomes and mtDNAs of both mouse and rat. Even after cultivation for about one year after cloning, the H2 cell population still retained both parental mtDNAs. However, when mtDNAs of H2 subclones were examined, it was found that some individual cells in the H2 cell population contained only mouse or only rat mtDNA, although they still retained complete sets of both kinds of parental chromosomes. This type of mtDNA segregation, named stochastic segregation, is bidirectional and may be caused by the repetition of random sharing of mouse and rat mtDNAs with daughter cells. This segregation occurred spontaneously during long-term cultivation. The second type of mtDNA segregation, named chromosome-dependent segregation, was found in the other mouse-rat hybrid cell lines that segregated either mouse or rat chromosomes. In these hybrid cells, chromosomes and mtDNA of the same species co-segregated. This second type of segregation is unidirectional. The types of mtDNA segregation appear to depend on the stability of the parental chromosomes in the hybrid cells. When both mouse and rat chromosomes retain stably, mtDNA shows stochastic segregation. On the contrary, when either species of chromosomes is segregated from the cells, mtDNA shows chromosome-dependent segregation.     

Chromosome replication, nucleoid segregation and cell division in archaea

Recent progress in cell cycle analysis of archaea has included the identification of putative chromosome replication origins, novel DNA polymerases and an unusual mode of cell cycle organization featuring multiple copies of the chromosome and asymmetric cell divisions. Genome sequence data indicate that in crenarchaea, the 'ubiquitous' FtsZ/MinD-based prokaryotic cell division apparatus is absent and division therefore must occur by unique, as-yet-unidentified mechanisms. The evolutionary and functional relationships between the archaeal Cdc6 protein and bacterial and eukaryal replication initiation factors are discussed.