1 MCB 502A-2015. Lecture #12 Genome logistics. Mutagenesis. DNA Damage DNA packing versus DNA condensation Recall that the four main genome functions are information maintenance, information expression, information replication and vertical information transfer, all embodied in the uniform chromosomal format of genomes (Lecture #4). The ~1,000-fold DNA compaction serves well the first two functions, but has to be augmented for the last two functions. It is evident that chromosomal DNA has to be completely decompacted and essentially naked for the purpose of replication. In contrast, the vertical information transfer function may require additional degree of DNA compression. This additional DNA compression comes as two very duifferent types: 1) DNA packing; 2) DNA condensation. DNA packing is employed when the final volume of the DNA-containing body is severely constrained, like phage capsid or sperm cell head. The objective here is to pack DNA as densely as physically possible in all three dimensions. This objective cannot be achieved without serious shielding of the huge overall DNA negative charge. In the phage capsids, the shielding comes from counterions, like Mg and spermidine. In the sperm head, the charge shielding is a function of small highly-basic proteins called protamines. The DNA compression in any one dimension appears modest in packed DNA, only ~300-fold, but it is equal in all three dimensions, so the actual 3D DNA compression equals cube of this number, ~3x10e7! DNA condensation pursues a different objective, which is to minimize length of already compacted DNA in two dimensions, for the purpose of bulk DNA movement within the cell. The obvious way to achieve this is to organize DNA in rosettes of radial loops, all originating from and returning back to the same point. As we have discussed before, this is exactly how chromosomes are ultimately organized both in prokaryotes and in eukaryotes. In fact, a modest DNA looping is the last stage of DNA compaction for the practical purpose of cellular metabolism (see the previous section). DNA condensation is promoted by specialized proteins called "condensins", circle-like proteins that can open up, encircle more than one individual DNA segment, and then close around them, bundling them up in loops. Condensins hold the eukaryotic chromosomal DNA together, making the chromosome occupy a particular space in the nucleus without intermingling with other chromosomes. What regulates the action of condensins in terms of timing and directionality is not known. Transcriptionally-inactive interphase heterochromatin is proposed to preserve the overall structure of mitotic chromosome, keeping its DNA as rosettes of radial loops in a spiral fashion along the chromosome core, with the total linear DNA compression level of around 10,000. For the purpose of chromosome segregation in eukaryotic cells, DNA condensation goes to the extreme. In preparation for mitosis, eukaryotic chromosomes undergo yet another round of condensation, so that the total linear DNA compression level reaches 50,000-fold. This additional condensation is still considered to be performed by condensins, although one would expect involvement of some "supercondensins" at this stage, operating with already condensed chromatin, rather than pure DNA. Such a degree of linear condensation is required to maximally shorten the chromosomes, to facilitate their efficient segregation by the mitotic spindle. As we will also see later, in addition to condensation, the replicated sister chromatids are held together through all the S-phase and G2 by condensin-like proteins called cohesins, which2 are removed at the beginning of anaphase, allowing sister chromatids to segregate from each other. Principles of bulk DNA transport The task of providing a newborn cell with its own genome complement demands an efficient way of transporting the genome-size DNA mass from its original position in the mother cell to the new designated position in the daughter cell. Three general principles of how to perform bulk DNA transport for significant distances are currently recognized: 1) pull (mitotic spindle in eukaryotes); 2) push (prokaryotic plasmids); 3) pump (naked DNA pumping, as during spore formation or filling up phage capsids). Although I have stated last time that the chromosomal segregation mechanisms in prokaryotes are still unknown, this is not completely true, because some bacterial chromosomes are derived from plasmids, and plasmid segregation mechanisms are actively studied. They are very different from the chromosome segregation mechanisms of eukaryotes, performed by mitotic spindle. The eukaryotic mechanisms segregate 105-fold condensed chromosomes by pulling them toward microtubule-organizing centers situated at the poles of the mother cell using a huge microtubule-based structure, called the mitotic spindle. The main features of the "pull" segregation principle are: 1) the spindle is based on microtubules and comprises from a few dozens to a few thousands of them, growing from the microtubule-organizing centers (MTOCs), which are clusters of microtubule polymerization buds (gamma-tubulin ring complexes) around centrioles; 2) chromosomes are attached via special structures, kinetochores, at the centromere, to the microtubules that will then pull them toward the pole; 3) pattern of microtubule interactions with the bivalents is such that sister chromatids are always segregated to the opposite poles; 4) the chromosomal DNA has to be maximally compressed in one dimension, so the length of the longest chromosomal arm in the karyotype is shorter than half the inter-centrosomal distance. By the way, this is the answer to the question about what is the most important denominator for the chromosome number in a eukaryotic karyotype — it is the inter-centrosomal distance! Thus, the eukaryotic DNA transport is dominated by the spindle that has dimensions of at least an order of magnitude longer than dimensions of average chromosomes segregated by it. In contrast, prokaryotic chromosome segregation mechanisms are so inconspicuous that there are serious arguments that there are none, and that bacterial chromosomes segregate by entropy. This argument is ludicrous, though, because bacterial chromosome segregation is precise in time and pattern. There are two possible examples of how prokaryotic segregation could be achieved in principle, illustrated by the
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