1 Chromosomes

1 CHROMOSOMES Metaphase chromosome In interphase cells stained for light microscopy, the chromatin usually appears as a diffuse mass within the nucleus, suggesting that the chromatin is highly extended. As a cell prepares for mitosis, its chromatin coils and folds up (condenses), eventually forming a characteristic number of short, thick metaphase chromosomes that are distinguishable from each other with the light microscope. Though interphase chromatin is generally much less condensed than the chromatin of mitotic chromosomes, it shows several of the same levels of higher-order packing. Even during interphase, the centromeres and telomeres of chromosomes, as well as other chromosomal regions in some cells, exist in a highly condensed state similar to that seen in a metaphase chromosome. This type of interphase chromatin, visible as irregular clumps with a light microscope, is called heterochromatin, to distinguish it from the less compacted, more dispersed euchromatin (“true chromatin”). Because of its compaction, heterochromatic DNA is largely inaccessible to the machinery in the cell responsible for transcribing the genetic information coded in the DNA, a crucial early step in gene expression. In contrast, the looser packing of euchromatin makes its DNA accessible to this machinery, so the genes present in euchromatin can be transcribed. The chromosome is a dynamic structure that is condensed, loosened, modified, and remodeled as necessary for various cell processes, including mitosis, meiosis, and gene activity. Figure 1. Classification of chromosomes Figure 2. Human metaphase chromosomes with regard to replacement of centromer Experiment: Examine the metaphase chromosome of mammalian cell in prepared microscope slide. Polytene chromosomes Polytene chromosomes (giant chromosomes) are oversized chromosomes which have developed from standard chromosomes. Specialized cells undergo repeated rounds of DNA replication without cell division (endomitosis), to increase cell volume, forming a giant polytene chromosome. Polytene chromosomes form when multiple rounds of replication produce many sister chromatids that remain fused together. In addition to increasing the volume of the cells' nuclei and causing cell expansion, polytene cells may also have a metabolic advantage as multiple copies of genes permits a high level of gene expression. In Drosophila melanogaster, for example, the chromosomes of the larval salivary glands undergo 2 many rounds of endoreduplication, to produce large amounts of glue before pupation. Another example within the organism itself is the tandem duplication of various polytene bands located near the centromere of the X chromosome which results in the Bar phenotype of kidney-shaped eyes. Polytene chromosomes have characteristic light and dark banding patterns that can be used to identify chromosomal rearrangements and deletions. Dark banding (band) frequently corresponds to inactive chromatin, whereas light banding (interband) is usually found at areas with higher transcriptional activity. Polytene chromosomes were originally observed in the larval salivary glands of Chironomus. Figure 3. Polytene chromosomes from larval salivary gland of Chironomus Experiment: Observe the Polytene chromosomes of salivary gland of Chironomus. Zoom in band and interband parts of the chromosomes. Compare the size of regular metaphase chromosomes with polytene chromosomes. MITOSIS When a cell is not dividing, and even as it replicates its DNA in preparation for cell division, each chromosome is in the form of a long, thin chromatin fiber. After DNA replication, however, the chromosomes condense as a part of cell division: Each chromatin fiber becomes densely coiled and folded, making the chromosomes much shorter and so thick that we can see them with a light microscope. Each duplicated chromosome has two sister chromatids, which are joined copies of the original chromosome. The two chromatids, each containing an identical DNA molecule, are initially attached all along their lengths by protein complexes called cohesins; this attachment is known as sister chromatid cohesion. Each sister chromatid has a centromere, a region of the chromosomal DNA where the chromatid is attached most closely to its sister chromatid. This attachment is mediated by proteins bound to the centromeric DNA; other bound proteins condense the DNA, giving the duplicated chromosome a narrow “waist.” The portion of a chromatid to either side of the centromere is referred to as an arm of the chromatid. (An unduplicated chromosome has a single centromere, distinguished by the proteins that bind there, and two arms.) Later in the cell division process, the two sister chromatids of each duplicated chromosome separate and move into two new nuclei, one forming at each end of the cell. Once the sister chromatids separate, they are no longer called sister chromatids but are considered individual chromosomes; this step essentially doubles the number of chromosomes in the cell. Thus, each new nucleus receives a collection of chromosomes identical to that of the parent cell. Mitosis, the Division of the genetic material in the nucleus, is usually followed immediately by cytokinesis, the division of the cytoplasm. One cell has become two, each the genetic equivalent of the parent cell. From a fertilized egg, mitosis and cytokinesis produced the 200 trillion somatic cells that now make up your body, and the 3 same processes continue to generate new cells to replace dead and damaged ones. In contrast, you produce gametes—eggs or sperm—by a variation of cell division called meiosis, which yields daughter cells with only one set of chromosomes, half as many chromosomes as the parent cell. Meiosis in humans occurs only in special cells in the ovaries or testes (the gonads). Generating gametes, meiosis reduces the chromosome number from 46 (two sets) to 23 (one set). Fertilization fuses two gametes together and returns the chromosome number to 46 (two sets). Mitosis then conserves that number in every somatic cell nucleus of the new human individual. Phases of the Cell Cycle Mitosis is just one part of the cell cycle. In fact, the mitotic (M) phase, which includes both mitosis and cytokinesis, is usually the shortest part of the cell cycle. The mitotic phase alternates with a much longer stage called interphase, which often accounts for about 90% of the cycle. Interphase can be divided into subphases: the G1 phase (“first gap”), the S phase (“synthesis”), and the G2 phase (“second gap”). The G phases were misnamed as “gaps” when they were first observed because the cells appeared inactive, but we now know that intense metabolic activity and growth occur throughout interphase. During all three subphases of interphase, in fact, a cell grows by producing proteins and cytoplasmic organelles such as mitochondria and endoplasmic reticulum. Duplication of the chromosomes, crucial for eventual division of the cell, occurs entirely during the S phase. Thus, a cell grows (G1), continues to grow as it copies its chromosomes (S), grows more as it completes preparations for cell division (G2), and divides (M). The daughter cells may then repeat the cycle. A particular human cell might undergo one division in 24 hours. Of this time, the M phase would occupy less than 1 hour, while the S phase might occupy about 10–12 hours, or about half the cycle. The rest of the time would be apportioned between the G1 and G2 phases. The G2 phase usually takes 4–6 hours; in our example, G1 would occupy about 5–6 hours. G1 is the most variable in length in different types of cells. Some cells in a multicellular organism divide very infrequently or not at all. These cells spend their time in G1 (or a related phase called G0) doing their job in the organism—a nerve cell carries impulses, for example. Mitosis is conventionally broken down into five stages: prophase, prometaphase, metaphase, anaphase, and telophase. Overlapping with the latter stages of mitosis, cytokinesis completes the mitotic phase. Many of the events of mitosis depend on the mitotic spindle, which begins to form in the cytoplasm during prophase. This structure consists of fibers made of microtubules and Figure 4. Mitosis in a plant cell. These light micrographs show mitosis in cells of an onion root. 4 Experiment: Take a small amount of brewers yeast solution by using Pasteur pipette. Put one small drop of brewers yeast solution onto microscope slide. Examine the amitosis of yeast cells. Experiment: Observe the mitosis in onion root cells in a prepared slide. MEIOSIS Prophase I Leptoten: Leptotene is of very short duration and progressive condensation and coiling of chromosome fibers takes place. Individual chromosomes—each consisting of two sister chromatids—become discrete. The two sister chromatids closely associate and are visually indistinguishable from one another. Zygoten: Occurs as the chromosomes approximately line up with each other into homologous chromosome pairs. At this stage, the synapsis (pairing/coming together) of homologous chromosomes takes place, facilitated by assembly of central element of the synaptonemal complex. The paired chromosomes are called bivalent or tetrad chromosomes. Pacytene: At this point a tetrad of the chromosomes has formed known as a bivalent. This is the stage when chromosomal crossover (crossing over) occurs. Nonsister chromatids of homologous chromosomes may exchange segments over regions of homology. Sex chromosomes, however, are not wholly identical, and only exchange information over

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