DNA Replication & Mutations PDF

DNA REPLICATION 1  EUKARYOTIC REPLICATION Eukaryotic genomes are: Larger in size than prokaryotic: 6 billion base pairs Have 100,000 origin of replication Rate of replication is 100 nucleotides per second Have 14 DNA polymerases: pol α, pol β, pol...

DNA REPLICATION 1  EUKARYOTIC REPLICATION Eukaryotic genomes are: Larger in size than prokaryotic: 6 billion base pairs Have 100,000 origin of replication Rate of replication is 100 nucleotides per second Have 14 DNA polymerases: pol α, pol β, pol γ, pol δ, and pol ε Yeast has Autonomously Replicating Sequence (ARS) which are analogous to origin of rep. DNA bound to histones (proteins) to form structure nucleosomes. Chromatin: complex between DNA and proteins may undergo chemical modifications so that DNA may be able to slide off proteins or be accessible to enzymes of DNA replication machinery. At origin of replication, pre-replication complex is made with initiator proteins. 2 Other proteins are then recruited to start replication process.  PROKARYOTE VS EUKARYOTE REPLICATION A 3  TELOMERES Eukaryotic chromosomes are linear, not circular like prokaryotes. DNA pol enzyme can add nucleotides only in 5’ to 3’ direction. Leading strand synthesis continues until end of chromosome is reached Lagging strand DNA is synthesized in short stretches each initiated by separate primer There is no place for a primer to be made for DNA fragment to be copied at end of chromosome when replication fork reaches there. As a result, these ends remained unpaired and over time the ends may get shorter as cells continue to divide. Telomeres: ends of linear chromosomes which have repetitive sequences that code for no particular gene. Protect genes from getting deleted as cells continue to divide. 4  CHROMOSOME ANATOMY Chromosome arms project from either end of the centromere may be designated as short, which is abbreviated as p (petite) or long as q (because it follows p alphabetically). Can further specify location by the locus/loci (which is a section of the chromosome) 21p3 = locus 3 on short arm of chromosome 21 Sample problem: What does (5q2) mean? Long arm of chromosome 5 locus 2 5  TELOMERASE Telomerase: enzyme which contains a catalytic part and built-in RNA template. It attaches to the end of chromosome and complementary bases to RNA template are added on the 3’ end of the DNA strand. Once 3’ end of lagging strand template is sufficiently elongated, DNA pol can add nucleotides complementary to end of chromosomes. Thus, they get replicated. Active in germ cells and adult stem cells, not somatic cells. 6  AGING Cells that undergo cell division continue to have their telomere shortened Most somatic cell do not make enzyme telomerase Since they lack the built-in RNA template, then DNA pol cannot add nucleotides to end of chromosome. Telomere reactivation, through the addition of telomerase, may have the potential for treating age-related diseases in humans as shortening is directly associated with aging. Cancer is associated with uncontrolled cell division of abnormal cells. They accumulate mutations and can migrate to different body parts in process called metastasis. They have shortened telomeres, but active telomerase. If enzyme can be inhibited in just these cells, they can be stopped from dividing. 7 MUTATIONS 8  OVERVIEW Mutation: a permanent change in the DNA During the process of DNA Replication mistakes can occur. However, these mistakes do not necessarily result in a mutation. A mutation occurs whenever the change to the DNA becomes permanent. In other words, if it is not fixed. If the cell proceeds without spotting it and repairing it. If a mutation takes place in the repair genes, it may lead to cancer. 9  SPONTANEOUS Spontaneous: this type of mutation occurs naturally within the body. It is the result of DNA Polymerase adding the incorrect nitrogenous base pair during the process of Replication. The enzyme is efficient, but not perfect. A mistake can occur randomly. Most mistakes are corrected, but if they are not then the change is permanent and it can be passed down to daughter cells through DNA replication and cell division. 10  SPOTTING THE MISTAKE During the process of replication, DNA Polymerase III reads the newly added base before matching the next one in a process known as Proofreading If an incorrect base was detected, the enzyme DNA Polymerase II makes a cut at the phosphodiester bond and releases the wrong nucleotide with exonuclease action. If the error was missed during Replication there is another chance to catch it during: G2 Checkpoint: proper chromosome duplication is assessed Mismatch repair: errors corrected after replication is completed. The repair mechanism used to replace incorrect bases by making a cut on both 3’ and 5’ ends of the incorrect base is known as Nucleotide Excision Repair 11  INDUCED Mutations can also occur when the organism interacts with the environment. Induced: mutations caused as a result of exposure to environmental factors Mutagens: physical or chemical agents in the environment that can cause mutations Radiation: Gamma: most of it is sent back to outer space by the ozone layer X-Ray: dentist and radiographies. Use apron for safety. Ultraviolet: exposure to sunlight for long periods of time Radio waves: prolonged exposure to cell phones may be caused for concern Gas: smoking or second-hand smoking contain carcinogens Liquid: contaminated source of water Solid: pesticides eaten from not washing your fruits and vegetables 12  EFFECTS OF MUTATIONS Mutations are not automatically bad. The majority of mutations are neutral, which we call silent, as they do not result in a positive or negative change to the organism. Mutations could also be beneficial. They result in a change that is positive for the organism. They could increase the chances of survival for an organism in the form of an adaptation, which increases their fitness compared to other members of the species. If the mutation has a negative effect we call it detrimental or deleterious. It could result in death. Or it could lower an organism’s fitness. Overall, mutations increase genetic diversity, which is the driving force for evolution: the change of species over time. 13  CANCER If a mutation takes place in the repair genes, it may lead to cancer. If it affects tumor suppressor genes, the checkpoints of the cell cycle may become non-functional. Tumor suppressor genes: prevent cell from uncontrollable cell division. Retinoblastoma proteins: Rb, p21 and p53. A common mutation is for a Cdk (Cyclin-dependent kinase) to be activated without partnering with cyclin (positive regulator) Subsequent cell division will result in an accumulation of mistakes as the mechanisms designed to prevent and repair them fail. 14  TUMORS Tumor: a swelling of a part of the body caused by an abnormal growth of tissue. Benign: grow slowly and do not invade surrounding tissue. Malignant: fast growth which invades surrounding tissue through the mechanism of apoptosis. Metastasis: tumors which migrate to other parts of the body. Stage of Cancer Description 0 Abnormal cells with potential to turn cancerous. I Small and contained in one area. II & III Larger and spreading to nearby tissue. IV Has spread to other parts of the body 15  CAUSE OF DEATH Rapid cell division results in shortened telomeres. When telomeres run out, encoded information starts to be affected. However, cancer cells have enzyme telomerase active. As a result, they are able to synthesize new telomeres. Cancer cells interfere with essential organ functions. They can cell signals to surrounding cells to carry out apoptosis (programmed cell death) for space to propagate or to use their internals as an energy source. Over time it depletes the ability of organs such as the liver, lungs or brain to carry out their functions. Treatment involves: Surgery if it is at an accessible site Radiation if it is contained Chemotherapy indiscriminately destroys cancer or healthy cells Gene therapies are looking to identify and target cancer cells exclusively. 16  INHERITANCE Can mutations be passed down to the next generation? If a mutation occurs in a somatic (body) cell during the life of the individual it will not be passed down to the offspring. However, there could be a genetic pre-disposition if environment is similar. If the mutation occurs in a gamete: The gamete may randomly not be used for fertilization It may not result in the formation of a zygote It may result in premature death If the mutation is present in the germ-line cells then all gametes will have the mutation It may not manifest itself as detrimental in the offspring because it requires both copies of the homologous chromosome to be defective: Carrier & Recessive lethal 17  CHROMOSOMAL MUTATIONS OVERVIEW Chromosomes are divided into two categories: Autosomes: Body chromosomes In human these are chromosome pairs 1 through 22 Sex: chromosomes involved in sexual characteristics of the organism In humans these are chromosome pair 23 Includes the X and Y chromosomes Chromosome disorders divided into two categories: Abnormalities in chromosome number Aneuploidy: mutation which results in an individual with an error in chromosome number Polyploidy: mutation which results in an individual with extra chromosome sets Chromosomal structural rearrangements: Inversion: part of a chromosome is detached, rotated 180 degrees and then reinserted Translocation: a segment of a chromosome is removed and it reattaches in another location 18  IDENTIFICATION OF CHROMOSOMES Cytogenetics: branch of science that involves the isolation and microscopic observation of chromosomes to detect abnormalities in humans Karyotype: Number and appearance of chromosomes, including: length, banding pattern and centromere position. Place into a chart called a karyogram. 19  ABNORMAL CHROMOSOME NUMBERS NonDisJunction: during cell division chromosome pairs may fail to separate from each other This occurs during Anaphase of Meiosis I or II The M-Checkpoint was bypassed The tetrads or sister chromatids were not attached to the meiotic spindle Euploid: An individual with the correct number of chromosomes Aneuploid: An individual with either one less or one extra chromosome Monosomy: loss of a chromosome Fatal for most living things as genetic information is missing Trisomy: possessing an extra chromosome Most common viable birth involves Trisomy 21, which is Down syndrome. Individuals posses short stature, stunned digits, broad skull, large tongue and significant developmental delays. Its incidence directly correlates with maternal age. Polyploid: An individual with extra chromosome sets Very rare in the animal kingdom 20 Very advantageous for plants as it results in very large and robust organism  SEX CHROMOSOMES NON-DISJUNCTION Polyploidy could be a result of fertilization of abnormal diploid egg with normal haploid sperm yielding a triploid zygote If it occurs in animals or plants that carry out sexual reproduction with other individuals they will be sterile If it occurs in organisms that self-fertilize it can be passed down to offspring Humans display dramatic deleterious effects with autosomal monosomies and trisomies Variations in the number of sex chromosomes are associated with mild effects. X Inactivation: early in development, female mammalian embryos have 1 X chromosome inactivated by tight condensing into a quiescent (dormant) structure called Barr body. Individual carrying abnormal number of X chromosomes will inactivate all but one in each of her cells. Associated with sterility. The inactivated ones continue to express a few genes If X chromosome is absent, individual will not develop, but able to have just one X. XXY genotype is one type of Klinefelter syndrome, males develop with some female traits 21  CHROMOSOMAL INVERSIONS Chromosome inversion: part of a chromosome is detached, rotated 180 degrees and then reinserted Pericentric: type of chromosomal inversion that includes the centromere If inversion is asymmetric about the centromere, it can change the relative lengths of the chromosome arms, making these inversions easily identifiable. Paracentric: type of chromosomal inversion that occur outside of the centromere Humans and chimpanzees are separated by pericentric inversion in chromosome 18 and a few others. The other difference humans (46) and chimpanzees (48) have is humans underwent a fusion of 2 chromosomes. 22  CHROMOSOMAL INVERSIONS Unless it disrupts a gene sequence, inversions only change orientation of genes and are likely to have mild effects. They can result in functional changes because regulators of gene expression could be moved out of position with respect to target so can lead to overproduction. When one homologous chromosome undergoes inversion, but the other does not, the individual is called an inversion heterozygote. To maintain point-for-point synapsis during meiosis, one homolog must form a loop, and the other mold around it. It ensures they are aligned correctly, but it forces stretching and can be associated with imprecise synapsis. 23  TRANSLOCATION Translocation: when a segment of a chromosome is removed and it reattaches in another location. It could re-attach to the top or bottom of the same chromosome It could re-attach to the homologous chromosome pair. It could re-attach to a non-homologous chromosome. Reciprocal: an exchange of chromosome segments between 2 chromosomes which results in no gain or loss of genetic information. If the fragments are the same size and it occurs between homologous chromosomes there is no change. It will have the same effect as recombination for gametes. Fragments may not be same size, but could involve an exchange between homologous pair Fragments might be exchanged between non-homologous pairs They can be benign or have devastating effects depending on positions of genes with regards to their regulatory sequences. 24