Types of DNA Repair, Mismatch, and Photoreactivation - Assignment Example

NAME) (COURSE NAME) Notes Assignment (15% Final Mark) (TUTOR) (DATE) 1. Compare and contrast the two types of DNA repair, mismatch and photoreactivation? (20 marks) There are numerous ways in which DNA may get damaged. This may occur due to cellular activities like replication and recombination or due to exogenous causes like exposure to mutagens. If the nucleotides are not repaired, it could lead to death of the cell Photoreactivation is a direct DNA system repair that helps in reversing a mutagenic event by reversing the pyrimidine dimer formation caused by UV light. It also removes methyl groups. In the process of the removal of the pyrimidine dimmers induced by UVm the photolyase enzyme recognizes the dimmers and binds to them splitting them in the presence of visible light with a range of between 300 and 500 mm. Mismatch DNA repair is essential for correction of any errors caused during DNA replication. Even though the replication of DNA is basically very accurate there are still chances that some errors might occur during the process replication or synthesis and nucleotide incorporation. When mistakes are made during the DNA replication process, they are supposed to be corrected during the proofreading phase. Any errors that escape without being detected during these processes are repaired via the mismatch repair process where the repair system has the capability of distinguishing between the parental DNA and the ones that are newly synthesized. This owes to the fact that parental DNA strands, unlike the newly synthesized ones, are methylated. However, hemimethylated DNA occurs when one of the strands is methylated while the other is not. DNA methylation results from the activity of dam methylase that is responsible for methylation of adenine bases in GATC sequence. Dam strains of E. coli have provided proof of the significance of the methylation process in the maintenance of bacterial DNA integrity. These strains have been found to increase the degree to which spontaneous mutation takes place. Mismatch repair system can accomplish its repair activities even from a distance. For instance, a hemimethylated site could be 1000 bp away and yet a mismatch system repair takes place to repair it. The mismatch repair system requires three basic genes (mutH, mutS and mutL) to provide the required products for the process. Generally, mismatch DNA repair system takes place in four phases. The first phase is where the mismatch is recognized by the proteins of mutS genes. The event that follows is a counter-measure where appropriate repair enzymes are recruited. Excision or removal of the incorrect sequence is the next phase and this is followed by the last phase where the DNA polymerase uses the parental strand as guide template to re-synthesize the nucleotide. 2. Give a general definition of the term ‘consensus nucleotide sequence’, and list five named prokaryotic/eukaryotic examples of such sequences, their location and functions. (20 marks) In bioinformatics and molecular biology, the term ‘consensus nucleotide sequence’ is used to imply the comparison of related sequences that results from multiple sequence alignment and serves the purpose of providing evidence or showing the residues that are most profuse in the alignment at every position. This pattern also serves an important purpose of showing where functional sequence motifs that are similar are found. In the contemporary fields molecular biology, genetics and other related fields the use of specially developed pattern recognition software has made recognition of sequence motifs little easier. Consensus nucleotide sequences are very important since particular sequence motifs may work as regulatory mechanisms for control or regulation of biosynthesis. In addition, the sequence motifs can function as signal sequences for regulation of molecule maturation or giving direction to a molecule. These sequences are conceived to be preserved for long periods of evolution due to the importance of their regulatory function. The amount of conservation of the consensus sequences sites can be used to estimate evolution relatedness. The consensus sequences (the conserved motifs) also help in revealing which residues are variable and which ones are conserved across the long periods of evolutionary process. For instance, in a DNA sequence of A[CT]N{A}, in the first A, an A will always be found in the position. The preceding notation [CT] implies that either C or T, N represents any base, and {A} implies that the base can be any base except A. Y stands for any pyrimidine while R represents any purine. The consensus sequence can also be represented graphically given that the above representation fails to indicate the frequency at which T or C occurs at [CT]. In graphical representation of the nucleotide sequence, the size of a given symbol gives a relative representation of the frequency of the occurrence of the nucleotide at a given position. The graphical sequence logos also give a relationship between the size of the symbol and the degree of conservation of residue. Larger symbols represent more conserved residue. Another example of the sequence is the intron found in the pre-mRNA in eukaryotes. These nucleotide sequence start with GT and end with AG. They are responsible for directing the intronic splicing mechanism to the appropriate donor and acceptor sites. Introns are actually regions in the DNA where the gene that contains the DNA is not translated into protein. Thus, the sections that are not coding are transcribed to the pre-mRNA. Prokaryotes also have the introns though not at the same locations as found in eukaryotes. In prokaryotes they are mainly found in rRNA and tRNA. They also preserve old codes that have since become inactive. In the kozak consensus nucleotide sequence, the mRNA of the eukaryote cell forms the main baseof the sequence. With a consensus sequence of (gcc)gccRccAUGG, the sequence helps in initiating the process of translation. Schizosaccharomyces pombe has a sequence motif, ATGACGTCA, recognized as M26 that is responsible for provision of recombination hotspot sites in the genome of the Schizosaccharomyces pombe (Steiner et al 2009). Lastly, the AUG consensus sequence found in plants plays a role in the initiation of the translation process. They are found in the translation initiation sites. 3. You have already studied several transcription factors in the lecture material. Using a named example of a transcription factor NOT previously discussed, describe how transcription factors regulate gene expression in eukaryotes? Examples should NOT include Gal4, steroid hormones, Myc/Max or CREB (20 marks) Transcription factors are very important DNA-binding proteins and help in regulating gene expression. In eukaryotes, a number of transcription factors work in conjunction with each other to fulfil this function. For example, the direct interaction between SP-1 and E2F help in delivering signals which activate the basic transcription machinery. However, the significance of the amount of dependencies varies between the background and the observed ones especially in human and rats (Tomovic 2009). E2F transcription factor plays another significant role in regulation of gene expression in the cell cycle of mammals. This takes place at G1/S phase transition. E2F transcription factors contribute to regulation of transcription and the promoters that provide sites for these processes are located in the genes that are responsible for synthesis of DNA. An example of such promoter is the DNA polymerase α. The promoters are also found in the genes that are responsible for control of cell cycle. Examples of genes that are responsible for cell cycle control and in which these promoters are found include cdc2 and B-myb. Intervention of E2F in the regulation of these genes results to differential expression in S and G0. This intervention causes high activity of the promoters at the G1/S phase border line while reducing promoter activity in the dormant cells. Despite the implication of the E2F transcription factors, some promoters have E2F sites that do not result to transcription activity that regulates growth. The reason why some sites regulate growth and others do not despite having the E2F transcription is still not well established. Nevertheless, a number of models have been produced to attempt in explain this. One of the model postulates that since E2F belongs to a group of transcription factors that characteristics of seven of the members have been unravelled. Of these, it is now known that functional E2F activity can be initiated when E2F1-5 hederodimerise with DP1 or DP2. In addition, not all the E2F’s are present at all stages of the cell cycle. For instance, though E2F1 is only present at certain cell cycle stages, E2F4 is present throughout the cell cycle stages. The activity of the E2F transcription factor is regulated by another family of proteins called the Rb proteins. To achieve this, the E2F1–3 specially binds Rb, while E2F4 and E2F5 principally bind p107 and p130. For this reason, there is possibility of having different compositions of the E2F protein complexes with respect to whether they regulate growth or not. The problem with this model is that it fails to account for the fact that deletion analysis of growth-regulated promoters points that even if E2F sites are kept intact, growth regulation can be eliminated if the dihydrofolate reductase promoter in mouse is deleted- especially deletion of sequences from -410 through -90. For this reason, and moreso because of the failure of the model, Farnham et al (1997), argue that: “..the fact that a particular E2F element can confer growth regulation in a given promoter, but not in a shorter version of the same promoter, suggests that subtle differences in the E2F site are not responsible for distinguishing growth-regulated versus non-growth-regulated promoters” (Farnham et al 18369) Another model is postulated on the fact that E2F bound pocket proteins interact with other upstream factors to achieve the growth-regulation function. That is, the proteins Rb, p107, and p130 have transcription domains and thus the model suggests that these transcription factors have a repression effect on the transcription activities of other factors. This model has supporting evidence from analysis of mutation activity in the E2F sites in some promoters like the B-Myb promoter. Bibliography: Paul R. van Ginkel, Kuang-Ming Hsiao, Hilde Schjerven and Peggy J. Farnham 1997 “E2F-mediated Growth Regulation Requires Transcription Factor Cooperation" Journal of Biological Chemistry 272: 18367-18374 Reef, R., Dunn S., Oren L. S., Eli D., Brickner S. I., Leggat W. & Ove H. 2009 “Photoreactivation is the main repair pathway for UV-induced DNA damage in coral planulae” Journal of Genetics, 182, 459-469 Steiner, W. W., Steiner, E. M., Girvin A. R. & Plewik L. E. 2009. “Novel Nucleotide Sequence Motifs That Produce Hotspots of Meiotic Recombination in Schizosaccharomyces pombe” Journal of Experimental Biology 212, 2760-2766 Tomovic, A., Stadler M., & Oakeley E. J, 2009 “Transcription factor site dependencies in human, mouse and rat genomes” Journal of Bioinformatics 10: 339. doi: 10.1186/1471-2105-10-339. Read More