DNA Replication, Mutations, and DNA Repair [Molecular Biology 2 of 11]

Overview of DNA Replication, Mutations, and Repair

Introduction to DNA Structure and Replication

• The lecture covers DNA replication, mutations, and repair mechanisms. It emphasizes that DNA is double-stranded with a five prime (5') end and a three prime (3') end.

• During replication, the template strand is read in the 3' to 5' direction while the new strand is synthesized in the 5' to 3' direction.

• Both strands are replicated simultaneously; however, for simplicity, focus on one strand as the template during explanation.

Mechanisms of DNA Replication

• DNA replication occurs bidirectionally from an origin of replication. This process involves both continuous and discontinuous synthesis.

• The semi-conservative nature of replication means each new double helix consists of one original strand paired with one newly synthesized strand.

Key Enzymes Involved in Replication

• Helicase unwinds the double helix by separating the two strands, allowing access for other enzymes involved in replication.

• Topoisomerase I alleviates strain caused by unwinding by creating breaks in one strand to allow rotation around another unbroken strand.

Synthesis Process

• DNA polymerase synthesizes new strands by adding complementary bases to the template strand while moving from 5' to 3'.

• Chemotherapy drugs can target this mechanism by mimicking nucleotides but lacking a necessary hydroxyl group for further base addition.

Leading vs. Lagging Strand Synthesis

• The leading strand is synthesized continuously while the lagging strand is synthesized discontinuously through short segments called Okazaki fragments.

• RNA primers are required for initiating synthesis on both strands; multiple primers are used on the lagging strand due to its discontinuous nature.

Completion of Lagging Strand

• Exonuclease removes RNA primers after synthesis; gaps left behind are filled with nucleotides by DNA polymerase.

• Ligase seals any remaining gaps between nucleotide sequences ensuring continuity in the newly formed DNA strands.

Proofreading and Telomere Maintenance

Proofreading Mechanism

• DNA polymerase has proofreading capabilities that correct base pairing errors using its exonuclease activity.

Understanding Telomeres

• Telomeres consist of repetitive sequences at chromosome ends that protect genetic material during replication. They shorten with each cell division.

Role of Telomerase

• Telomerase adds repeat sequences back onto telomeres during replication, preventing loss of essential genetic information over time.

Mutations: Definition and Impact

Defining Mutations

• Mutations occur when changes in DNA sequence go undetected during proofreading or repair processes becoming part of a cell's permanent genome.

Consequences of Mutations

• A mutation can affect RNA transcription which may lead to defective proteins impacting cellular functions or causing inherited diseases.

Mechanisms for DNA Repair

Mismatch Repair Enzymes

• Mismatch repair enzymes detect structural distortions caused by mismatched bases and remove incorrect sequences before filling them correctly.

Base Excision Repair

• This mechanism identifies single incorrect bases using specific glycosylases that remove damaged bases followed by filling gaps with correct ones via ligation.

Nucleotide Excision Repair

• Nucleotide excision repair addresses bulky distortions like those caused by UV light damage through removal and replacement processes involving endonucleases.

Direct Repair Mechanism

• MGMT repairs methylated guanine bases caused by alkylating agents through direct removal of methyl groups restoring normal base structure.

The Role of Methylation in DNA Damage and Cancer

Methylation and DNA Repair Mechanisms

• Alkylating agents, such as carcinogens, add methyl groups to DNA, which can damage it and potentially turn off important genes involved in DNA repair mechanisms.

• If a gene responsible for apoptosis is turned off due to methylation, cells may continue to divide uncontrollably, leading to cancer development.

• Direct damage to DNA from alkylating agents can hinder replication and cell division processes.

Chemotherapy and MGMT Function

• Chemotherapy uses alkylating agents to target rapidly dividing cancer cells by damaging their DNA; this exploits the fact that cancer cells often skip repair mechanisms.

• MGMT (O6-methylguanine-DNA methyltransferase) repairs damaged DNA by removing added methyl groups; however, its levels are finite within the cell.

• Continuous chemotherapy dosing is necessary until MGMT is depleted so that the treatment can effectively kill cancer cells.

Double Strand Break Repair Mechanisms

• Non-homologous end joining (NHEJ) repairs double strand breaks by bringing together two fragmented ends of DNA without requiring perfect base pairing.

• This process may result in loss of some genetic material during repair but focuses on rejoining fragments quickly rather than accurately.

Clinical Implications of Defective DNA Repair

Xeroderma Pigmentosum

• A condition caused by defective nucleotide excision repair leads to increased risk for skin lesions and various skin cancers like basal cell carcinoma.

Lynch Syndrome

• Also known as hereditary nonpolyposis colorectal cancer (HNPCC), this syndrome results from defective mismatch repair, increasing risks for early-onset colorectal and ovarian cancers.

Ataxia Telangiectasia

• Caused by mutations in the ATM gene affecting NHEJ, patients experience neurological symptoms alongside immunodeficiencies and heightened cancer risk.

Fanconi Anemia

• Another disorder linked with defective NHEJ involves different enzymes affecting hematological function, leading to progressive anemia and congenital abnormalities.

Bloom Syndrome

• Resulting from mutated helicase impacting both replication and double strand break repair; patients face short stature, facial rashes, immunodeficiency, and increased risk for various cancers.