Name: repairing dna double-strand breaks
Uploaded: 2023-04-30T18:28:49.000Z
Duration: 2 min 4 s
Channel: JoVE

K12 Standards

north-carolina-k-12-science-standard-course-of-study

NC K-12 Science Standard Course of Study

9-12

Biology

heredity-inheritance-and-variation-of-traits

Heredity- Inheritance and Variation of Traits

LS.Bio.6.2

Fixing Double-strand Breaks

When both strands of DNA are damaged, there is no intact template left for accurate repair but if left unrepaired, this scenario can lead to cell death.&#160;
There are two mechanisms to repair double-strand breaks. The first type, non-homologous end joining, permits joining of ends even if there is no sequence similarity between them - and takes place before DNA duplication when the DNA needs quick repair.&#160;
In mammalian cells, this is carried out by the DNA end-binding heterodimeric protein Ku which forms a complex with the catalytic subunits of DNA-dependent protein kinase.&#160;
This complex holds the broken chromosome ends in place while a DNA polymerase inserts nucleotides to bridge the gap between these ends. Next, DNA ligase IV forms a complex with its cofactor XRCC and another protein called XLF, and rejoins and seals these ends.
Quick fixes like these can lead to mutations at the repair site, or genomic rearrangements including deletions, translocations of genetic material, and fusions which may result in chromosomes with two centromeres, or lacking centromeres altogether.&#160;
Mutations are widespread, and human somatic cells can tolerate as many as 2000 of these. Genomic rearrangements, on the other hand, are rare - but can be found in cancerous cells.&#160;
Most DNA double-strand breaks lead to single-stranded overhangs and the second repair type, homologous recombination, fixes these breaks. This form of recombination is much more accurate than non-homologous end joining and requires DNA from a sister chromatid as a template&#8211; so it typically occurs after gene duplication during cell division.

fixing double-strand breaks

The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages both copies of genetic information but also disrupts the continuity of DNA, making the chromosome fragile.
In a cell, there are an estimated ten double-strand breaks (DSBs) per day. The primary source of damage is metabolic by-products such as Reactive Oxygen Species and environmental factors such as ionizing radiations. Although less common, malfunctioning nuclear enzymes can also cause DSBs. Failure of enzymes like type II topoisomerases, which cut both strands of DNA and rejoin them while disentangling chromosomes, can inadvertently result in DSBs. Mechanical stress on the DNA duplex can also lead to DSBs. In prokaryotes, prolonged desiccation strains DNA, causing DSBs.
Of the two mechanisms for DNA repair, homologous recombination depends on a sister chromatid being nearby, which happens during the S and G2 phases. Due to this restriction, in the absence of a homology donor, cells have to resort to Nonhomologous end joining (NHEJ), even though it is much less accurate. It has been hypothesized that the reason higher eukaryotes can afford to preferentially utilize NHEJ for DSB repairs is that they have abundant non-coding DNA, which permits nucleotide substitutions, deletions or additions without grievous consequences.

The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the ...

Education

JoVE Core

Cell Biology

Unit 1

DNA Replication and Repair

KEY TERMS AND CONCEPTS

repairing dna double-strand breaks

Repairing DNA double-strand breaks is an important part of DNA maintenance. A double-strand break happens when both strands of the DNA molecule are cut. Cells must fix this damage to keep genetic information stable.

The repair process can happen by more than one pathway. Some breaks are repaired by direct joining of the broken ends. Other breaks are repaired by using a matching DNA sequence as a template. These pathways help cells respond to damage in different ways.

This topic is closely linked to genetic recombination and DNA repair. It shows how cells handle serious DNA damage and restore the chromosome structure. Understanding these repair steps helps explain how cells protect their genomes.

Repairing DNA Double-Strand Breaks

base-pairing and dna repair

DNA Pairing Rules in Repair

DNA pairing rules help guide DNA repair. In DNA, bases pair in a specific way, and those matching pairs support accurate copying and fixing of the molecule. When DNA is damaged or copied incorrectly, repair systems can use the normal base-pairing pattern as a reference. This helps cells find mismatched bases and restore the correct sequence. The process depends on the idea that each base has a preferred partner. Understanding base-pairing is important because it explains how cells protect...

the dna replication fork

DNA Replication Fork in Action

The DNA replication fork is the active region where DNA copies itself. It forms when the two strands of the double helix separate so each strand can serve as a template for a new partner strand. At the fork, the copying process moves in a set direction. The two strands are built in different ways as replication continues, so the fork helps organize how the new DNA is made. The replication fork is a key part of accurate DNA duplication. It shows how the cell opens the double helix, copies the...

lagging strand synthesis

Okazaki Fragments on the Lagging Strand

Lagging strand synthesis builds new DNA in short pieces called Okazaki fragments. This happens during DNA replication because the lagging strand runs in the opposite direction from the direction the replication machinery moves. As the DNA double helix opens, DNA polymerase adds nucleotides only in the 5' to 3' direction. On the lagging strand, this forces the cell to copy the template in segments instead of one continuous stretch. Each segment begins near the replication fork and extends away...

the replisome

Replisome Components in DNA Copying

The replisome is the protein machine that copies DNA during cell division. It works at the replication fork, where the double helix opens so new strands can be made. DNA polymerase is the core enzyme in the replisome. It adds nucleotides to the growing DNA strand. In bacteria, two DNA polymerases work together at the fork. These enzymes are linked to the helicase by a clamp-like protein so they stay attached to the DNA. Helicase unwinds the DNA ahead of the fork. Single-stranded binding...

proofreading

DNA Polymerase Error Correction

DNA polymerase uses proofreading to help correct errors during DNA replication. As it builds the daughter strand, the enzyme adds nucleotides that match the template DNA strand. This matching helps keep DNA copying accurate and supports replication fidelity. DNA is made in the 5' to 3' direction, and several DNA polymerases help with this process. In nuclear DNA replication, DNA polymerase delta and DNA polymerase epsilon have proofreading ability. After a new base is added, these polymerases...

replication in prokaryotes

How Prokaryotic DNA Copies Itself

Prokaryotic DNA copies itself through a set of coordinated steps called initiation, elongation, and termination. In bacteria, replication starts when initiator proteins bind to a single origin of replication, or ori, on the circular chromosome. The DNA is then copied in both directions until two DNA molecules are made. Several proteins help open and stabilize the DNA during replication. Topoisomerase cuts one side of the double-stranded phosphate-sugar backbone so the helix can unwind faster.

replication in eukaryotes

Eukaryotic DNA Replication Steps

Eukaryotic DNA replication copies the genome before cell division. It happens in a tightly regulated way so each daughter cell gets a complete set of chromosomes in the nucleus. The process has three stages: initiation, elongation, and termination. Eukaryotic cells use many proteins to manage replication because their chromosomes are linear and their genomes are larger than those of prokaryotes. Replication begins at multiple origins of replication along each chromosome. Initiator proteins...

telomeres and telomerase

Telomeres and Telomerase in DNA Replication

Telomeres and telomerase help protect chromosomes during DNA replication. Telomeres are repeated DNA sequences at chromosome ends, and telomerase is the enzyme that can extend them. Together, they help keep important genetic information from being lost when cells divide. Each time a chromosome is copied, the very end of the DNA strand can be difficult to replicate fully. Telomeres act like a buffer zone at the tips of chromosomes. Telomerase adds new repeats to these ends, which helps maintain...

overview of dna repair

DNA Repair and Genetic Recombination

DNA repair and genetic recombination help cells respond to damage in DNA. DNA can be harmed by normal cell activity or by outside factors. When damage is not fixed, it can affect how genetic information is copied and used. Cells use repair pathways to detect and correct many kinds of DNA damage. Some repair steps remove damaged bases or replace missing pieces of DNA. Other pathways help keep the DNA sequence stable after damage has occurred. Genetic recombination is also part of this process.

base excision repair

Base Excision Repair Pathway

Base excision repair is a DNA repair pathway that fixes small base damage. It helps cells correct changes that can appear in DNA before they cause larger problems. In this pathway, a damaged base is removed first. The cell then processes the site and replaces the missing part with the correct DNA building block. This repair step helps keep genetic information accurate. Base excision repair is one of the main ways cells handle everyday DNA damage. It is an important part of maintaining DNA...

nucleotide excision repair

UV Damage and DNA Repair by NER

UV damage can bend DNA and block replication or transcription. Cells often face mutagens, which are outside factors that damage DNA and can create mutations. If these changes are not repaired, they can lead to disease or cancer, depending on which DNA sequences are affected. Nucleotide excision repair, or NER, is the pathway that helps cells find and remove damaged DNA. In prokaryotes, three proteins carry out this work: UvrA, UvrB, and UvrC. UvrA and UvrB move along the DNA together and look...

mismatch repair

DNA Mismatch Repair and Cancer Risk

DNA mismatch repair fixes copying errors that happen during DNA replication. It helps cells find the wrong nucleotide, remove it, and replace it with the correct one before the cell divides. Human cells carry more than 3 billion base pairs of DNA, so replication must be very accurate. Even with DNA polymerase proofreading, a copying error can still occur about once every 1 million base pairs. One common error is a mismatch, such as A paired with G or T paired with C. The Mutator protein family...

dna damage can stall the cell cycle

DNA Repair Checkpoints in the Cell Cycle

DNA repair checkpoints help cells pause the cell cycle after DNA damage. This pause gives the cell time to check for breaks and repair them before division continues. Cells must do this at critical stages of the cycle. The amount of DNA in the cell depends on when the pause happens. If the cycle stops before DNA replication, the cells will contain twice the normal amount of DNA. If cells stop after DNA replication but before mitosis, they will contain four times the normal amount. Cells use...

Repairing DNA double-strand breaks is an important part of DNA maintenance. A double-strand break happens when both strands of the DNA molecule are cut. Cells must fix this damage to keep genetic information stable. The repair process can happen by more than one pathway. Some breaks are repaired by direct joining of the broken ends. Other breaks are repaired by using a matching DNA sequence as a template. These pathways help cells respond to damage in different ways. This topic is closely...

homologous recombination

Homologous Recombination in DNA Repair

Homologous recombination is a DNA repair process that uses a matching DNA sequence as a template. It helps cells fix breaks in DNA and keep genetic information accurate. The process begins when a broken DNA strand finds a similar, or homologous, piece of DNA. That matching sequence guides repair and allows the damaged DNA to be restored. Because the repair uses a copied sequence, the final DNA can closely match the original information. Homologous recombination is also important in genetic...

gene conversion

Gene Conversion During DNA Repair

Gene conversion is a DNA repair event linked to genetic recombination. It can change the sequence of one DNA region by using a related DNA strand as a template. This process is important for understanding how cells repair damage and reshuffle genetic information. During recombination, gene conversion can copy information from one DNA molecule to another. The result is not a simple swap of DNA segments. Instead, one sequence is corrected or replaced while the related partner sequence remains...

restarting stalled replication forks

Restarting DNA Replication After Stalls

DNA replication can stop before it is finished when the fork stalls. A stalled replication fork is a point where the DNA-copying machinery slows down or pauses during chromosome duplication. Cells can restart these forks to keep copying DNA. This process helps the replication machinery move forward again after the pause. It is an important part of maintaining normal DNA replication when problems arise during copying. The term stalled replication fork describes the stopped or delayed state of...

key terms and concepts

Learn how cells repair DNA double-strand breaks using direct joining or a template-based pathway. Clear, high school-friendly biology content from JoVE K12.

Repairing DNA Double-Strand Breaks | JoVE K12

cell biology

JoVE Core Cell Biology illustrates complex cellular processes in easy-to-understand yet comprehensive animated video lessons. This collection focuses on the essential components and functions of cells, including organelles and membranes, cellular transport, and cell energetics. It is complemented by scientist-in-action videos that demonstrate the application of cell biology in real-world research experiments.

Louisiana Student Standards for Science

High School

Life Sciences

Heredity: Inheritance And Variation Of Traits

HS-LS3-3

Texas Essential Knowledge and Skills

BIO.7

BIO.7.C

What is Meiosis?

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Meiosis I

Meiosis II

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Crossing Over

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Nondisjunction

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Spermatogenesis

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Oogenesis

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Fertilization

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Cleavage and Blastulation

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Gastrulation

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Neurulation

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Cell Migration

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Determination

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Use models to explain how DNA is passed from parents to offspring through the processes of meiosis and fertilization in sexual reproduction.

LS.Bio.6.1

Mutations

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genome-size-and-the-evolution-of-new-genes

Genome Size and the Evolution of New Genes

types-of-genetic-transfer-between-organisms

Types of Genetic Transfer Between Organisms

Base-pairing and DNA Repair

The DNA Replication Fork

Lagging Strand Synthesis

The Replisome

Proofreading

Synthesis of new DNA molecules is carried out by the enzyme DNA polymerase, which adds nucleotides on the daughter strand complementary to the template ...

Replication in Prokaryotes

DNA replication has three main steps: initiation, elongation, and termination. Replication in prokaryotes begins when initiator proteins bind to the ...

Replication in Eukaryotes

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Overview of DNA Repair

Base Excision Repair

Nucleotide Excision Repair

DNA Distortion and Damage
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Mismatch Repair

Organisms are capable of detecting and fixing nucleotide mismatches that occur during DNA replication. This sophisticated process requires identifying the ...

Homologous Recombination

Gene Conversion

Restarting Stalled Replication Forks

Construct an explanation to summarize how inheritable genetic variations may result from: new genetic combinations in meiosis, mutations during replication, or mutations caused by environmental factors.

An Overview of Genetic Analysis

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Genetic Crosses

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Punnett Squares

Monohybrid Crosses

Dihybrid Crosses

Trihybrid Crosses

Trihybrid Crosses
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Law of Independent Assortment

Chi-square Analysis

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Pedigree Analysis

Multiple Allele Traits

Incomplete Dominance

Gregor Mendel&#39;s work (1822 - 1884) was primarily focused on pea plants. Through his initial experiments, he determined that every gene in a diploid ...

Lethal Alleles

Agouti: A Lethal Allele
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Polygenic Traits

background-and-environment-affect-phenotype

Background and Environment Affect Phenotype

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Use mathematics and computational thinking to predict the variation and distribution of expressed traits based on: Mendelian inheritance, co-dominance, incomplete dominance, multiple alleles, and sex-linked inheritance.

LS.Bio.7.1

Overview

A pedigree is a diagram displaying a family&#x2019;s history of a trait. Analyzing pedigrees can reveal (1) whether a trait is dominant or ...

Probability Laws

Overview

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The Concept of Multiple Allelism

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Analyze and interpret data to explain how polygenic traits result in a wide range of phenotypes.

LS.Bio.7.2

Pleiotropy

Pleiotropy is the phenomenon in which a single gene impacts multiple, seemingly unrelated phenotypic traits. For example, defects in the SOX10 gene cause ...

Nature and Nurture

Many human characteristics, like height, are shaped by both nature&#x2014;in other words, by our genes&#x2014;and by nurture, or our environment. For ...

Construct an explanation to summarize how traits result from interactions of genetic factors (multiple genes and/or alleles) and environmental factors.

LS.Bio.7.3

PCR: The Polymerase Chain Reaction

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DNA Gel Electrophoresis

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DNA Isolation

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Restriction Enzymes

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DNA Agarose Gel Electrophoresis

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dna-isolation-and-restriction-enzyme-analysis

DNA Isolation and Restriction Enzyme Analysis

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Preparation of Solutions and Materials
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First ...

DNA Isolation
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Analyze and interpret data to compare DNA samples.

LS.Bio.8.1

Transgenic Organisms

Reproductive Cloning

CRISPR

Synthetic Biology

Synthetic biology is an interdisciplinary science that involves using principles from disciplines such as engineering, molecular biology, cell biology, ...

Ribosome Profiling

Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences ...

Transgenic Plants

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Gene Therapy

biofuels-producing-ethanol-from-cellulosic-material

Biofuels: Producing Ethanol from Cellulosic Material

Source: Laboratories of Margaret Workman and Kimberly Frye - Depaul University
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Obtain and communicate information that summarizes the impact of biotechnology applications on the individual, society, and the environment, including agriculture and medicine.

LS.Bio.8.2

Understand genetic mechanisms for variation.

Repairing DNA Double-Strand Breaks

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