Molecular Biology - 04 Gene Mutation

MOLECULAR BIOLOGY AND DIAGNOSTICS GENE MUTATION HEREDITARY MUTATIONS • OUTLINE • Molecular Definition of Gene • Causes of Gene Mutation Mutation o What are the Causes of Gene o What is Gene Mutation? Mutation? o Mutations & the Environment • Gene Mutation Occurs in 4 o Mutations caused by Ways Chemicals o What is a Gene Mutation and o Spontaneous Mutations How do Mutations Occur? o Errors during DNA Replication o Hereditary Mutations o Acquired/Somatic Mutations • Types of Gene Mutation o De Novo/New Mutations o What are the Types of Gene o Mosaicism Mutations? o Types of Mutation • • ACQUIRED (OR SOMATIC) MUTATIONS • • L1: MOLECULAR DEFINITION OF GENE MUTATION WHAT IS GENE MUTATION? • It is the changing of the structure of a gene, resulting in a variant form that may be transmitted to subsequent generations, caused by the alteration of single base units in DNA, or the deletion, insertion, or rearrangement of larger sections of genes or chromosomes. • Occur at some time during a person’s life and are present only in certain cells, not in every cell in the body. These changes can be caused by environmental factors such as ultraviolet radiation from the sun or can occur if an error is made as DNA copies itself during cell division. Acquired mutations in somatic cells (cells other than sperm and egg cells) cannot be passed to next generation. DE NOVO (NEW) MUTATIONS • • • • ` Inherited from a parent and are present throughout a person’s life in virtually every cell in the body. These mutations are also called germline mutations because they are present in the parent’s egg or sperm cells, which are also called germ cells. When an egg and a sperm cell unite, the resulting fertilized egg cell receives DNA from both parents. If this DNA has a mutation, the child that grows from the fertilized egg will have a mutation in each of his or her cells. Genetic changes that are described as can be either hereditary or somatic. In some cases, the mutation occurs in a person’s egg or sperm cell but is not present in any of the person’s other cells. In other cases, the mutation occurs in the fertilized egg shortly after the egg and sperm cells unite. (It is often impossible to tell exactly when a de novo mutation happened.) As the fertilized egg divides, each resulting cell in the growing embryo will have the mutation. De novo mutations may explain genetic disorders in which an affected child has a mutation in every cell in the body but the parents do not, and there is no family history of the disorder. MOSAICISM • • • L2: GENE MUTATION OCCURS IN 4 WAYS WHAT IS A GENE MUTATION AND HOW DO MUTATIONS OCCUR? • • • A gene mutation is a permanent alteration in the DNA sequence that makes up a gene, such that the sequence differs from what is found in most people. Mutations range in size; they can affect anywhere from a single DNA building block (base pair) to a large segment of a chromosome that includes multiple genes. Gene mutations can be classified in four ways: o Hereditary o Acquired o De novo o Mosaicism These are Somatic mutations that happen in a single cell early in embryonic development. T hese genetic changes are not present in a parent’s egg or sperm cells, or in the fertilized egg, but happen a bit later when the embryo includes several cells. As all the cells divide during growth and development, cells that arise from the cell with the altered gene will have the mutation, while other cells will not. Depending on the mutation and how many cells are affected, mosaicism may or may not cause health problems. • L3: CAUSES OF GENE MUTATION • WHAT ARE THE CAUSES OF GENE MUTATION? • DNA in any cell can be altered by way of a number of factors, including environmental influences, certain chemicals, spontaneous mutations, and errors that occur during the process of replication. MUTATIONS CAUSED BY CHEMICALS ERRORS IN DNA REPLICATION • On very, very rare occasions DNA polymerase will incorporate a non-complementary base into the daughter strand. During the next round of replication, the misincorporated base would lead to a mutation. This, however, is very rare as the exonuclease functions as a proofreading mechanism recognizing mismatched base pairs and excising them. • ERRORS IN DNA RECOMBINATION • DNA often rearranges itself by a process called recombination which proceeds via a variety of mechanisms. Occasionally DNA is lost during replication leading to a mutation. • CHEMICAL DAMAGE TO DNA • Many chemical mutagens, some exogenous, some manmade, some environmental, are capable of damaging DNA. Many chemotherapeutic drugs and intercalating agent drugs function by damaging DNA. RADIATION • Gamma rays, X-rays, even UV light can interact with compounds in the cell generating free radicals that cause chemical damage to DNA. MUTATIONS AND THE ENVIRONMENT • • • DNA interacts with the environment, and sometimes that interaction can be detrimental to genetic information. In fact, every time you go outside, you put your DNA in danger, because ultraviolet (UV) light from the Sun can induce mutations in your skin cells. One type of UV-generated mutation involves hydrolysis of a cytosine base to a hydrate form, causing the base to mispair with adenine during the next round of replication and ultimately be replaced by thymine. Researchers have found an extremely high rate of occurrence of this UVinduced C-to-T fingerprint-type mutation in genes associated with basal cell carcinoma, a form of skin cancer UV light can also cause covalent bonds to form between adjacent pyrimidine bases on a DNA strand, which results in the formation of pyrimidine dimers. Repair machinery exists to cope with these mutations, but it is somewhat prone to error, which means some dimers go unrepaired. Furthermore, some people have an inherited genetic disorder called xeroderma pigmentosum (XP), which involves mutations in the genes that code for the proteins involved in repairing UV-light damage. In people with XP, exposure to UV light triggers a high frequency of mutations in skin cells, which in turn results in a high occurrence of skin cancer. As a result, such individuals are unable to go outdoors during daylight hours. In addition to ultraviolet light, organisms are exposed to more energetic ionizing radiation in the form of cosmic rays, gamma rays, and X-rays. Ionizing radiation induces double-stranded breaks in DNA, and the resulting repair can likewise introduce mutations if carried out imperfectly. Unlike UV light, however, these forms of radiation penetrate tissue well, so they can cause mutations anywhere in the body. Oxidizing agents, commonly known as free radicals, are substances that can chemically modify nucleotides in ways that alter their base-pairing capacities. For instance, dioxin intercalates between base pairs, disrupting the integrity of the DNA helix and predisposing that site to insertions or deletions. Similarly, benzo[a]pyrene, a known carcinogen and a component of cigarette smoke, has been demonstrated to induce lesions at guanine bases in the tumor suppressor gene P53 at codons 157, 248, and 273. These codons are the major mutational hot spots seen in clinical studies of human lung cancers Mutations such as these that are fairly specific to particular mutagens are called signature mutations. A variety of chemicals beyond those mentioned here are known to induce such mutations. SPONTANEOUS MUTATIONS • Mutations can also occur spontaneously. For instance, depurination (Figure 1), in which a purine base is lost from a nucleotide through hydrolysis even though the sugarphosphate backbone is unaltered, can occur without an explicit insult from the environment. If uncorrected by DNA repair enzymes, depurination may result in incorporation of an incorrect base during the next round of replication. o • Figure 1: Depurination is a spontaneous mutation that occurs when a nucleotide loses a purine base. During replication, two strands of DNA separate. If a nucleotide on one strand has lost a purine base, the apurinic site on this strand cannot provide a template for a complementary base on the newly-synthesized strand. An incorrect nucleotide (most often adenine) is inserted into the newly-synthesized strand, across from the empty apurinic site on the template strand. The result is a normal double-stranded DNA molecule that does not contain a mutation, and a mutant double-stranded DNA molecule. When the mutant DNA undergoes a second round of replication, the incorrectly incorporated base (adenine) acquired during the previous replication round is used as a template for synthesizing a new DNA strand. The two resulting double-stranded DNA molecules each contain a permanent mutation in both of their strands. Deamination, or the removal of an amine group from a base, may also occur. Deamination of cytosine converts it to uracil, which will pair with adenine instead of guanine at the next replication, resulting in a base substitution. Repair enzymes can recognize uracil as not belonging in DNA, and they will normally repair such a lesion. However, if the cytosine residue in question is methylated (a common modification involved in gene regulation), deamination will instead result in conversion to thymine. Because thymine is a normal component of DNA, this change will go unrecognized by repair enzymes (Figure 2). • • o Figure 2: Deamination is a spontaneous mutation that occurs when an amine group is removed from a nitrogenous base. The nitrogenous base cytosine is converted to uracil after the loss of an amine group. Because uracil forms base-pairs with adenine, while cytosine forms base-pairs with guanine, the conversion of cytosine to uracil causes base substitutions in DNA. As previously mentioned, repeats also occur in nonmitotic tissue, and CAG repeats have further been shown to accumulate in mice defective for individual DNA repair pathways, suggesting that multiple repair mechanisms must be operative in repeat expansion in nonproliferating cells (Pearson et al., 2005). In agreement with this hypothesis, studies have revealed increased repeat instability following induction of double-stranded breaks and UV-induced lesions, which are corrected by nucleotide excision repair. To date, all diseases associated with TNRs involve repeat instability upon transmission from parent to offspring, often in a sex-specific manner. For example, the CAG repeats that characterize Huntington's disease typically exhibit greater expansion when inherited paternally. This expansion has been shown to occur prior to meiosis, when germ cells are proliferating. Contraction of other TNRs has been linked to sex-specific differences in germ-line DNA methylation patterns (Pearson et al., 2005). L4: TYPES OF GENE MUTATION WHAT ARE THE TYPES OF GENE MUTATIONS? ERRORS DURING DNA REPLICATION • • Errors that occur during DNA replication play an important role in some mutations, especially trinucleotide repeat (TNR) expansions. It is thought that the ability of repeat sequences to form secondary structures, such as intrastrand hairpins, during replication might contribute to slippage of DNA polymerase, causing this enzyme to slide back and repeat replication of the previous segment (Figure 7). Supporting this hypothesis, lagging-strand synthesis has been shown to be particularly sensitive to repeat expansion. For instance, the secondary structure of some TNR DNA has been shown to inhibit an enzyme (FEN1) necessary for proper resolution of the Okazaki fragments generated during lagging-strand replication; as a result, FEN1 mutant yeast cells demonstrate increased expansion of CAG repeats. • A mutation, which may arise during replication and/or recombination, is a permanent change in the nucleotide sequence of DNA. Damaged DNA can be mutated either by substitution, deletion, or insertion of base pairs. Mutations, for the most part, are harmless except when they lead to cell death or tumor formation. Because of the lethal potential of DNA mutations cells have evolved mechanisms for repairing damaged DNA. POINT MUTATION • • • • A point mutation or a single base modification is a type of mutation that causes a single nucleotide base substitution, insertion, or deletion of the genetic material, DNA or RNA. The term frameshift mutation indicates the addition or deletion of a base pair. Point mutant - individual that is affected by a point mutation. Repeat induced point mutations are recurring point mutations. CHROMOSOMAL MUTATION NONSENSE MUTATION • • A mutation involving a long segment of DNA. These mutations can involve deletions, insertions, or inversions of sections of DNA. In some cases, deleted sections may attach to other chromosomes, disrupting both the chromosomes that loses the DNA and the one that gains it. Also refereed to as a chromosomal arrangement. A nonsense mutation is also a change in one DNA base pair. Instead of substituting one amino acid for another, however, the altered DNA sequence prematurely signals the cell to stop building a protein. This type of mutation results in a shortened protein that may function improperly or not at all. COPY NUMBER VARIATION • CNVs is a relatively new field in genomics and it is defined as a phenomenon in which sections of the genome are repeated and the number of repeats in the genome varies between individuals in the human population. SOMATIC MUTATION • Genetic alteration acquired by a cell that can be passed to the progeny of the mutated cell in the course of cell division. Somatic mutations differ from germ line mutations, which are inherited genetic alterations that occur in the germ cells. TYPES OF MUTATION INSERTION • • • What kinds of gene mutations are possible? o The DNA sequence of a gene can be altered in a number of ways. Gene mutations have varying effects on health, depending on where they occur and whether they alter the function of essential proteins. The types of mutations include: MISSENSE MUTATION • An insertion changes the number of DNA bases in a gene by adding a piece of DNA. As a result, the protein made by the gene may not function properly. This type of mutation is a change in one DNA base pair that results in the substitution of one amino acid for another in the protein made by a gene. DELETION • A deletion changes the number of DNA bases by removing a piece of DNA. Small deletions may remove one or a few base pairs within a gene, while larger deletions can remove an entire gene or several neighboring genes. The deleted DNA may alter the function of the resulting protein(s). DUPLICATION • A duplication consists of a piece of DNA that is abnormally copied one or more times. This type of mutation may alter the function of the resulting protein. FRAMESHIFT MUTATION • This type of mutation occurs when the addition or loss of DNA bases changes a gene's reading frame. A reading frame consists of groups of 3 bases that each code for one amino acid. A frameshift mutation shifts the grouping of these bases and changes the code for amino acids. The resulting protein is usually nonfunctional. Insertions, deletions, and duplications can all be frameshift mutations. REPEAT EXPANSION • Nucleotide repeats are short DNA sequences that are repeated a number of times in a row. For example, a trinucleotide repeat is made up of 3-base-pair sequences, and a tetranucleotide repeat is made up of 4-base-pair sequences. A repeat expansion is a mutation that increases the number of times that the short DNA sequence is repeated. This type of mutation can cause the resulting protein to function improperly. TYPES OF DNA MUTATIONS AND THEIR IMPACT Class of Mutation Type of Mutation Description One base is incorrectly added during replication and Substitution replaces the pair in the corresponding position on the complementary strand One or more extra nucleotides are Point inserted into Insertion mutation replicating DNA, often resulting in a frameshift One or more nucleotides is "skipped" during Deletion replication or otherwise excised, often resulting in a frameshift One region of a Inversion chromosome is flipped and reinserted A region of a chromosome is lost, Deletion resulting in the absence of all the genes in that area Chromosomal A region of a mutation chromosome is repeated, resulting in Duplication an increase in dosage from the genes in that region A region from one chromosome is Translocation aberrantly attached to another chromosome The number of tandem Gene copies of a locus is amplification increased Copy number The normal number of variation Expanding repeated trinucleotide trinucleotide sequences is repeat expanded • Human Disease(s) Linked to This Mutation Sickle-cell anemia One form of betathalassemia Cystic fibrosis Opitz-Kaveggia syndrome Cri du chat syndrome Some cancers One form of leukemia Some breast cancers Fragile X syndrome, Huntington's disease