Gene Mutation, Repair And Transposition Flashcards
1
Q
What is the function of the DNA?
A
Stores information for cellular function
2
Q
How can variations/changes in DNA affect someone or populations?
A
- Phenotypic variability
- Adaptation to environmental changes
- Evolution
- Genetic disease, cancer, cell death
3
Q
How has phenotypic variation from mutation helped scientists?
A
Allows geneticists to identify and study the involved gene
-sometimes we only know the normal activity of a gene when we see what changes in a cell or organism after the mutation has occurred
Mutations can also act as markers for genes
4
Q
What is DNA repair?
A
A cell has various mechanisms to correct base pair changes: DNA repair
5
Q
What are the types of mutations?
A
- Point or base substitutions
- Missense mutation
- Nonsense mutation
- Silent mutation
6
Q
What is a point/base substitution mutation?
A
A change from one base pair to another
7
Q
What is a missense mutation?
A
A change in one nucleotide of a triplet within the protein coding portion of a gene resulting in a new triplet that codes for a different amino acid
8
Q
What is a nonsense mutation?
A
A triplet is changed to a stop codon, resulting in the termination of translation of the protein (premature stop codon)
-defective protein product: extent of deficit depends on of the stop codon is at the beginning or later on in the sequence
9
Q
What is a silent mutation?
A
There is an alteration of a codon but it doesn’t change the amino acid at that position
-if the change affects a splice site, it could affect the protein product
10
Q
Objective: in what scenario can a mutation be found?
A
-An alteration in DNA(not major alteration in chromosome)
-Can be found in coding or noncoding
regions of a gene
- Mutations can occur in somatic cells during cell duplication
- Mutations can occur with germ cells which are heritable
11
Q
What is the effect of mutations in germ cells
A
These are heritable and increases genetic diversity and evolution or disease
12
Q
What are the effects of mutations in somatic cells during cell duplication?
A
Not heritable but may lead to altered cellular function or tumors
13
Q
What are coding regions of a gene?
A
Coding region of a gene- the regions which contain the information to be translated into protein
14
Q
What are the noncoding regions of a gene?
A
Noncoding regions may include a gene regulatory region like the promoter or introns or splice signals
15
Q
What are transitions?
A
Pyrimidine replaces pyrimidine or a purine replaces purine
16
Q
What are transversions ?
A
Purine and pyrimidine are interchanged
17
Q
What are frameshift mutations?
A
- Results from the insertion of deletion of a base pair
- This causes a shift in the reading frame
- The codons for the amino acids are all changed and no longer make a proper protein
- Almost always causes a premature stop codon soon after the frameshift
18
Q
What do frameshift mutations usually lead to?
A
- usually lead to a premature stop codon, resulting in shorter than normal protein
- Worse if this change happens neat the start of the sequence
- The RNA with a premature stop codon is often degraded by a special pathway called Nonsense Medicated Decay where the affected RNA is degraded
19
Q
What are the types of loss of function mutations?
A
- Null mutation
- Recessive mutation
- Dominant mutation
- Dominant negative mutation
20
Q
What is a loss of function mutation?
A
Reduces or eliminates the function of the gene product
21
Q
What is a null mutation?
A
Complete loss of function
22
Q
What is a recessive mutation?
A
Wild type phenotype is the other allele is wild type, 50% gene product (from the good allele) is enough to bring the wild type phenotype
23
Q
What is a dominant mutation?
A
Mutant phenotype even when the other allele is wild type, 50% of gene product (from the good allele) is NOT ENOUGH and the defective phenotype is shown
-Also known as haploinsufficiency
24
Q
What is dominant negative mutation?
A
The one mutant allele is inactive and directly interferes with the function of the product from the good allele
-Example of dominant negative is when the nonfunctional gene product binds to the wild type product in a homodimer, inactivating or reducing the activity of the homodimer
25
What are gain of function mutations?
Mutation codes for an altered gene product with enhanced, negative or new functions
-amino acid change leads to new activity
26
What is a suppressor mutation?
A second mutation in the same gene that reverts or relieves the effects of the first mutation (first mutation is gain of function)
Example: one deletion soon followed by insertion which restores the reading frame
27
If a gain of function mutation is in a regulatory region of a gene, what may be the result?
- higher expression of the gene
- The gene turned on at the wrong time
- The gene turned on in the wrong tissue or place
28
What are the types of lethal mutations?
Lethal mutations
Lethal condition mutations
29
What are lethal mutations?
-Interrupt an essential process and result in an early death
- Typical of the mutations which lead to biochemical disorders
- e.g., tay Sachs results in death in early childhood
30
What are lethal condition mutations?
- Usually where the viability of the organism depends on the environment
- E.g., the organism grows normally in total medium (with all nutrients) and not on minimal medium (with restricted nutrients)
31
What are neutral mutations?
- Majority of these types occur in noncoding regions
| - Neither beneficial or detrimental
32
What are somatic mutations?
- Mutations restricted to the somatic cells and not transmitted to future generations
- Mutation occurs in any cell of the bidy except germ cells
- a Can occur on any chromosome
33
What are inherited mutations?
- Autosomal mutations occur in the germ cells and are found on the autosomal chromosomes (1-22 in humans)
- Sex-linked mutation can occur in the germ cells and are found on the X or Y chromosomes
34
What are inherited dominant mutations?
Phenotype seen in the first generation to inherit the mutation
35
What are inherited dominant mutations?
Phenotype not seen till the chance mating of two similarly affected persons brings two copies of the mutation into a single zygote which will now be homozygous for the mutation
36
Mutations occur...
Spontaneously and randomly
37
What are spontaneous mutations?
These are changes in nucleotide sequence that occurs naturally
Most arise from normal biological or chemical process in the organism that changes the structure of the nitrogenous bases
38
At what cellular function/genetic process dies spontaneous mutation occur?
Most often this occurs during DNA replication
39
What is the frequency of spontaneous mutations?
Rates vary among organisms, but in all extremely low
Some regions of DNA appear to be more prone to mutation that others FYI, nothing here to memorize
40
What are induced mutations?
Result from the influence exogenous factors, whether natural, or artificial
Radiation
UV light
Natural or synthetic chemicals(intrudes in processed foods)
41
What was the methodology of the Iceland Germ-line mutation rate study? What did they find out about mother’s?
In 2012, DNA sequencing of 78 parent/offspring sets -DNA sequences compared
- Looking for single nucleotide changes called Single Nucleotide Polymorphisms(SNPs)
- Newborn baby genome contains the average of 60 nucleotides changed (new mutations) compared to parents
- Mothers contribute about 15 new mutations to baby at any age
42
What did the Iceland Germ-line mutation rate study find out about fathers?
The number of new mutations from fathers depends significantly on the age of the father at the time of conception
Father 20 years old= about 25 new mutations
Father 40 years old about 65 new mutations
Both of these indicate about 2 mutations per year of life
May occur because make germ cells go through more cell divisions during a life time compared to females
43
How do spontaneous changes occur?
DNA Replication errors
Tauromeric shifts
44
Obj: What are the mechanisms of DNA replication errors?
Slippage; tautomeric shifts; depurination and deamination; oxidative damage; other mutagenic agents
45
How are nucleotides miscorporated into new strand?
A form of DNA replication errors
-occasionally DNA polymerases insert incorrect nucleotides even after proof reading (3’ to 5’ exonuclease proof reading activity)
46
What is replication slippage?
Small inserts or deletions can occur when DNA polymerase slips or stutters during replication
- DNA template makes a loop during replication, a small deletion in the new strand occurs
- DNA polymerase introduces nucleotides that are not present in the template strand and a small insertion in the new strand occurs
- Replication slippage can occur anywhere but seen more commonly in regions containing tandemly repeated sequences
47
Can purines and pyrimidines exist in tautomeric forms?explain this
Purines and pyrimidines can exist in tautomeric forms
- An alternate chemical form that differs by a proton
- Keto-enol forms of thymine and guanine
- Amino-imino forms of cytosine and adenine
48
What are the results and effects of tautomeric shifts?
- Tautomers increase the chance of mispairing during DNA replication
- Can change the bonding structure allowing noncomplementary base Pairing
- May lead to permanent base-pair changes
Results: A transient tautomeric shift in adenine during round 1 of replication
-Results in the formation of an A-T to C-G transition mutation in round 2 of replication
Daughter strand showing heritable mutation
49
What is depurination?
The loss of one of the nitrogenous basis in intact DNA
-usually a purine (A or G)
- The glycosidic linkage from the C1’ of the sugar to atom 9 of the purine ring is broken
- Result: DNA polymerase May add a random nucleotide
50
What is deamination ?
The amino group on cytosine or adenine is converted to a keto group
Cytosine —> uracil
Adenine —> hypoxanthine
51
What causes oxidative damage to DNA?
Due to by-products of normal cellular processes: aerobic respiration
Also generated by exposure to high-energy radiation
- Super oxide (O2- radical)
- hydroxyl radicals (OH- radical)
- Hydrogen peroxide
52
What are the common types of modifications ?
- Modification of bases
- Loss of bases
- Single stranded breaks
- Many others less common
53
What mutations are caused by UV radiation?
Mainly causes thymine dimers
| -covalent crosslinks Shawn in red
54
What are intercalating agents ?
These Chemicals have the tight shape to allow them to wedge between the base pairs
-Causes DNA to distort and unwind, leading to frame shift mutations
55
What are base analogues?
5-bromouracil behaves like a thymine analogue but nature favors the tautomeric structure so causes A-T to C-G transition after one round of replication
56
What do single gene mutations cause?
Mutations in one gene cause disease
Disease is monogenic disease
57
Obj: What is allergic heterogeneity ?
Mutations in the same gene all lead to the same disease
Example here is the HBB gene that cause b-thalassemia - one disease, many mutation sites in the one gene
58
Describe the transposable elements “jumping genes”
Barbara McClintock studded the genetics of maze, first to describe recombination and in 1950, developed the theory that mobile elements can move within and between chromosomes and regulate genes
- Research was conceptually difficult, not immediately understood
- in 1950’s Jacob and Monad discovered gene regulation of E. Coli and scientific community was ready to understand gene regulation
59
In what organisms have transposable elements been found?
In all organisms found
60
What are transposable elements?
DNA elements that move within or between genome
61
Transposable elements can be from .......
| Bp and can include.....
50-10,000 base pairs
Includes ORFs(open reading frames)
62
How much of the human genome is derived from the transposable elements?
Almost 50% of human genome is derived from Transposabke elements
63
What can be caused from transposable elements (negatively)?
Inversions, translocations, double stranded breaks
64
What mechanism controls the functioning of transposable elements?
Mechanism thought to act as naturally occurring mutagens
65
What can result by transposable elements insertions?
They may insert into a new location of the genome:
- If they insert into the coding region of a gene, they can alter the reading frame
- If they insert into the regulatory region of a gene, they can disrupt proper expression of the gene
66
What are the steps involved in DNA transposon transposition : cut and paste?
Step 1: transposase cleaves DNA at ITRs
Step 2: Transposase makes staggered cuts at target site in chromosomal DNA
Step 3: transposase inserts transposing at target site
Step 4: gaps in target site are filled in by DNA polymerase and DNA ligase
Donor DNA is repaired with no trace of TE
67
What are inverted terminal repeats (ITR)?
These are located on each end if the transposable elements (TE)and an open reading from codes for enzyme Transposase
-ITRs are between 9-40 bp long and are ifentical but inverted relative to each other
68
Explain the functioning of the transposase enzyme
- ITR is recognized, bound and cleaved by the transposase enzyme between ITR and DR, brings the two together
- The transposing loop will be inserted into new target DNA which has a target site DNA site
- Transposase will cut the target DNA at the target site leaving 5’ single stranded overhangs
- Gaps are filled in by DNA pol and DNA ligase
69
What are the types of transposable elements ?
Autonomous transposons
Nonautonomous transposons
70
What are autonomous transposons?
- TE contains transposase ORF (encoding for transposase enzyme) plus/minus another ORF that encodes for a gene
- Intact ITR to the left and right of the ORFs (flanking the ORF)
71
What are nonautonomous transposons?
- contains DNA but NOT transposase gene
| - To function there has to be transposase enzyme available
72
What are the types of retrotransposons?
- Long terminal repeat (LTR) retrotransposons
| - Non-LTR Retrotransposons
73
What are long-terminal repeat(LRT) transposons?
- Long term repeats flank the retrotransposon
- Contains transcription promoters in the 5’ untranslated region and polyadenylation sites in the 3’ untranslated region
- Can be autonomous or non-autonomous (+/- integrate and reverse transcriptase)
74
What are Non-LTR Retrotransposons?
- Lacks the LTR region
- Contains transcription promoters in the 5’ untranslated region and polyadenylation sites in the 3’ untranslaated region
- Can be autonomous or nom-autonomous (+/- integrate and reverse transcriptase
75
What are the functions of Retrotransposons?
- RNA is made
- integrate and reverse transcriptase enzymes produced
- The RNA is converted back to dsDNA by reverse transcriptase
- Integrase inserts dsDNA into chromosomal DNA
76
What are the steps of Retrotransposon functioning?
Step 1 and 2: Retrotransposon is transcribed and translated by cellular enzymes
Step 3: reverse transcriptase creates double-stranded DNA copies of each Retrotransposon RNA
Step 4: integrase inserts double-strandedDNA-copies into chromosomal DNA
77
What are the functions of DNA Repair?
- counteract spontaneous and induced DNA damage
- Maintain the integrity of genetic material
- Counteract genetic damage that would result in genetic diseases and cancer
78
How often dies DNA pol. 3 male a mistake in bacteria?
Once every 100,000 base pairs (10^-5)
79
How does DNA pol. 3 prevent mistakes? How useful is this?
1. DNA pol. 3 proofreads each step and can “reverse its direction, cut out the bad nucleotide (3’-5’ exonuclease activity) and replace with the proper nuccleotide
2. This improved the efficiency of replication 100 fold, to a mistake every 10,000,000 (10^-7)
80
What polymerase proofreads DNA in eukaryotes?
DNA polymerase delta and epsilon deal with elongation have proofreading ability (3’-5’ exonuclease)
81
What is mismatch repair?
After proofreading, there may be base-base mismatching, small insertions or deletions
-repair based on DNA methylation
82
What enzyme in bacteria can be used for mismatch repair?
In bacteria, the enzyme DNA adenine methylase will recognize the DNA sequence and methylated the adenine nuccleotide
83
How is DNA mismatch fixed?
- Endonuclease will nick the phosphate backbone of the new un-methylated strand of DNA
- Exonuclease will unwind and degrade the nicked DNA till the mismatch is removed
- DNA polymerase will fill in the gap using the correct DNA strand as a template
- DNA ligase will seal the nicked backbone
84
Contrast endonuclease and exponuckease
Endonuclease -internal cuts in DNA
Exonuclease-Nucleotides moved from the ends
85
What bacterial proteins are involved in the DNA repair?
- MutH recognizes the mismatch and has endonuclease activity
- MutL helps form a stable complex of DNA binding proteins
- MutS assists in the recognition of the mutation. Mutation in these proteins and the bacteria is deficient in MMR
86
What proteins in humans are involved in DNA repair?
- hMSH2 recognizes the mismatch and recruits other repair proteins
- hMLH1 helps form a stable complex of DNA binding proteins
Mutations in these genes are associated with hereditary nonpolyposis colorectal cancer
87
Mutations in bacterial and human proteins involved in DNA repair ....
Mutations in any of these result in any of the genes that code for MMR can be associated with many other cancers as well
88
Explain post-replication repair
If DNA repair has skipped over a lesion (damage or abnormal change) such as a thymine dimer
- Replication skips over the lesion and continues
- The correct complimentary sequence is taken from the other parental strand
- New gap is filled in by DNA polymerase and backbone is sealed by DNA linkage
89
Explain the SOS Repair system in E. Coli
- Last resort to minimize DNA damage
- DNA synthesis becomes error-prone; inserts random/incorrect nucleotides in places that normally would stall replication
- Bacteria induce expression of 20 genes; their products allow replication to move forth
- SOS repair cam itself become mutagenic; allows cells to survive with DNA damage (cell would otherwise kill itself)
90
What organisms use photoreactivation DNA repair?
- Bacteria, fungi and plants
| - Not in humans
91
Explain how photoreactivation repair functions?
- UV light induces the formation of pyrimidine dimers
- Following this radiation, if the DNA is exposed to visible light, particularly in the blue range, the pyrimidine dimers can be reversed
- The blue light activates the activity of the Photoreactivation Enzyme also called PRE or photo lyase
92
What organisms use Base excision repair(BRE)?
Repair mechanism found in prokaryotes and eukaryotes
93
Give an example of Base excision Repair (BER) repairing U-G mismatch
- Repairs the most common mutation is the deamination of cytosine to uracil
- Here, uracil DNA glycolase recognizes and excises the incorrect base and created an apurinic/apyrimidinic.
- The sugar with the missing base is recognized by an enzyme called AP endonuclease makes a cut in the backbone
- DNA polymerase fills and DNA ligase repairs the gap
94
What is the function Nucleotide Excision Repair(NER)?
Repairs the bulky lesions in DNA that alter or distort the double helix such as UV induced pyrimidine dimers
95
Give the history of nucleotide excision repair(NER)
UV sensitive E. Coli mutants
- A group of genes were identified, ultraviolet repair (uvr)
- uvrA, uvrB and uvrC were found mutated in these UV sensitive E. Coli
- the activity of the uvr genes have been worked out and the pathway shown to the right
- uvrA, uvrB recognize and bind to the lesion and the recruit uvrC and then uvrA dissociates
- uvrB cleaves the phosphodiester bond 4 nucleotides downstream and uvrC cleaves the phosphodiester bond 8 nucleotides upstream of the DNA damage
The gap is filled by DNA polymerase and backbone lighted by DNA ligase
96
What organisms use a Nucleotide Excision Repair (NER)?
Prokaryotes but similar system is found in eukaryotes
97
What causes Xeroderma pigmentosa in humans ?
- NER in eukaryotes involves about 30 genes
| - If any of these genes are mutated, it can lead to xeroderma pigmentisa (XP)
98
What is Xeroderma Pigmentosa?
- XP is an autosomal recessive disorder that predisposes individuals to severe skin abnormalities and skin cancers as well as developmental and neurological defects
- by age 2, freaking of the skin in sun-exposed regions
- Skin coloring changes
- 2000-fold higher rate of cancer
- Persons with XP are very sensitive to UV light, can get a sunburn after just a few minutes in the sun
99
How can Xeroderma Pigmentosa be arrested?
Early detection and protection from sunlight can arrest it
100
What 3 diseases are caused by defects in NER?
- Cockayne syndrome (CS)
- Xeroderma Pigmentosa (XP)
- Trichothiodystrophy (TTD)
101
What causes Cockayne syndrome (CS)?
- Autosomal recessive inheritance
- Developmental and neurological defects
- Sunlight sensitivity but no increase in cancers
- Premature aging, with death by age 20
102
What causes Trichothiodystrophy ? What are the effects?
- Mainly autosomal recessive inheritance
- rare X linked case found
- Sulfur-deficient brittle hair, brittle skin and nails
- Hair has tiger banding under polarized microscopy
- Dwarfism, retardation
- Facial deformities and sensitivity to sunlight
- No increase in cancers- six year life span
