Topic 1 - Nucleic Acids Flashcards
Lectures 1-5
1
Q
What are Megakaryocytes the parent cells of?
A
Blood Platelets
2
Q
How many nucleaotides does the human genome consist of?
A
About 3x10^9
3
Q
How many chromosomes in (most) human cells?
A
46
4
Q
Which cells can copy their DNA without dividing?
A
Megakaryocytes, Hepatocytes etc.
5
Q
What is the Barr body?
A
Women inactivate one of their XX chromosomes & push it to the edge of the nucleus
6
Q
Why do Chimps have a greater chromosome number?
A
Human chromosome 2 is a fusion of chimp chromosomes 12 & 13
7
Q
What is Synteny?
A
Where long DNA sequences are present in the same order across species
8
Q
What is translocation?
A
Chromosomes breaking and reforming
9
Q
What causes Chronic Myelogenous Leukemia?
A
Faulty translocation in the Philadelphia chromosome (22)
10
Q
What are genes transcribed into?
A
- mRNA (to be translated to proteins)
- structural RNAs (rRNA or tRNA)
- regulatory RNAs (microRNAs or Xist)
11
Q
What is Xist?
A
X Inactivation Specific Transcript, a regulatory RNA that switches off a copy of X in XX cells
12
Q
What are introns and exons?
A
Introns - Non-coding, spliced out.
Exons - Coding, make it to the mature mRNA.
13
Q
Why was the Puffer Fish genome sequenced?
A
Fugu is very economic in it’s DNA - few kilobases per gene
14
Q
What is a ‘gene desert’?
A
Large intergenic regions in the genome, may be control elements for diseases such as Huntingtons.
15
Q
What is Imprinting?
A
When one copy of a gene inherited from a parent is switched on but the other parents copy is switched off.
16
Q
What is a retrovirus?
A
RNA viruses that insert a DNA copy of their genome into the genomic DNA of the cells that they infect.
17
Q
What is a Pseudogene?
A
Stretches of DNA that have sequence in common with functional human genes but which are non-functional
18
Q
How do pseudogenes arise?
A
Sometimes genes are duplicated and acquire mutations that make them inactive.
Reverse Transcription can also occur, copying an mRNA back to DNA, but these pseudogenes lack promoters so are unlikely to be transcribed.
19
Q
What is a VNTR?
A
Variable Number Tandem Repeat - multiple copies of short sequences that can be amplified by PCR
20
Q
How are VNTRs generated?
A
When DNA polymerases copy highly repeated sequences slippage can occur - they insert too many or too few copies of a repeat sequence
21
Q
What is Huntingtons caused by?
A
Expansion of a long CAG repeat in the Huntington protein gene
22
Q
How do we look for genes causing single gene disorders?
A
Exome sequencing.
23
Q
What is a telomere?
A
A type of repeat sequence which protects the ends of chromosomes and is involved in regulating the number of divisions a cell can make
24
Q
What are SNPs?
A
Single Nucleotide Polymorphisms. A polymorphism is a difference present in 1% or more of the population
25
What is a GWAS?
Genome Wide Association Study - works out the disease rink in many individuals with various conditions.
26
What does ANRIL appear to regulate/influence?
Risk of heart disease and diabetes
27
What is the evidence for DNA's biological role?
-Griffith
-Avery
-Hershey and Chase
(experiments ranging from the 1920s to the 50s)
28
What was Griffiths experiment? (1928)
Used two strains of Streptococcus pneumoniae - one rough (non-virulent) and one smooth (virulent). Figured out that when heat killed smooth strain was combined with smooth strain it could somehow pass the virulence onto the non-deadly strain.
29
What was Avery's experiment? (1944)
Showed that DNA was a transforming agent because out of purified DNA, RNA, protein, lipid and carbohydrate only DNA could induce virulence in non-virulent strain.
30
What was the Hershey & Chase experiment? (1952)
Confirmed the role of DNA as the genetic material by labeling the protein and DNA components of bacteriophages with different radioactive molecules.
31
What was the Chargraff experiment? (1952)
Base composition of DNA - found that % of adenine and thymine and guanine and cytosine were always very similar.
32
What did Watson and Crick do? (1953)
Figured out the 3D structure of the DNA molecule + suggested copying mechanism. Rosalind Franklin managed to get X-Ray diffraction image of DNA
33
What is the DNA structure?
- Antiparallel strands form double helix
- Sugar phosphate backbone
- Base pairs join complementary strands together by hydrogen bonding
34
What are the building blocks of DNA?
Nucleotides. Consist of base, sugar and phosphate. Sugar is deoxyribose.
35
What is the phosphate backbone?
- Backbone of DNA is a sugar phosphate polymer
- Adjacent deoxyribose sugars are linked by phosphodiester bonds
- 5' and 3' ends give the strands directionality
36
What is the larger scale packaging of DNA?
- DNA is tightly coiled around histone proteins, forming chromatin
- Active genes more loosely coiled than silent ones
37
What is meant by semiconservative DNA replication?
Each new double stranded DNA molecule contains an original template strand and a newly synthesised complementary strand.
38
What is the replication fork?
The origin of replication, site which utilises a dynamic structure.
39
What happens at the replication fork?
- helicase unwinds the double stranded DNA to allow replication to occur
- single strand binding proteins stabilise the denatured DNA
- DNA primase synthesisses a short RNA primer to allow replication to commence
- DNA polymerase III carries out the elongation of the new strand of DNA which forms by complementary base pairing to the template strand
40
Which direction does DNA synthesis occur in?
5' to 3' end, requiring a free 3'-OH initially provided by the RNA primer
41
How is the lagging strand of DNA synthesised from?
As DNA can only be synthesised 5' to 3' end, short strands must be synthesised on the lagging strand, each with it's own RNA primer.
DNA polymerase I then replaces the RNA primers with DNA and DNA ligase seals the gaps between fragments.
42
How are errors in DNA replication corrected?
DNA polymerase has a 3' to 5' editing function to remove incorrectly inserted bases. (reducing error to 1x10^7)
A further check for mismatched bases is made by other enzymes (reducing error to 1x10^9)
43
How do mutations arise?
- Spontaneously due to error in replication
| - Induced by DNA damage (eg by radiation or mutagens)
44
What is Ethyl Methane Sulphonate (EMS)?
A mutagen which alkylates DNA, causing a GC-->AT mutation.
45
What are the types of gene mutation?
- Base substitution
- Base insertion
- Base deletion
- Base rearrangement
46
What are the consequences of mutation?
Germ cell - can be inherited
| Somatic cell - may lead to cancer
47
How is DNA repaired?
- Base excision repair proteins cut out damaged bases - they are specific to specific damage types
- Nucleotide excision repair proteins are less specific and cut out sections of the damaged DNA strand
- DNA polymerase I replaces the DNA by copying the intact strand, and DNA ligase seals the gaps.
- Uracil N-Glycolase recognises Uracil in DNA and cuts it out
48
What is a Polysome?
mRNA in the cytosol become covered in ribosomes - several proteins made of the same mRNA simultaneously.
49
Exocytosis vs Endocytosis
Exo- out
| Endo- in
50
What is a lysosome?
Full of degenerative enzymes, they degrade old damaged cellular components and some molecules imported into the cell.
51
Where are most mitochondrial proteins made?
The cytosol, as the majority are encoded by nuclear genes and then imported into the mitochondria. However mitochondria has it's own ribosomes that translate mRNA synthesised in mitos.
52
What are the 3 mammalian RNA polymerases?
RNA Polymerase I - transcribes ribosomal RNA
RNA Polymerase III - transcribes tRNA
RNA Polymerase II - transcribes mRNA, microRNAs and non-coding RNAs.
53
What is a TATA box?
Eukaryotic promoter element, short run of T and A bases. Used as they form the lowest energy base-pairs and so are the easiest to unwind. TATA box is usually located about 25bp (very close) upstream from the start of transcription.
54
What is a CpG island?
Stretch of DNA where there are multiple points where a C in followed by G. These occur upstream of many genes and may have promoter activity.
The C's within a CpG island will be protected from methylation.
55
How can you inactivate a gene expression?
Methylate it's CpG island (happens in X-inactivation)
56
What is polyadenylation?
In eukaryotes transcription continues past the point where multiple polyadenylation sequences are present. The mRNA is then cut near the poly A sequence and a polyA tail is added.
57
What does a mature eukaryotic mRNA looks like?
5' end cap nucleotide -> non-coding -> coding sequence -> non-coding -> polyA tail
58
What is polycistronic mRNA?
In prokaryotes, one mRNA will often encode multiple proteins which are required for a particular task. They are separated by non-coding sequences.
59
What is splicing?
Removal of intronic sequences from the pre-mRNA.
60
How are introns removed?
By the spliceosome. There are specific splice sites at the start and end of each intron.
GU dinucleotide at the 3' end of the upstream intron and AG dinucleotide at the 5' end of the downstream intron.
61
What is an LCR?
Locus Control Region, special sequences that isolate a gene or group of genes from external influences.
62
What is wobble?
Some amino acids are coded for by multiple codons - some tRNAs recognise more than 1 codon. THE FIRST NUCLEOTIDE OF THE ANTICODON RECOGNISES THE THIRD POSITION OF THE CODON
63
What is the start codon for protein synthesis?
AUG - methonine
| or N-formyl methionine in bacteria
64
What are the three sites on the ribosome?
- Aminoacyl-tRNA binding, or A-site.
- Peptidyl-tRNA binding, or P-site.
- tRNA exit site, or E-site.
65
What are the stop codons for protein synthesis?
UAA
UAG
UGA
66
How does termination occur?
At a stop codon, release factors bind and the polypeptide chain is released from the ribosome.
67
Which antibiotics target prokaryotic ribosomes?
```
Small subunit:
-tetracycline
-spectinomycin
-hygromycin B
-streptomycin
Large subunit:
-Streptogramin B
-Erythromycin
-Chloramphenicol
```
68
What if an mRNA codes for a secretory protein?
A cytosolic ribosome begins translating the mRNA, the first bit of the protein is a stretch of (around) 20 hydrophobic amino acids. This is a signal sequence. Signal recognition particle binds to this and causes the ribosome to dock on the ER. The newly synthesized protein is fed through a channel into the ER as it is translated.
69
What happens to newly-synthesised polypeptides in the RER?
ER proteins are folded.
Disulfide bridges are formed if present.
Sugar side chains are added if the appropriate signals are present.
70
What happens to a newly-synthesised polypeptide in the Golgi body?
Some ER sugars are trimmed and replaced.
71
What is Prenylation?
1- protein is made in cytosol, particular amino acid sequence at the carboxyl terminus.
2- this sequence results in part of the carboxul terminus being cleaved off and a fatty acid chain being transferred to the sulfur atom of a cysteine residue.
3- fatty acid tail is believed to bury in the lipid bilayer.
72
What are TOM and TIM?
If a protein is destined for the mitochondrial matrix it is threaded through two channels called TOM (translocase of the outer membrane) and TIM (translocase of the inner membrane)
73
What is cancer?
A group of diseases that includes solid tumours at almost any site of the body, as well as leukaemias. Affects the bodys own cells.
74
What is cancer caused by?
An uncontrolled division of abnormal cells in a part of the body.
75
What are the features of cancer on a cellular level?
- Uncontrolled cell division
- Change in morphology
- Dedifferentiation of cells
- Cell mitigation into adjacent and distant tissues
76
What allows cancer cells to divide indefinitely?
Cancer prevents cell senescence by switching enzyme telomarase back on.
77
What is cancer morphology?
Cancer cells are visually different to normal cells.
78
What are the characteristics of cancer cells grown in the lab?
- Uncontrolled growth
- Loss of contact inhibition
- Immortality
79
What does 'loss of contact inhibition' mean?
Cancer cells do not stop growing once they reach the edges of the dish. They do not have have controls to stop them growing.
80
What are the stages of cancer development?
Normal cells - Hyperproliferative cell population -Early adenoma - Late adenoma - Carcinoma
81
What does metastatic mean?
Cancer cells can move to other parts of the body.
82
What is proliferation?
The growth of cells. Therefore hyperproliferation refers to abnormally fast growth of cells.
83
What are the common genetic abnormalities?
- Point mutation (base substitutions)
- Deletion
- Insertion
- Gene amplification
- Chromosomal translocation
- Aneuploidy
84
What are point mutations?
A base substitution. Smallest change in DNA sequence that can change gene function. Can result in amino acid substitution or introduce a stop codon.
85
What are insertions/deletions?
Can range in size from a single pair to a huge swathe of chromosome. Can result in a frameshift mutation.
86
What is gene amplification?
Can result in a cell having anything up to a hundred copies of a gene it is only meant to have two copies of.
87
How can chromosomal translocations cause genetic mutations?
Genes may be moved to more transcriptionally active regions of the chromosome, or two genes may be combined into a new gene fusion.
88
What is anueploidy?
Any departure from the normal structure or number of chromosomes.
89
What does mutation induction require?
- Chemical modification of DNA
- Replication of DNA
- Resulting in misincorporation by DNA polymerase
90
What differentiates normal cells from cancer cells?
- they are induced to divide by +ve growth factors
- they are prevented from dividing by -ve growth factors
- have a finite lifespan
- have a self destruct mechanism (apoptosis)
- have no influence on blood vessel formation
- are tightly joined together and immobile
- are genetically stable
91
What are the 'hallmarks' of cancer cells?
- genetic instability
- invasive and metastatic
- angiogenic
- resistant to apoptosis
- immortal
- independent of +ve growth factors
- resistant to -ve growth factors
92
What does angiogenic mean?
Refers to cancer cells that have the ability to make blood vessels grow.
93
What is an oncogene?
Mutated versions of normal genes that play a key role in promoting growth and division of cells - mutations lead to increased activity/inappropriate switching on of cell division
94
What are tumour suppressor genes?
Genes that play a role in controlling growth/protecting cells against damage - mutations that inactivate both copies of these genes are involved in carcinogenesis
95
What causes cancer?
- Old age
- Genetics (cancer syndromes)
- Environmental agents
96
How may age cause cancer?
Your risk of cancer increases with age - the more often cells are replicated the higher the chance of a cancer causing mutation - these risks accumulate.
97
What are two examples of cancer syndromes?
- Li Fraumeni Syndrome
| - Xeroderma pigmentosum
98
What percentage of cancers are accounted for by cancer syndromes?
5%
99
What does Xeroderma pigmentosum cause?
High susceptibility to sunlight and risk of skin cancer, as DNA is unable to repair damage caused by UV light.
100
What types of environmental agents cause cancer?
- biological
- chemical
- physcial
101
What are some examples of biological environmental agents that cause cancer?
- Hepatitis B virus
- Human Papilloma virus
- H. pylori
102
Which cancer is Hep B associated with?
liver cancer
103
Which cancer is HPV associated with?
Cervical cancer
104
Which cancer is H. pylori associated with?
Stomach cancer
105
What are some examples of chemical environmental agents that cause cancer?
- Cigarette smoke
| - Heterocyclic amines in cooked meat
106
Which cancer is cigarette smoke associated with?
-lung
-bladder
others
107
Which cancer are heterocyclic animes in cooked meat associated with?
Colon cancer
108
What are some examples of physical environmental agents that cause cancer?
- UV in sunlight
| - X-rays/gamma rays
109
Which cancers are X and gamma rays associated with?
Leukaemia
110
What is a carcinogen?
Any agent that significantly increases the risk of developing cancer - they are often genotoxic.
111
What does genotoxic mean?
How some carcinogens damage DNA and cause mutations
112
How do we know that something causes cancer?
- Experiments (animal/in-vitro)
| - Epidemiological studies
113
What are case-control studies?
Compare disease groups with matched control groups to look for factors more common in the people with the disease
114
What are prospective studies?
Follow a population over time to confirm that the disease is linked to the suspected cause
115
What is a cohort study?
Healthy people are recruited and followed over time. Data collected about exposure is later used to establish whether there is an association between the exposure and the disease.
116
How does smoking cause cancer?
Benzo[a]pyrene binding to deoxyguanosine to form a DNA adduct, this chemical can be edited in a way that it can react with DNA
117
Why has the incidence of melanoma skin cancer increased 4x in men and 3x in women in the UK over 25 years?
It has coincided with a huge increase in the number of holidays abroad
118
What is one way UV damages DNA?
The thymine dimer - where the input of energy from the UV light causes the formation of covalent bonds between adjacent thymine molecules
119
What are steroid hormones?
Derived from cholesterol
120
What are thyroid hormones?
Derived from tyrosine within the protein thyroglobulin
121
What is an androgen?
Any of a group of steroid hormones which promote the development and maintenance of male characteristics of the body
122
What is an oestrogen?
Any of a group of steroid hormones which promote the development and maintenance of female characteristics of the body
123
What are the subgroups of steroid hormones?
Oestrogens and Androgens
124
What is an example of an androgen?
Testosterone
125
What is an example of an oestrogen?
Oestradiol
126
Where are receptors for a specific hormone found?
Only target cells will have a receptor specific to the hormone - the receptor may be in the cytosol or nucleus.
127
What is the mechanism of steroid hormones?
- Steroid hormone crosses cell membrane
- Binds to intracellular receptor in nucleus/cytosol
- If bound in the cytosol moves to nucleus
- The hormone-receptor complex acts as a transcription factor
- mRNA is transcribed
128
What are zinc fingers?
Loops of protein that contain a Zn2+ ion and in the case of steroid hormone receptors are coordinated with 4 cysteine residues
129
How do hormone receptors interact with other proteins that regulate transcription?
They have a transcription regulation domain at the amino terminus.
130
What makes nuclear hormone receptors specific?
They have a hormone binding domain at the carbonyl terminus.
131
What is a dimerization domain?
Two dimerization domains on two nuclear hormone receptors come together with the factors facing opposite directions, so that they are interacting with opposite strands on the DNA
132
What does healthy breast tissue express more of?
ER-beta oestrogen receptors rather than ER-alpha. Some types of breast cancer strongly express ER-alpha.
133
What can oestrogen promote in breast tissue?
Rapid division of ER-alpha positive breast cancer cells.
134
How can ER-positive breast cancers be detected?
By staining sections of tumour biopsies with antibodies against the oestrogen receptor.
135
What is tamoxifen?
Ab anti-oestrogen cancer drug that inhibits the growth and spread of ER+ breast cancers.
136
How does tamoxifen work?
It is a pro-drug that becomes hydroxylated within the body and activates. It then binds to oestrogen receptors.
137
What is a pro-drug?
A biologically inactive compound which can be metabolized in the body to produce an active drug.
138
What are aromatase inhibitors?
Drugs used to treat oestrogen sensitive tumours by blocking the formation of the aromatic ring in oestrogens
139
What are the two ways of treating ER+ breast cancer?
- tamoxifen as an inhibitor to the oestrogen receptors
| - aromatase inhibitors to block the formation of the aromatic ring in oestrogens
140
Which cell surface receptor do some breast cancers change the amount of?
HER2 - this binds to a epidermal growth factor
141
What is Trastuzumab and how does it work?
An anti-cancer drug that attaches to HER2, stopping the cancer cells from growing and dividing
142
How are thyroid hormones synthesized?
- By iodinating tyrosine molecules on the thyroglobulin
- Cross-linking iodotyrosine molecules in the thyroglobulin
- Cutting out the thyroxine
143
Where are all thyroid hormone receptors found?
Bound to the DNA
144
What is the TRE?
Thyroid hormone recognition/response element , where thyroid hormone receptors bind on the DNA
145
Why are there two separate genes for thyroid hormone receptors?
They have slightly different actions from each other
146
How do retroviruses become capable of causing cancer?
By capturing a host version of a protein that is involved in growth control, and moving onto the next host with it.
