1.6 - DNA Replication; Regulation and Repair Flashcards
origin of replication in bacteria
DnaA
origin of replication in eukaryotes
Orc/Cdc6
initiator
DNA binding protein complex that prepares origin for first step of replisome assembly
what does initiator promote in some prokaryotes?
some unwinding, necessary for recruitment of helicase into DNA
single start site on bacterial chromosome
oriC
effect of binding of initiator protein to DnaA
triggers process of opening by modifying local organisation of initiator sequence to promote DNA unwinding and helicase binding
how many proteins are involved in start of DNA synthesis in bacteria
at least 10
how is DNA replication controlled in bacteria?
DNA methylation, another round will not start until new strand (of oriC) is methylated
yeast sequences vs humans (2)
- yeast - consensus sequences
- humans - do not appear to have consensus sequence but frequently have same chromatin modifications
role of ORC and CDC6 proteins
(similar to DnaA) bind to origin and recruit replisome proteins
role of cyclin proteins
“license” initiation and ensure each origin only used once per cell cycle
how do cyclin proteins ensure each origin is only used once per cell cycle?
in part by phosphorylating ORC proteins
telomeres
eukaryotic chromosome ends
what is present at telomeres?
multiple end-to-all repeats of a short TG-rich sequence TTAGGG (most repeats double stranded but 3” end extends beyond 5” as single strand
how can the 3” end of telomeres be extended?
by telomerase (enzyme containing a short RNA molecule complementary to the TTAGGG repeats
what does the RNA molecule in telomerase act as?
a template for “reverse transcription”, copying the RNA into DNA (thus extending the 3” strand of the telomere)
how is the 5” strand at the telomere extended?
usual way, with an RNA primer and DNA polymerase
difference between telomeres in young/old people
longer in young people
telomerase activity in adult somatic cells/ stem cells and germ cells (2)
- adult somatic cells - lose telomerase activity
- stem/germ cells - retain telomerase activity
what do cancer cells have to do once they start dividing?
activate telomerase to escape rapid senescence
effect of mutagens?
(chemicals, UV light etc) - continuously damage DNA generating thousands of lesions per cell per day
cancer cause
somatic cell mutations
why does risk of cancer increase over lifetime?
cell mutations build up
cause of early life cancers/ other genetic diseases
often due to faulty DNA repair mechanisms (cause early onset pathologies due to a high mutation burden)
how does persistent chromosomal damage lead to genome instability?
chromosomal breaks can lead to miss-repair and re-arrangements with other chromosomes
primary non-replicative causes of DNA damage (4)
- alkylation
- depurination
- deamination
- UV irradiation
DNA repair mechanisms (3)
- Base-excision repair
- Nucleotide-excision repair
- Double strand break repair
common damaging event to DNA
deamination of C to U
why is deamination of C to U common? (3)
- in ancient cells U was likely only base that could pair with A
- DNA repair enzymes would not have been able to distinguish “real” from “damages” U (derived from C)
- leading to frequent mutation
result of replacing U with T
means all U in DNA must be damage and therefore repaired by base excision repair
base excision repair (3)
- detects uracil (or other altered bases)
- an endonuclease excises the abasic site
- “hole” in chain filled by DNA polymerase/ligase
(substantial length of one strand removed and replaced)
importance of nucleotide excision repair
important for removing lesions such as thymidine dimers that distort double helix (critical for survival)
disease arising from defects in nucleotide excision repair (3)
- xeroderma pigmentosum
- patients very sensitive to light and UV-induced DNA damage
- skin cancers common
exogenous causes of DNA double strand break (2)
- radiation
- chemicals
endogenous causes of DNA double strand break (2)
- oxygen - free radicals
- DNA replication
specialised causes of DNA double strand break (3)
- V(D)J recombination
- class switching
- meiosis
cellular outcomes after DNA break formation (3)
- incorrect -> genome instability -> carcinogenesis
- cell death (apoptosis)
- repair
one of the earliest markers of uncontrolled cell growth
accumulation of broken chromosomes
how do DNA lesions affect DNA replication?
leading strand polymerase cannot get past block (stalls replication)
how can DNA lesions be bypassed? (3)
- bypass DNA polymerase binds to lesion
- bypass polymerase makes DNA opposite the lesion
- replication restarts
positive notes on mutation (3)
- if 95% of DNA is “junk” most mutations wont be in coding regions (1-2 still will be and some junk matters)
- even in coding regions many mutations don’t change protein function
- 2 copies of each gene
