chapter 11 part 3 Flashcards
when do we do non-homologous end joining?
when there is a DSB that occurs during G1 (before replication) which prevents completion of DNA replication
what does non-homologous end joining allow for?
cells to regain the ability to complete DNA replication, though we will have mutations for sure
non-homologous end joining process
4 steps
-DSB happens
-DSB are recognized by the proteins PKcs, Ku70, and Ku80 that attach to the broken ends of the DNA
-the complex trims back the free ends of the break- LOSS OF GENETIC INFO
-the blunt ends produced by resection are ligates by ligase IV
when does synthesis-dependent strand annealing SDSA occur?
we do it when there are DSBs that occur after replication by an error-free process
what is synthesis-dependent strand annealing also called?
homology dependent repair (HDR)
what does the strand invasion process do?
displaces one strand of the DNA duplex on the sister chromatid, forming a displacement (D) loop
-replication within the loop synthesizes new DNA from the intact template strand
what is the process of SDSA
-SDSA begins with trimming the broken ends, followed by attachment of protein Rad51 (a homolog of bacterial RecA protein)
-Rad51 facilitates the invasion of the intact sister chromatid by the resected end of the broken strand
-stand invasion process
-sister chromatids refrom
how do sister chromatids reform in SDSA?
by dissociation & annealing of the new strand to the other side of the break, resulting in replacement of the excised DNA with a duplex identical to the sister chromatid
homologous recombination
the exchange of genetic info between homologous DNA molecules
homologous recombination in bacteria
occurs during conjugation & as a consequence of DSB repair (archaea as well)
homologous recombination in eukaryotes
takes place in prophase 1 of meiosis
what is homologous recombination initiated by in eukaryotes?
double-strand DNA breaks
what does proper chromosome segregation in meiosis depend on?
homologous recombination
-without it, errors like nondisjunction are likely to occur
what is the system of homologous recombination in bacteria?
RecBCD pathway
what does RecBCD rely on?
DNA double-strand breaks to innitiate the process
-this attracts the RecA protein (homolog of human Rad51 protein)
-RecBCD then attaches to the region where RecA is bound, leading to single-strand invasion & D loop formation (similar to SDSA)
-RuvAB & RuvC proteins bind & complete homologous recombination
DSB model of meiotic recombination simplified
- recombination initiated by Spo11
- Spo11 degrades, Mrx cuts single strand of cut chromatin
- Rad51 & Dmc1 facilitate strand invasion & D loop formation
- holliday junction formed between 2 strands that appear to cross over
- heteroduplex forned
- invading strand in extended with DNA synthesis guided by intact template strands assisted with Rad52 & Rad59
- 3’ end of invading strand joins 5’ end of a strand segment that was initially invading strand (ligation)
- non-sister chromatids are connected by double holliday junctions (DHJs)
heterodulex
a double-stranded DNA formed from single stranded pieces of DNA of different homologs
how many ways can holliday disjunction be resolved
2
what are the way holliday disjunction can be resolved
-same sense resolutions
-opposite sense resolution
same sense resolution
when two north-south (NS) resolution cuts or two east-west (EW) resolution cuts occur
-flanking markers DO NOT recombine though heteroduplex regions remain only in between junction points
opposite sense resolution
a resolution in which one Holliday junction is resolved by a NS cut & the other by an EW cut, is much more common
-resulting chromosomes are recombinant, & lead to production of recombinant progeny = major changes in chromosome segments
-ends swap
what of the two holliday junction resolutions is more commin?
opposite sense resolution
what are transposable genetic elements?
DNA sequences that can move within the genome by an enzyme-driven process, transposition
what do transposable genetic elements vary in?
length, sequence composition, number
what ways does movement occur in transposable genetic elements?
- excision of the elements from its original location & insertion in a new location
- duplication of the element & insertion of the copy in a new location
insertional activation
when a transposable element can cause a mutation if it inserts into a wild-type allele & disrupts its function
what do all transposable elements have in common?
- transposable element contains terminal inverted repeats on its ends
- the inserted transposable element is bracketed by flanking direct repeats
typical transpostion event
-staggered cuts are made in both strands of DNA at the new target site for transposable element insertion, leaving short single stranded overhangs on each end of cut
-staggered cuts are made by transposase, produced by the transposable element
-the double-stranded transposable element is inserted into its new site
-dna is replicated at the new site of insertion to fill the single-stranded gaps, producing flanking direct repeats
what are the categories of transposable elements?
DNA transposons
Retrotransposons
DNA transposons
(class II transposable elements)
-transpose as DNA sequences & may be replicative or non-replicative
Retrotransposons
-(Class I transposable elements)
-composed of DNA but transpose through an RNA intermediate
-rna is coped back into DNA by reverse transcriptase
-the reverse-transcribed DNA is then inserted into a new location where flanking directs repeats are formed
non-replicative transposition
excises a transposable elements from one position & inserts it into a new location
-cut & paste !!!!
-the process moves transposable elements around the genome but results in NO increase in the number of transposable elements
replicative transposition
increase the number of elements per genome
-COPY & paste !!!!!
-a recombination-like process resolves the cointegrate, & leaves both plasmids with a copy of the element
what are the three categories of bacterial transposons
-insertion sequences (ISs)
-composite transposons (Tn in bacteria)
-non-composite transposons
insertion sequences
simple transposable elements containing terminal inverted repeats surrounding a gene (sometimes two genes) encoding transposase
composite transposons (Tn in bacteria)
carry a transposase gene, two flanking IS elements, & one or more additional genes
-larger than IS element
-often contain resistance genes
non-composite transposons
-similar to composite transposons but lack the IS element
-typically have additional genes like composite transp.
are there a lot of eukaroytic transposable elements?
yes, they are highly varied with two types
-short sequences with inverted repeats
-retrotransposons
how much of the human genome is composed of transposable DNA?
nearly half
what do retroviruses do?
infect eukaryotic cells & have genomes composed of single stranded RNA (ssRNA)
-on infection the RNA is transcribed into double stranded DNA (dsDNA) by reverse transcriptase, allowing the DNA to parasitize the host cells
-the gene(s) carried on retrotransposons are flanked by long terminal repeats (LTRs)
what genes are needed to produce new retroviral particles
gag & env encoded by the integrated virus
what gene encodes reverse transcriptase?
pol
do all viral genes contain gag?
no, but all contain pol
are retrotransposons & retroviruses related?
yes
what types of retrotranposons are found in yeast?
Ty
-these can cause insertional mutations
-central element is about 6kb long & is flanked by long terminal repeats of about 330 bp
-both LTRs contain promoters that direct transcription fo different genes in the central region
multiple forms of ______ elements are found in the drosophila genome
copia
-have a central element of 5-8.5 kb that contain gag & pol genes & are flanked by 250-600 bp LTRs
*more than 5% of the total genome is comprised of various types of copia elements
how much of the human genome is composed of transposable DNA?
more than 45%
LINEs
- Long interspersed elements
SINEs
short interspersed elements
what do LINEs & SINEs do
they are relatively abundant & can cause mutations in humans
what is a particularly common LINE
L1 elements (600,000/genome)
-associated with spontaneous human mutations
-encode proteins w nuclease & reverse transcriptase function & RNA-binding protein
-6.5-8.0 kb
what is the most common SINE
Alu elements - they actively generate mutations
-vary in length from 100-300 bp & are flanked by 7-20 bp direct repeats
-more than 1 million of these elements
-active in 1 in 200 ppl & responsible for 0.3% of human hereditary disease
