Lecture 22 Flashcards
TMV
- tobacco mosaic virus
- RNA virus
- develops a helical structure as RNA and protein interact
- if you dissociate the RNA and protein it loses its helical structure
Lambda
- bacteriophage
- temperate
- DNA in long strings of copies attached end to end
- as the DNA fills the protein coat, the interaction between the two leads to the coat getting its shape
Esther Lederberg
- first to isolate Lamda
- discovered specialized transduction
concatemer
- strings of copies of Lambda’s DNA attached end to end
E.coli DNA
- 4.6 million base pair DNA
- DNA is associated with positively charged proteins due to high arginine and lysine content
E. coli chromosome structure
- Circular DNA is compacted into 40-50 twisted loops
- each loop is supercoiled and held in place with positive proteins
chromatin
the complex of DNA, chromosomal proteins and RNA within the nucleus
euchromatin
- lighter staining parts of the chromosome during interphase
- consists of actively transcribed genes
- condense and relaxes
heterochromatin
- darker staining parts of chromosome
- fewer genes
- remain condensed
- usually not involved in crossing over
- replicates late in S phase
constitutive heterochromatin
- regions within heterochromatin that are always condensed
facultative heterochromatin
- regions of heterochromatin which may be euchromatic at times
- ex. X chromosome condensed into a barr body
nucleosome
where the DNA wraps around histone proteins in the most basic part of chromatin structure
describe histones
- basic pH proteins
- lots of positively charged amino acids
- highly conserved
histone functions
- involved in packaging the eukaryotic chromosome
- their positive charge binds electrostatically to negatively charged phosphates in DNA
list the histones
H1 H2A H2B H3 H4
Highly conserved
- same or very similar amino acid sequence between organisms
If two proteins from diverse organisms have the same sequence…
it is likely they have the same function in both species
why are histone’s positively charged
lysine
arginine
When the entire amino acid sequence of the proteins in two diverse species are identical…
the entire sequence is important to the proteins functions
Which histone proteins are highly conserved?
H3 and H4
Which histone proteins show species to species variation?
H2A and H2B
Which histone proteins show tissue to tissue variation between species?
H1
chromatsome
nucleosome + H1 histone
nucleosome
nucleosome core + ~53 base pair linker DNA
200bp/nucleosome
nucleosome core
core histones + ~147 bp DNA
core histones
2 each of H2A, H2B, H3, and H4
linker DNA
- the DNA that joins the chromatasomes along the length of a chromatin fiber
A nucleosome core is ___ nucleotide pairs wrapped ___ turns around an octomer of histons
146 nucleotide pairs
1.74 turns
how long is linker DNA normally?
30-40 bp
describe the in situ chromatin structure
- nucleosomes coilin a zig-zagging manner to form a fiber 30 nm in diameter called the 30 nm fiber
- the 30 nm fiber is held in loops in the metaphase chromosome
- it is folded to have loops 300nm in length
- these loops are anchored by non-histone proteins at the loop bases
- they coild into about 700 nm in diameter in the metaphase chromosome
30 nm fiber
the fiber formed by nucleosomes coiling in a zig-zagging manner in the in situ model
in vivo chromatin models
- chromatin fibers are flexible and range from 5-24 nm, smaller than the proposed in situ model
- les organized than the proposed model
nuclear scaffold
- non-histone scaffold of proteins seen after removing histones from chromosome
loops of DNA anchor to the scaffold and DNA is anchored to the EC matrix
nuclear matrix
fibrous network throughout the nucleus that anchors a series of DNA loops
each loop is thought to have 20,000 to 100,000 base pairs
MARs
matrix attachment regions
DNA sequences that bind the nuclear matrix and anchor DNA loops
Proteins that bind to MARs…
may be important for getting the proper conformation for expression
topoisomerase II
- a protein in the nuclear scaffold
- can manipulate coiling of DNA by cutting the backbone - the amount of coiling is important in DNA expression
supercoiling
occurs when DNA coils back on itself when it is over or underwound
it is relieved by topoisomerases
positive supercoiling
- over-rotated
- supercoils in the opposite direction
- left-handed coiling
negative supercoiling
- underwound
- same direction
- right-handed coiling
banding patterns
- produced by treating human chromosomes with enzyme trypsin followed by stain giemsa - most common method - G bands
R bands
reverse of G bands
produced by using microscopy after the G band production
C bands
centromere is standed
treated with sodium hydroxide before the giemsa
polytene chromosomes
chromosomes seen in the salivary glands of drosophila larvae due to endoploidy
endopolyploidy
several rounds of dna replication without separation of replicate chromsomes
puffs and balbiani rings
areas where DNA is loosely coiled so that transcription can occur
centromeres
- must attach to the kinectochore which then binds to spindle fibers
- not a single centromere sequence for eukaryotic chromosomes
an example of an organism with specific centromere sequences
yeast
very AT rich with central region surrounded by conserved regions 11-14 bp long
describe centromere
- heterochromatic
- short, repeated DNA sequences
CenH3
different histone that replaces the normal H3 in the centromere of humans, flies, and Arabidopsis
- seem to change chromatin structure to allow kinectochore binding
telomeres
provide stability for ends of chromosomes so they are not degraded by exonucleases
prevent chromosomes from joining due to ligases
provide proper replication of end of chromosome
tend to be GC rich
human telomeric sequence
5’- TTAGGG -3’
repeated 300-5000 times
telomeric structure
- 3’ end extends further on one strand than the other; a 3’ overhang
- this extension coils back towards the interior and displaces one strand of the DNA to form a loop at the end of the chromosome by pairing with the nondisplaced strand
T-loop
formed by the 3’ overhang coiling back toward the interior of the chromosome and displacing one strand pairing with the non-displaced loop
thought to help protect the ends from degradation and to keep from joining with other chromosomes
Who discovered transposable genetic elements?
Barbara McClintock
she is the only woman to receive an unshared nobel prize in physiology or medicine
transposable genetic elements
“jumping genes”
move from one site to another
can move to different chromosome
can alter phenotypes when they move
How did McClintock discover transposable elements?
Ac-Ds system in Maize
noticed splotches of color in kernels that should be solid based on genotypes
types of transposition
replicative
nonreplicative
conservative
retrotransposons
replicative transposition
uses transposase to move a copy
causes an increase in the number of these sequences over time
nonreplicative transposition
transposase moves the original sequence from its original location but does not repair the DNA from the site it leaves
conservative transposition
transposase moves the sequence from the original position and repairs the original DNA
retrotransposons
uses reverse transcriptase to create DNA from element’s RNA
RNA is produced from DNA sequence in the chromosome
reverse transcriptase produces DNA from this RNA
the new DNA integrates into another space on the chromosome
components of a simple transposable element
- sequence
- terminal inverted repeated
sits between direct repeats which are not part of the element
inverted repeat
the sequence reads the same from 5’ to 3’ on one strand as it does from 5’ to 3’ on the other strand
part of a simple transposable element
direct repeat
the sequence reads the same 5’ to 3’ on one place on a strand as it does later on the same strand in the 5’ to 3’ direction
outside of the simple transposable element
Mechanism for insertion of transposable elements
- staggered cuts are made in target sequence
- the strands separate
- the insertion sequence with inverted repeats on the ends is stuck in the opening
- the gaps are filled by addition of complementary nucleotides producing direct repeats
Ti Plasmid
- a tumor inducing plasmid in plants that causes crown gall disease
- inserts itself into the plant chromosome
- opportunistic pathogen
How does Ti plasmid work?
- contains T-DNA which is a transposable element
- after infection, the T DNA causes DNA from the plasmid to insert into the plant’s chromosome
Do humans have transposable elements?
yes
~45% of genome appears to be remnants of transposons - no longer move
Do transposons have a purpose?
have the potential to move regulatory sequences to new places which could affect gene expression
can cause mutations
purpose of transposons in arbidopsis
Transposase derived proteins regulate plant genes required for plant growth
purpose of transposons in humans
mechanism for antibody formation may have evolved from transposons
purpose of transposons in drosophila
telomerase enzyme is not present in drosophila but ends of chromosomes have transposon like sequences and mechanism is similar
transposons and bacteria
- bacterial genes can move between chromosome and plasmids
- bacterial transposons can carry multiple genes for antibiotic resistance
- once drug resistance has evolved in one species of bacteria, it can spread to another by plasmids
