24: Genes and Chromosomes

Question 1

what is encoded in our genome?

Answer 1

1. coding RNA: RNA transcribed from DNA that code for proteins (only 1% of genome)
2. non-coding RNA (ncRNA): functional RNA molecule that is transcribed from DNA but not translated into proteins. regulatory role
3. transposons: mobile DNA elements (45% of genome!)
   less than 30% of the human genome contains genes, and only a small portion is used to code protiens

Question 2

define gene. where do genes start/stop? how does a gene encode a protein?

Answer 2

gene: all the DNA that encodes the sequence needed to produce some final gene product, RNA or protein product.
specific sequences indicate the machinery to start/stop
how to encode a protein: each amino acid of a polypeptide is encoded by 3 consecutive bases of a single stranded DNA (codon)

Question 3

how does our nucleotide/amino acid system display colinearity?

Answer 3

colinearity: consistent translation of DNA to RNA to Protein. this is achieved through the three base codon system that consistently achieves protein production out of a DNA code

Question 4

what are introns?

Answer 4

DNA sequences that are removed during transcription, almost all eukaryotic genes have them. bacteria have some introns, but not interrupting coding genes like eukaryotes do.

Question 5

what is the open reading frame?

Answer 5

ORF is the part of a reading frame that has the ability to be translated. An ORF is a continuous stretch of codons that begins with a start codon (AUG) and ends at a stop codon

Question 6

describe the packaging of genes (very general)

Answer 6

genes are packaged in chromatin; Genomic DNA with lots of proteins that together can form a condensed structure. Chromatin forms into even more dense structure called chromosomes.

Question 7

describe genome differences between prokaryotes and eukaryotes

Answer 7

size: generally much larger in eukaryotes (BUT number of genes is not a predictor of complexity!)

Question 8

what is the natural intrinsic coiling of DNA?

Answer 8

DNA is intrinsically supercoiled, when purified it still remains supercoiled.

Question 9

how is supercoiling induced?

Answer 9

strand separation: if strands pull apart in one section, it causes supercoiling in areas around it. strand separation occurs with replication/transcription and so both affect and are affected by supercoiling
underwinding: most common. remove a turn from DNA to cause strain, strain is accommodated by allowing the DNA to bend on itself i.e. supercoil. strain could also be relieved by separation of strand

Question 10

how is underwound DNA actively maintained?

Answer 10

DNA must either be circular or bound and stabilized so strands are not free to rotate. this is controlled by topoisomerases.

Question 11

explain linking number

Answer 11

DNA underwinding is defined by linking number: the number of times the 2nd strand pierces the imaginary surface created by the 1st strand. Or, the # of base pairs / # base pairs per turn
to change Lk, you must break the DNA!

Question 12

ways to measure coiling

Answer 12

superhelical density
specific linking difference
sigma = ∆Lk/Lk0

Question 13

what are topoisomers

Answer 13

2 forms of DNA that differ only in linking #. topoisomers can be positive or negatively coiled or relaxed

Question 14

define topoisomerases (type I and II)

Answer 14

topoisomerases change the linking number of DNA by breaking the strand
type I: transiently breaking 1 strand, changes by increments of +1 Lk. bacterial (?)
type II: breaks both strands, changes by increments of -2 Lk. eukaryotic (?)

Question 15

mech of type I topoisomerase

Answer 15

slide 20-22
DNA binds. an active site Try attacks the phosphodiester DNA bond, cleaving it and creating a covalent 5’-Tyr protein DNA linkage. Enzyme switches to open conformation and the unbroken DNA strand passes through the break. Enzyme switch to closed conformation, 3’ hydroxyl group attacks the 5’-Tyr linkage to ligate the cleaved strand

Question 16

mech of type II topoisomerase

Answer 16

slide 23

same mech as type I but cleaves two strands and moves double strand through break. consumes 2 ATP!!

Question 17

what is DNA gyrase

Answer 17

a type II topoisomerase that introduces negative supercoils (decreases Lk). I think its used during replication?

Question 18

what is plectonemic and solenoidal?

Answer 18

types of DNA compaction, slide 25 for visual

plectonemic: twisted like a double helix, is naturally more stable and requires no proteins to form
solenoidal: twisted like slinky, stabilized by proteins and provides more compaction. best option for storage

Question 19

what constitutes chromatin?

Answer 19

DNA, protein (histones especially), some RNA. these elements from nucleosome ‘beads’ which form chromatin.
one nucleosome contains one histone core and some DNA wrapped around it

Question 20

what are histones? what’s their structure?

Answer 20

histones are small basic proteins with lots of Arg and Lys residues. tails can be highly posttranslationally modified, which contributes to a histone code
there area 5 classes in all eukaryotic cells. these compose the nucleosome histone core with 2 of each except H1, which interacts with linker region

Question 21

describe the importance of the histone tails

Answer 21

the tails serve as ‘landing pads’ for a variety of proteins to bind that could affect the rate of transcription of nearby genes. tails are super long and highly modified

Question 22

what does tight DNA wrapping form

Answer 22

left handed solenoidal supercoil. requires removal of about 1 helical turn in the DNA by topoisomerase

Question 23

how does histone binding affect supercoiling?

Answer 23

solenoidal negative supercoils are induced by DNA binding to the histone. to balance this out, a topoisomerase relaxes by one plectonemic positive supercoil. end result is 1 net negative supercoil
slide 32

Question 24

describe interactions that affect DNA binding to histones

Answer 24

composition and modification of histones (histone variants exist and can be utilized to alter DNA binding)
topography of DNA
DNA composition. higher local abundance of A=T pairs in contact with histones

Question 25

describe packaging steps from nucleosomes to chromsomes

Answer 25

nucleosomes pack into a 30nm fiber
30nm fibers make loops (75,000 bp)
loops associate with a nuclear scaffold and make a rosette of 6 loops
30 rosettes form one coil
10 coils form one chromatid
4 chromatids form a chromsome

Question 26

what is epigenetic and what are some techniques that tell you about histone modifications

Answer 26

epigenetic information: is passed from one generation to the next but is NOT encoded in DNA sequence (includes histone mods or DNA methylation)
ChIP: chromatin immuniprecipitation. show changes in histone PTMs associated with a particular gene, use QPCR to quantify
ChiP-Seq: quantitatively tell which PTMs are associated with which genes in a whole set.
Chip-CHIP: quantitatively tell which PTMs are associated with which genes in a whole set
see slide 37-38 idk

Question 27

compacting of DNA in bacteria

Answer 27

DNA compacted into structure called nucleoid, analogous to a chromosome. nucleoid is formed by tons of loops of DNA. there is no nucleosome or histone core association.
structure is more dynamic and irregular, reflecting shorter cell cycle and more active metabolism