Genetics Exam 2

Main features of the eukaryotic transcription unit

promoter, coding region, terminator

start sequence in bacteria

AUG

Shine-Dalgarno sequence

bacteria only

recruits 16S rRNA to bind to the sequence

Common structural features for prokaryotic and eukaryotic transcription unit

RNA polynucleotide, all 5’-3’ from non-coding template, 5’ and 3’ UTR, internal coding region, start and stop codons for translation of coding region

difference between prokaryotic and eukaryotic structural features in transcription

prokaryotics - no processing

eukaryotic - processing including 5’ cap, poly-A tail, intron splicing

covalent modifications to eukaryotic mRNA

addition of modified m7G to 5’ end (methyl-7-guanesine)

addition of poly-A tail to 3’ end, 50-250 As

cleavage/trimming to generate ends in eukaryotic mRNA

involved in 3’ poly-A tail addition

splicing (removal of introns)

introns are spliced out

joins exons covalently

gets pre-mRNA to mature mRNA

addition of 5’ cap

adds methyl-7-guanosine to 5’ end

adds 2’ methyl groups to first few nucleotides of RNA strand

functions of 5’ cap m7G

stabilizing (resembles 3’ end, prevents degredation by exonucleases)

nuclear export (cap binding complex)

translation

addition of polyA tail

cleavage near the polyadenylation signal in the 3’ UTR

addition of 50-250 A ribonucleotides

function of the poly A tail

stabilizing, translation

spliceosome

joins exons during or following transcription

consensus sequence

short stretches of common nucleotides, comprises the most commonly encountered nucleotides at each site

how are the introns spliced out of the pre-mRNA

mRNA cut at the 5’ splice site

5’ end of intron attaches to branch point (an A in the intron)

cut is made at 3’ splice site

intron released as lariat

exons spliced together

bond holding lariat is broken, linear intron degraded

spliced mRNA exported to cytoplasm and translated

alternative splicing

splicing of variable exons

isoforms

product of alternative splicing

tRNAs

contain modified bases

amino acid attachment site (3’)

anticodon arm of the cloverleaf

prokaryote stop codon

UAG

Kozak Sequence

ACCAUGG

surrounds start codon AUG

Shine Dalgarno sequence

UAAGGAGGU located about 6 bp before start codon

eukaryote stop codon

UAA

amino acids in protein joined by

peptide bond

directionality of protein

peptide bonds between carboxy (C) and amino (N) ends of the AA

protein is always N terminus - C terminus

degeneracy

one amino acid may be specified by more than one codon

why do 61 sense codons encode 20 AA total

degeneracy, synonymous codons, same amino acid

first and second pase pairs

strict paring

third base pair

wobble

ability to base pair with alternate bases

some tRNAs can pair with more than one codon

how do tRNAs function in the cell

cell has more tRNAs than amino acids

each type of tRNA is charged with specific amino acid

charging done by aminoacyl transferase

A site

aminoacyl tRNA site

holds tRNA carrying next amino acid

P site

peptidyl tRNA site

holds tRNA carrying growing polypeptide chain

E site

empty tRNA

regulated genes

not always expressed

constitutive genes

always expressed, not regulated

housekeeping genes

regulatory genes

encode RNA or proteins that interact with other DNA sequences to affect transcription and translation

lacI

regulatory elements

not transcribed, affect expression of DNA (like structural or regulatory genes) to which they are physically linked

lacO

operator

DNA sequence situated upstream of genes and downstream of promoter

Regulatory element

operator is recognized by

regulatory protein

presence of regulatory protein at operator determines

whether transcription takes place

regulator and promoter

overlap

regulator protein two sites

DNA binding, small molecule binding

negative control

regulation inhibits gene expression

positive control

regulation stimulates gene expression

regulator proteins have two types

repressor and activator

regulator proteins either activate or repress operon, this is

inducible or repressible

if a regulator is allosteric, it

undergoes a confirmation change

inducible operon

transcription is normally off and must be induced

repressible operon

transcription is normally off and must be repressed

negative inducible

repressor normally binds to DNA to turn off transcription, substrate binds to repressor to make inactive so transcription can take place

negative repressible

repressor normally can’t bind to DNA to turn off transcription, substrate binds to repressor to make active so transcription is turned off

positive inducible

activator normally can’t bind to DNA to turn on transcription, substrate binds to activator to make active so transcription is turned on

positive repressible

activator normally can bind to DNA to turn on transcription, substrate binds to activator to make inactive so transcription is turned off

lac operon

negative inducible, encodes genes that metabolize lactose when lactose is present in the cell

what does it mean that the lac operon is basal

lac operon is transcribed always, background or noise, a small amount of lac proteins are always present

inducer for lac operon

allolactose

product of lactose from lacZ

what happens when allolactose binds to repressor

repressor can’t bind to operator, transcription of proteins needed to metabolize lactose takes place

catabolite repression

glucose has inverse relationship with cAMP (cyclic AMP)

when CAP bound to cAMP, promotes transcription

when cAMP not available, CAP does not bind and transcription is low

when lactose is absent

there is very little Lac gene production

when lactose is present but a preferred carbon source is also present

then a small amount of enzyme is produced

when glucose is absent

CAP-cAMP binds to a specific DNA site upstream of the promoter and makes a direct protein-protein interaction with RNAP that facilitates the binding of RNAP to the promoter

mechanisms for altering of chromatin structure

chromatin remodeling

modifications to the histone tails

DNA methylation of cytosine bases

covalent modification of histone tails

addition or removal of phosphate, methyl, acetyl, and ubiquitin groups

histone code

changes (individual or combinations) encode information that affects gene expression

acetyl groups on a histone tail

promote transcription

methyl groups on a histone tail

activate or repress transcription

phosphate groups on a histone tail

have a role in miotic chromatin condensation and apoptotic chromatin

DNA methylation

represses transcription

eukaryotic promoters

core promoter, upstream of gene to be transcribed, binding site for basal transcriptional apparatus

regulatory promoter, located upstream of core promoter, binding site for transcriptional activator proteins

basal transcriptional apparatus

binds core promoter

comprised of general transcription factors and RNA polymerase

capable of minimal levels of transcription

transcriptional activators and repressors

bind elements within the upstream regulatory promoter

associated with the basal transcriptional apparatus via direct contact or through indirect interactions

activators stimulate transcription, repressors inhibit transcription

enhancer

regulatory element that is not physically associated with the core promoter and is located elsewhere on the chromosome

silencer

regulatory element that is recognized and bound by repressor proteins that act to repress transcription

somatic cell mutation

body cells, cannot be passed to offspring, can cause cancer in genes that control cell division

germ cell mutation

sex cells, can be passed to offspring

transition

purine to purine (AG, CT)

transversion

purine to pyrimidine (A to C or T)

tautomers

alteration of base in DNA sequence

spontaneous mutations

occur naturally under normal conditions

depurination

loss of purine base due to breakage of 1’C bond

deamination

loss of AA group from base

thymine dimer

disrupts replication via UV rays, leads to cell death

nucleotide excision repair

pathway has specialized enzymes that recognize thymine dimer distortion

bases are excised

DNA polymerase fills gap

ligase seals nick in phosphodiester backbone