Eukaryotic Gene Expression Flashcards
define gene expression / the central dogma?
the process in which the information within a gene is used to synthesise, first RNA (in transcription) then a polypeptide (in translation) eventually to express the phenotype of an organism
what are the similarities between RNA and DNA? MEMORISE
- both polynucleotides
- sugar-phosphate backbone joined by phosphodiester bonds
- 3 bases: AGC
- complementary base pairing
- formed via condensation reaction
compare the differences between RNA and DNA MEMORISE
- molecular mass: RNA smaller (20-2000 kDa), DNA larger *100-150000 kDa)
- RNA is single-stranded, DNA is double-stranded
- monomers: RNA’s is ribonucleotides, DNA’s is deoxyribonucleotides
- chemical stability: RNA is less stable with additional reactive 2’ OH group, DNA lacks it and is more stable
- Nitrogenous bases: RNA has uracil and DNA has thymine
- ratio of purines to pyrimidines: RNA not 1:1, DNA is 1:1
- basic form: RNA has multiple (eg. mRNA, tRNA, rRNA), DNA has one
- location: RNA throughout the cell, DNA in nucleus (and mitochondria and chloroplasts, endosymbiotic theory)
- amount per cell: RNA varies, DNA constant
what are the roles of the different types of RNA (mRNA, tRNA, rRNA, snRNA, si/miRNA) MEMORISE
mRNA: carries information which codes for pp chains from DNA to ribosomes (only coding RNA)
tRNA: adaptor molecule in protein synthesis, translates mRNA codon sequence into aa sequence
rRNA: catalytic and structural roles in ribosomes
snRNA: catalytic and structural roles in spliceosomes
si/miRNA: regulation of gene expression
define transcription
the process by which a complementary RNA copy is made under the direction of the template strand of a specific region of the DNA molecule, catalysed by the enzyme RNA polymerase
define gene
a section of DNA that encodes information in the form of a specific base sequence to direct the synthesis of one polypeptide chain or RNA molecule
a unit of inheritance located at a locus on the chromosome, which specifies a particular character of an organism
state the three key components of a eukaryotic gene
promotor, coding region, termination sequence
what is the role of the promoter in a eukaryotic gene?
contains the TATA box and transcription state site (+1: the nucleotide where RNA synthesis begins
- TATA box serves as binding site for general transcription factor (TFIID), which facilitates binding of RNA polymerase
- TATA box typically located 25 base pairs upstream (closer to 3’ direction)
- promoter determines which DNA strand used as template, not transcribed except for transcription start site
what is the role of the coding region in a eukaryotic gene?
- segment of DNA transcribed into single-stranded RNA molecule (pre-mRNA)
- bounded by transcription start and termination site
- only 1 strand serves as transcription template, read in 3’ to 5’ direction to facilitate RNA synthesis in 5’ to 3’ direction
- RNA synthesis occurs within transcription bubble, DNA transiently separated into single strands
what is the role of the termination sequence in a eukaryotic gene?
codes for polyadenylation signal sequence (AAUAAA) in the pre-mRNA
whole termination sequence is transcribed
- transcription termination (no more phosphodiester bonds formed)
describe the structure of RNA polymerase during transcription
RNA polymerase is an enzyme comprising of several protein subunits, found in the nucleoplasm
- responsible for RNA synthesis using ribonucleoside triphosphate (RTP) as its substrate
what are general transcription factors, and what is its role in transcription?
protein required for RNA polymerase molecule to bind to promoter and initiate transcription
help position RNA polymerase correctly at promotor and release RNA polymerase from promoter
what happens during the initiation stage of transcription?
general transcription factors (TFIID) binds to TATA box within promoter,mediating the binding of RNA polymerase to the promoter and forming a transcription initiation complex
RNA polymerase binding causes DNA double helix to unwind and separate both strands of DNA, breaking hydrogen bonds between complementary base pairs and creating a transcription bubble
one of the strands acts as a template for complementary base pairing to direct the assembly of incoming ribonucleotides, and RNA polymerase catalyses the formation of the first phosphodiester bond (marking the end of transcription initiation)
what happens during the elongation stage of transcription?
RNA polymerase moves along template DNA in 3’ to 5’ direction (reading it), transiently unwinding DNA double helix
ribonucleotides form complementary base pairs with DNA template, phosphodiester bond formed between free 5’ phosphate group of incoming nucleotide and free 3’ hydroxyl (OH) group of growing pre-mRNA chain
mRNA synthesised in 5’ to 3’ direction
RNA polymerase reanneals unwound DNA, and proofreads it
what happens during the termination stage of transcription?
RNA polymerase transcribes a termination sequence (polyadenylation signal sequence, AAUAAA) in DNA, triggering release of RNA chain and dissociation of RNA polymerase from DNA
in eukaryotic cells, RNA polymerase continues transcription until a point about 10-35 nucleotides downstream of polyadenylation signal sequence
cleavage site of mRNA is also the site of addition of poly (A) tail
what post-transcriptional modifications does pre-mRNA have to undergo?
- addition of 5’ methylguanosine cap
- RNA splicing
- addition of 3’ poly (A) tail
how and why does pre-mRNA undergo the addition of a 5’-methylguanosine cap?
5’ end of new RNA has a methylated guanine nucleotide / methylguanosine triphosphate “cap” added
protects mRNA from enzymatic degradation
defines the 5’ end of mRNA, serves to recruit the small subunit of the ribosome from translation initiation
distinguishes mRNAs from other types of RNA molecules
how and why does pre-mRNA undergo RNA splicing?
both exons (protein-coding sequences) and introns (non-coding sequences) are transcribed into pre-mRNA
RNA splicing removes introns and splices / joins remaining exons together to form mature mRNA, via the hydrolysis of ATP for energy
splicing is carried out by spliceosomes, a large complex with several subunits known as small nuclear ribonucleoproteins (snRNPs), each with small nuclear RNAs and a set of proteins
how and why does pre-mRNA undergo the addition of a 3’ poly (A) tail?
3’ end of pre-mRNA modified by addition of a series of approximately 200 adenine (A) nucleotides, referred to as the poly (A) tail, catalysed by the enzyme poly (A) - polymerase
protects mRNA from degradation, more stable
required to facilitate export of mRNA out of nucleus via nuclear pores
what are the general features of the genetic code (how does it work)?
the set of rules by which the DNA base sequence of a gene, through the intermediary mRNA codon sequence, is translated into the amino acid sequence of a pp
a list of the codons of mRNA, the sequence of triplet bases in non-template / non-transcribed strand of DNA, each codon is written as it appears in mRNA and reads in 5’ to 3’ direction
4^3 = 64 possible codons, three nucleotides per codon and four possible bases for each nucleotide position
of 64 possible codons, 61 code for the twenty possible amino acids and include a start codon, and three termination signals / stop codons
what are the key features of the genetic code (TUND)?
triplet: three nucleotide bases for one aa
universal: used by (almost) all organisms
non-overlapping: read continuously in 5’ to 3’ direction as successive groups of 3 nucleotides, one codon at a time without skipping, 3 possible reading frames for every mRNA sequence
degenerate: a single aa can be coded by multiple codon, only methionine (AUG) and tryptophan (UGG) coded for by a single codon each
but one codon = one aa (unambiguous)
silent mutations: most aa encoded by degenerate codons that differ in the 3rd position, mutations in this position can not be expressed, leading to wobble base phenomenon (base-pairing at third bp is not so specific, since tRNA can still recognise even if it’s wrong)
what does the start codon and stop codons define?
start signal is AUG, codes for incorporation of methionine, defining the first aa of pp chain and reading frame used from that point on
stop signals (UAA, UAG, UGA) has no tRNA with anticodon complementary to it
define translation
the process in which a polypeptide chain is synthesised by ribosomes using genetic information encoded in a mature mRNA template
state all the components of translation machinery
mature messenger RNA
transfer RNA
amino acids
aminoacyl-tRNA synthetase
ribosomal RNA and ribosome
translation factors