lect 16: DNA to protein I Flashcards
what is gene expression?
DNA codes for product (RNA or protein) to be used by cells
-doesn’t always refer to both transcription and translation
what are the learning objectives of this lecture?
-compare the process of transcription in prokaryotic and eukaryotic cells
-describe the synthesis and structure of eukaryotic mRNA
what are the differences between RNA and DNA?
differ at carbon 2
what is complex about RNA?
RNA folding=complex 3D shapes
RNAs have specific folding
-diverse functions depending on the folding (regulation, catalyzing)
-driving force=complementary base pairing
-forms double stranded stems and connected by loops (stem and loop structure)
-non-standard base pairing modified bases (like methylation)
what are the types of RNA?
what is RNA polymerase?
in both prokaryotes and eukaryotes
-DNA-dependent (needs DNA as template)
-does not require primer (can bind and synthesize)
-synthesizes complementary RNA strand (20-50 nucleotides/second, fast but not as fast as DNA polymerase)
-reads in 3’ to 5’ direction (same as DNA polymerase), synthesizes in 5’ to 3’
-gene transcription tightly regulated
-proof reading function
-multiple transcripts from one gene
-processive: once it binds, it will not dissociate but continue to end
what is the directionality of RNA synthesis?
RNA synthesized 5’ to 3’
-i.e. nucleotides added to 3’ end
RNA polymerase reads DNA 3’ to 5’
what are the stages of transcription?
- initiation (what to know where to bind, promoter region)
-RNA polymerase binds to promoter region of DNA sequence, DNA unwound, RNA synthesis begins - elongation (RNA polymerase continues along and adds nucleotides)
-RNA polymerase moves along DNA synthesizing RNA - termination
-RNA polymerase encounters transcription stop signal in DNA (have stop sequences so RNA polymerase dissociates)
what are promoters?
where transcription of gene starts
-promoter is before (upstream) of transcribed gene
-each gene (group) has own promoters
-specific DNA sequence allows RNA polymerase binding
downstream=transcribed region
+1=transcription start site unit (to RNA) is downstream
-because strands are antiparallel, we need to chose which one is template and coding strand (depends on orientation of promoters)
-promoters read in specific direction by RNA polymerase which tells it which is template strand
why is the promoter _______
asymmetrical
how can different genes be transcribed from different DNA strands?
gene “a” and gene “b” transcribed from different DNA strands
-depends on orientation of promoter region (chooses which strand is template strand)
-can have opposite so both strands can be used as templates for different genes
-RNA polymerase reads 3’ to 5’, synthesizes in 5’ to 3’ direction
what are the differences in transcription in prokaryotic and eukaryotic cells?
-cytoplasm vs nucleus
-prokaryotes: simultaneous transcription/translation (both occur in cytoplasm)
-eukaryotes need pre-mRNA to do translation
-pros have 5 subunits and euks have 10-17 subunits in RNA polymerase
-euks: multiple proteins to recognize promoter (general transcription factors)
where does transcription and translation occur in prokaryotic cells?
remember:
-no nuclei
-no physical separation of DNA, RNA, ribosomes (why it is simultaneous transcription and translation in the cytoplasm)
-mRNA translated while transcription in progress
operons
-cluster of genes transcribed together into single mRNA molecules (in euks, different promoters for different genes)
what are the typical promoters in bacteria?
how many RNA polymerase does bacteria have?
what is prokaryotic transcription?
RNA polymerase recognize -35 and -10 promoters
conformational change in RNA polymerase after 10 nucleotides
-sigma factor dissociates (only need to bind at beginning)
-becomes a transcriptional elongation complex
transcription stops when terminator sequence reached
-completed RNA released
-map require p factor
two different factors at beginning and end of transcription
-RNA polymerase unwinds DNA in prokaryotes (helicase unwinds DNA in DNA replication)
what are the RNA polymerases used in the initiation of eukaryotic gene transcription?
three types of RNA polymerase
-RNA polymerase I: most rRNA
-RNA polymerase II: mRNA
-RNA polymerase III: tRNA
-requires large set of accessory proteins (very complex)
-mechanisms that control transcription initiation are elaborate
-initiation must deal with higher-order chromatin structure (euk chromosomes have more complex structure)
what is the overview of stages of transcription?
- initiation
-RNA polymerase binds to promoter region of DNA sequence, unwinds DNA, begins RNA synthesis - elongation
-RNA polymerase moves along DNA synthesizing RNA - termination
-RNA polymerase encounters transcription stop signal in DNA
what is the elongation step in transcription of eukaryotes?
what is the closer look at RNA polymerase in eukaryotes?
RNA polymerase is multi-subunit complex
-“groove” for DNA
RNA transcript exits via channel (yellow)
-complementary NTP
-Mg+ ion: neutralizes phosphate
-RNAPII is processive
what is the machinery for mRNA transcription i eukaryotes?
what is the most known promoter region in eukaryotes?
critical portion located 24-32 bases upstream of initiation site
-contains TATA box
what are some other known promoter elements and what are their general transcription factors in eukaryotes?
eukaryotes have multiple promoters and are recognized by transcription factors
what is the graph of the initiation of transcription from eukaryotic RNA polymerase II promoter?
