Transciption Flashcards
In prokaryotes, the coding region of a gene is often a
Single, continuous unit (CO-LINEAR)
Only the untranscribed part is called the
Shine-Dalgarno sequence
Do transcription and translation occur at the same time in prokaryotes
yes :)
What is a protein coding segment
Exon
What segments do not code for proteins
Introns
Do transcription and translation occur at the same time in eukaryotes
No :(
Types of RNA
Messenger RNAs (mRNA) - intermediates that carry genetic info from DNA to ribosomes Transfer RNAs (tRNA) - adaptors between AA and codons in mRNA Ribosomal RNAs (rRNA) - structural and catalytic components of ribosomes Small nuclear RNA (snRNA & snoRNA) - spliceosomes, rRNA, tRNA modification, respectively MicroRNAs (miRNAs, siRNA & Crispr RNA) - short RNAs that block expression of complementary mRNAs Long noncoding RNAs - long RNAs that regulate gene transcription
What is the RNA synth to the DNA template strand
Complementary and antiparallel
RNA synthesized in _’ to _’ direction while DNA is the opposite
5,3
Transcription requires a _’ to _’ template and 4 ribonucleoside triphosphates and a DNA-dependent RNA polymerase.
3,5, rNTPs
Initiation in Prokaryotes cells
RNA polymerase binds to promoter, unwinds it and joins the first 2 nucleotides
Initiation of RNA synthesis DOES NOT require a primer
Recognition of the gene promoter region requires the intact RNA polymerase holoenzyme
σ factor recognizes and binds to the -35 element in promoter region properly position the RNA polymerase to begin transcription
2 important promoter sequence elements are the -35 element to which σ factor binds and -10 element which is prone to unwinding due to A/T rich content (fewer H bonds)
-35 sequence: 5’ TTGACA 3’, separated from -10 by ~16-19 bp
-10 sequence: 5’ TATAAT 3’ (aka “Pribnow” box)
Transcription initiates about 5-9 bp downstream from the end of the -10 sequence by looking for a purine (A or G) on the 5’ to 3’ strand, finding opposing pyrimidine (3’ to 5’ strand - template), and attaching the first purine onto template strand (synthesize in 5’ to 3’ direction)
Elongation in prokaryote cells
complementary nucleotides continue to be added during elongation process
Elongation occurs when σ factor is released and RNA polymerase begins to move along 3’ to 5’ DNA template strand
Localized DNA unwinding ahead of RNA polymerase generates “transcription bubble” (18 bp)
Transcription bubble moves with the RNA polymerase and the unwound DNA rewinds behind it
Positive supercoils formed in dsDNA ahead of advancing RNA polymerase are removed by topoisomerases (gyrase)
RNA polymerase has both helix UNWINDING & REWINDING activities
Termination in prokaryote cells
complementary nucleotides continue to be added during elongation process
Elongation occurs when σ factor is released and RNA polymerase begins to move along 3’ to 5’ DNA template strand
Localized DNA unwinding ahead of RNA polymerase generates “transcription bubble” (18 bp)
Transcription bubble moves with the RNA polymerase and the unwound DNA rewinds behind it
Positive supercoils formed in dsDNA ahead of advancing RNA polymerase are removed by topoisomerases (gyrase)
RNA polymerase has both helix UNWINDING & REWINDING activities
Transcription in eukaryotes, Five RNA polymerases
Refer to the notes for an answer its a big chart.
In eukaryotes, ____ recognize each of these specific types of promoters (through interaction with their DNA sequences) and ____ the appropriate polymerase for transcription
accessory proteins, bind/recruit
Initiation in eukaryotic cells
(for pol. II) - involves stepwise assembly of general Transcription Factors of pol. II (TFII A, B, D, E, F, and H)
TFIID assembles first at the TATA box followed by remaining general transcription factors (TFs) and finally, RNA polymerase II
This forms the preinitiation complex (PIC) which initiates transcription
Elongation in eukaryotic cells
many general transcription factors remain at promoter region, making for quick re-initiation with a new Pol. II
An ~8 bp transcription bubble is generated by RNA:DNA binding
This with DNA unwinding ensures that free RNA 3’-OH terminus is free for new rNTP addition
Termination in eukaryotic cells
involves cleavage of pre-mRNA and 5’ to 3’ degradation of remaining RNA by Rat1 exonuclease
RNA po. II transcribes well past the coding sequence of the gene
Transcription terminates when Rat1 catches up to pol. II (follows up behind it degrading excess mRNA as it goes)
Transcript is polyadenylated (adds a bunch of A bases on 3’ end)
An important inhibitor of transcription is α-amanitin (found in Amanita mushrooms) - inhibits RNA pol. II at both initiation & elongation states of transcription (genes are not expressed)
The removal of introns from pre-mRNA are required to form
mature mRNA that is translated into a polypeptide
Removal of introns
- Addition of 7-Methyl Guanosine (7-MG) Cap - unique linkage to pre-mRNA by 5’ phosphate of 7-MG and 5’ phosphate of first ribonucleotide in the RNA (5’ to 5’ phosphate linkage)
Addition of 7-MG cap occurs early in elongation process - Addition of PolyA tail - Eukaryotic pre-mRNA is cleaved 11-30 nt following 5’ AAUAAA 3’ sequence
PolyA polymerase adds ~200 A residues at cleaved end - Removal of Introns - introns in pre-mRNA removed by specialized process called “RNA splicing”
Removal of introns must be precise in order to properly fuse 3’ end of one exon to 5’ end of next
Every intron has 2 conserved sequences that are required for its precise removal:
5’ & 3’ splice sequences containing junction sequences “5’ GU & 3’ AG”, respectively
Intron Branch point - conserved “A” residue
Splicing is accomplished by RNA/protein complex known as the spliceosome
Spliceosomes contain five snRNAs - U1, U2, U4, U5, and U6
snRNAs associate with about 40 small proteins to form small nuclear ribonucleoproteins (snRNPs, pronounced “snurps” lol)
snRNPs U1, U2, U4, U5, and U6 assemble to form a complete spliceosome
snRNP U1 binds to 5’ splice site, snRNP U2 binds to branch site
Complete spliceosome assembles and cleaves transcript at 5’ splice site
5’ end of intron joined to A in branch site to form a lariat structure, U1 & U4 are released
Lariat structure - linkage between 5’ phosphate of G and 2’ OH of A (fused loop)
3’ splice site is cleaved and 5’ end of exon 2 is joined to 3’ end of exon 1, lariat-shaped intron is released along with U2, U5, and U6
Summary - snRNP assembly → 5’ splice site cleaved → lariat loop forms → 3’ splice site cleaved → exons joined and intron removed
Alternate splicing
Only one type of pre-mRNA made, can result in exons being removed along with introns, different mature mRNAS
Multiple 3’ cleavage sites
One exon may have more than one 3’ cleavage cute which results in 2 different chunks of that exon
Rna editing is carried out by
guide RNAs (gRNA)
Cytidine deaminase converts _ to _ and in one specific case transforms a glutamine codon (CAA) into a ____ codon (UAA)
C,U,Termination
Transfer RNAs
Transfer RNAs (tRNAs) - specialized RNAs that have a role as adaptor molecules in translation First postulated by Crick in 1956 as adaptors between AA and codons in mRNAs At 3’ end of tRNA there is a tag: 5’ CCA 3’ that binds to AA covalently Specificity of tRNA to AA is dictated by the anticodon tRNAs have modified bases in some parts that assist with structure & function
Ribosomal RNAs
rRNAs) - key component of ribosome
Ribosomes composed of large and small subunits that are assembled from many different proteins and rRNAs
Prokaryotes - 5S and 23S rRNA in 50S (large) subunit of prokaryotic ribosomes, 16S rRNA in 30S (small) subunit (ribosome is 70S, densities are NOT additive)
16S rRNA has a sequence complementary to shine-dalgarno sequence that is found at the 5’ end of prokaryotic mRNA
Small subunit is essential for initiation of translation in prokaryotes
rRNA synthesis and ribosome assembly occur in the cytoplasm
tRNA is encoded in rRNA gene and trimmed out in rRNA processing
Eukaryotes - 5S, 5.8S, and 28S rRNA associate with 60S (large) subunit, 18S rRNA in 40S (small) subunit (ribosome is 80S)
rRNAs are synthesized and ribosomes are assembled in nucleolus (eukaryotic)
rRNAs are processed through methylation, cleavage, and trimming before being put to use
Small nuclear RNAs (snRNAs)
snRNAs act in complexes with proteins, play roles in post-transcriptional processing of RNA such as splicing (spliceosome assembly)
Small nucleolar RNAs (snoRNAs)
also act in complexes with proteins, involved in chemical modifications of rRNAs, tRNAs, and snRNAs
Small micro RNAs
(siRNA, miRNA, Crispr RNA) - act differently in eukaryotes than in prokaryotes
Eukaryotes - act as short (~22nt) ssRNAs that bind to complementary sequences in mRNA
Produced by cleavage of mRNAs, RNA transposons, and RNA viruses
Regulate and control gene expression in different ways
siRNA bind to mRNA and inhibit translation, miRNA bind to mRNA and degrade it
Prokaryotes - Crispr RNA is encoded by DNA in prokaryotes, works in association with Cas9 nuclease to cleave foreign DNA that enters host cell
Bacterial defense system that prevents incorporation of foreign DNA into host genome
Long noncoding RNA
Comprise about 80% of mammalian genome, regulate and control gene expression
