Lecture 3: Transcription

Question 1

RNA predominantly exists in which form?

Answer 1

single strands, primary structure

Question 2

can RNA form secondary structures?

Answer 2

yes! they can fold back on themselves using complimentary base-pairing

Question 3

_____ is both a genetic and a functional molecule, how?

Answer 3

RNA
contains genetic information (mRNA) but also works as an enzyme (ribozymes) and structural elements (rRNA and tRNA)

Question 4

what are the three majopr types of RNA?

Answer 4

mRNA (1-2% cytosol)
tRNA (15% cytosol)
rRNA (85% cytosol)

Question 5

T/F: Structure of RNA affect longevity of molecules

Answer 5

true! mRNA only exists for a few mins because it almost only exists in primary structures
tRNA and rRNA exist longer because they have secondary structures that prevent ribonuclease attack!

Question 6

what are the roles of the three RNA molecules?

Answer 6

1.mRNA carries genetic information from the genome to the ribosome
2.rRNA has a functional and structural role in ribosomes (protein synthesis)
3.tRNA has an active role in carrying amino acids for protein synthesis

Question 7

transcription is catalyzed by…

Answer 7

RNA polymerase

Question 8

RNA polymerase Forms _______ between the ribonucleotides ATP, GTP, CTP, and
UTP

Answer 8

phosphodiester bonds

Question 9

_______ is driven by the energy released from the hydrolysis of two
energy-rich phosphate bonds of incoming ribonucleoside triphosphates

Answer 9

Polymerization

Question 10

RNA ________ involves the incorporation of ribonucleoside triphosphates
which are added to the 3’-OH of the ribose of the preceding nucleotide

Answer 10

chain elongation

Question 11

in what direction is RNA synthesis?

Answer 11

5’ to 3’ and antiparallel to the DNA template strand

Question 12

describe the pverview of transcription:

Answer 12

1. binding of RNA poly and sigma factor to promoter
2. transcription is initiated and sigma factor dissociates
3. elongation of mRNA
4. termination site is reached
5. disassociation of RNA poly and newly synthesized mRNA

Question 13

T/F: RNA polymerase uses DNA as a template, but only one of the two
strands is transcribed for any given gene

Answer 13

true!!

Question 14

T/F: genes are present on both strands of DNA, thus DNA sequences on
both strands can be transcribed, although at different locations

Answer 14

true!

Question 15

T/F: RNA polymerase can initiate synthesis on its own
* Does not need a primer to start from

Answer 15

true!

Question 16

which strand of DNA does RNA poly read?

Answer 16

template strand (3’-5’)

Question 17

which strand of DNA will have the exact same sequence as the mRNA strand? just with T instead of U

Answer 17

the coding strand (whichever DNA strand is 5’ to 3’)

Question 18

T/F: Transcript is
complementary to
template strand, and has
the same sequence
(except U’s in place of T’s)
as non-template strand

Answer 18

true!

Question 19

what are the two components to the transcription apparatus?

Answer 19

RNA polymerase
sigma factor

Question 20

______: Large multimeric enzyme (a.k.a.
core enzyme) → catalytic activity
* 2 α, β, β′, ω
* Is responsible for RNA synthesis
* The β subunit is the target site of
rifamycin class of antibiotics
thereby inhibiting RNA synthesis

Answer 20

RNA polymerase (the plane)

Question 21

which subunit of RNA poly is the target for rifamycin? (antibiotic)

Answer 21

the beta subunit

Question 22

______: Controls binding of RNA polymerase to promoter
* Detaches once the first few RNA nucleotides have been joined
together
* Many bacteria have multiple types

Answer 22

sigma factor (the pilot)

Question 23

RNA polymerase + sigma factor = ________

Answer 23

holoenzyme

Question 24

what are the three stages of transcription?

Answer 24

initiation, elongation, termination

Question 25

many bacteria have multiple types of sigma factors, how is this possible?

Answer 25

each type of sigma initiates the binding of RNA polymerase to a particular set of promoters

Question 26

how many sigma factors does E. Coli have?

Answer 26

sigma 70 and sigma 38, so two!

Question 27

______: Located upstream of gene, contains promoter elements essential for transcription

Answer 27

promoter region

Question 28

what two components does the promoter region consist of always, though the sequences may change?

Answer 28

-35 box and -10 box (Pribnow/TATA box)

Question 29

what is the consensus sequence for the -35 promoter box?

Answer 29

TTGACA

Question 30

what is the consensus sequence for the -10 promoter box?

Answer 30

TATAAT

Question 31

The -______ box is recognized by diverse sigma factors

Answer 31

35

Question 32

which sequence does the sigma 70 factor recognize? (consensus sequence and name)

Answer 32

-35 box, TTGACA sequence

Question 33

what are the five regions of a bacterial structural gene?

Answer 33

1. Promoter : recognition and binding site for RNA polymerase holoenzyme – Feature -35 and -10 boxes 2. Leader sequence: region transcribed into mRNA but not translated – Transcription begins at position +1 [usually A or G (purines)] 3. Coding region: transcribed and translated into polypeptide (i.e. protein) 4. Trailer: region transcribed into mRNA but not translated – Prepares RNA polymerase to release the template strand 5. Terminator: RNA polymerase releases the template strand

Question 34

_________ component of RNA ________ recognizes and binds to promoter region on the DNA coding strand (i.e. non-template strand)

Answer 34

Sigma factor holoenzyme

Question 35

what is the +1 site?

Answer 35

the location on template strand of the first nucleotide of RNA chain

Question 36

T/F: The -35 box sequence is sigma factor-specific

Answer 36

true!!

Question 37

Variations in sequence of ______ determine promoter strength (ease of RNA polymerase holoenzyme recognizing/binding promoter region)

Answer 37

-35 box i.e. the conserved/consensus sequence will bind the RNA poly best!

Question 38

Recognition and binding of RNA polymerase holoenzyme to promoter region forms a....

Answer 38

closed complex

Question 39

how does the open complex form? during transcription initiation

Answer 39

melting for -10 box region

Question 40

the open complex forms the....

Answer 40

transcription bubble, Region of 16-20 bp is unwound within complex Causes the Polymerization of the first few RNA nucleotides

Question 41

whats the only thing we need for the actual elongation of RNA transcript?

Answer 41

RNA polymerase core, sigma factor is released

Question 42

Core RNA poly enzyme reads _______ of DNA in 3’ to 5’ direction

Answer 42

template strand

Question 43

the elongation fo transcription involves a temporary...

Answer 43

dsRNA/RNA hybrid

Question 44

T/F: When conditions permit, transcription is a repetitive process; initiation is not dependent on completion of previous round of transcription

Answer 44

true! we can transcribe multiple mRNA at the same time!

Question 45

what will inverted repeats in the DNA sequence form in mRNA?

Answer 45

hairpin/stemloop structures, which can cause termination of transcription

Question 46

Hairpin structure presents as a physical obstruction to cause RNA polymerase to temporarily stall, what two mechanisms can then remove RNA poly from the template strand?

Answer 46

rho-independant termination rho-dependant termination

Question 47

is RNA poly recycled once removed from the template strand?

Answer 47

yes! can re-associate with sigma factor to form holoenzyme to once again initiate transcription

Question 48

describe how rho-independant termination occurs

Answer 48

A-rich region present after inverted repeats in DNA * String of “U”s (i.e. poly-U) in RNA following hairpin structure * Causes destabilization of RNA/DNA hybrid in open complex Release of RNA polymerase from template

Question 49

rho-independent termination is an _______ terminator, there's no special protein needed- encoded right in the genome!

Answer 49

intrinsic

Question 50

why would it be important to have a long strand of AAAAAA forming a poly-U tail in mRNA when using rho-independent termination?

Answer 50

weker bonds of A-U make it a good place for dissociation

Question 51

describe how rho-dependent termination functions

Answer 51

Once transcribed, rut site will be bound by multi-Rho hexameric protein that assembles around the RNA transcript * Features helicase activity fueled by ATP hydrolysis * Moves in the 5’ to 3’ direction along RNA transcript Hairpin structure will cause RNA polymerase to slow down and pause * Rho protein catches up and knocks off RNA polymerase off the template strand by destabilizing the RNA/DNA hybrid region

Question 52

Termination requires a special protein → ______ and a rut site located within the mRNA transcript

Answer 52

rho (ρ) factor

Question 53

the rut site is active only in... but encoded in...

Answer 53

RNA DNA!

Question 54

in what direction does the rho protein move (displaying helicase activity)

Answer 54

causing termination, so moving 5' to 3' along mRNA strand

Question 55

genetic information is organized into ______. Segments of DNA that are transcribed into a single RNA molecule bounded by their initiation and termination sites

Answer 55

transcriptional units

Question 56

what three different things can transcriptional units be?

Answer 56

1. be RNA transcribed from a single gene; has its own promoter and terminator → monocistronic transcript 2. form from two or more genes (co-transcribed genes) → Polycistronic transcript 3. be gene(s) that encode non-translated RNAs * tRNAs, rRNAs, non-coding RNAs (most have regulatory role)

Question 57

T/F: genes (as well as operons) can be located on either strand

Answer 57

true

Question 58

An ______ contains two or more genes (co-transcribed genes) flanked by one promoter and one terminator

Answer 58

operon structure

Question 59

T/F: Polycistronic mRNA transcript contains multiple ribosome binding sites upstream of coding sequences to facilitate translation (i.e. protein synthesis)

Answer 59

true!!

Question 60

Polycistronic RNA molecule that is not translated, but processed to form individual RNA molecules... what's an example of this?

Answer 60

rRNA operon (seven rRNA operons in E Coli!)

Question 61

describe the structure of the rRNA operon in E Coli, containing all three rRNA genes

Answer 61

promoter, gene encoding 16S rRNA, gene encoding tRNA, gene encoding 23S rRNA, gene encoding 5S rRNA, transcription terminator plus spacers always!

Question 62

Primary transcript (pre-RNA) is cleaved by _________ allowing spacers to be discarded

Answer 62

ribonucleases

Question 63

mature RNA molecules like rRNA and tRNA are ready to aid in which process?

Answer 63

translation!