Genetics comprehensive exam summary

Ch. 12 - Transcription
a. Requirements
i. ssDNA template
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ii. Substrates to make RNA (ribonucleoside triphosphates)
iii. Transcription machinery
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1. RNA polymerase
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a. Core enzyme
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b. Sigma factor
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b. Directionality
i. Only synthesize RNA in the 5’ to 3’ direction
ii. Different direction of transcripts → gene specific
c. Template vs. Nontemplate
d. Transcriptional Unit 
i. Stretch of DNA that codes for an RNA molecule AND the sequences necessary
for transcription
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ii. Keys
1. Promoter:
a. DNA sequence that transcriptional apparatus recognizes
b. Indicates which strand of DNA will be transcribed
c. Determines start site
d. NOT transcribed but essential for transcription
2. RNA coding sequence
a. The template for transcription
3. Terminator
a. IS transcribed
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iii.
e. Polymerase
i. Bacterial RNA polymerase
1. Usually only one type
2. Synthesizes all classes of RNAs (m, t, and r)
3. 4 key subunits to Core enzyme
a. 2x alpha (α)
b. 1 beta (β)
c. 1 beta prime (β’)
4. Sigma factor
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a. Added to core subunits

, b. Holoenzyme]]]]]
i. Initiate transcription at promoter
c. Different sigma factors direct initiation at different promoters
5. Have to have the sigma factor to be complete
6. Can be multiple sigma factors but only one polymerase




7.
ii. Eukaryotic RNA polymerases
1. Result of large multiprotein complexes
2. Polymerase II most common
3.
f. Steps (Bacterial)
i. Initiation
1. Machinery assembles on promoter, begins synthesis of RNA
2. Polymerase does NOT need a primer
3. No functional promoter = no transcribe = no protein product
4. Steps:
a. Promoter recognition/binding
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i. Key to determining frequency that a gene is transcribed
(sigma factor)
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ii. Important consensus sequences
1. Are NOT transcribed
2. Binds to holoenzyme
3. -10 consensus sequence = Pribnow Box (TATA
Box)
a. Site where unwinding begins
b. Only 2 H bonds between As and Ts → less
energy and easier to break
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4. -35 consensus sequence
b. Formation of transcription bubble
c. Generation of first bonds between ribonucleotides
d. Escape of machinery from the promoter
i. Sigma factor keeps it there because there is a promoter
ii. Once it escapes it kicks the sigma factor out and the core
enzyme continues to transcribe the RNA 
ii. Elongation

, 1. RNA polymerase reads DNA, adds ribonucleotide to growing RNA
2. Steps:
a. RNA polymerase unwinds DNA ahead of transcription bubble and
rewinds behind it
b. 5’ to 3’ extension of RNA
c. DNA/RNA duplex within bubble
d. RNA trails behind
e. Farther away on the strand = transcribed first
iii. Termination
1. End of transcription
2. Separation of RNA from DNA template
3. Rho-independent
a. Contain inverted sequences
i. Triggers termination
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ii. Allows RNA to fold over and match up
iii. Forms right behind polymerase
b. “Hair pin” (stem-loop) structure formation is required
i. This structure causes RNA polymerase to pause
ii. Also may destabilize RNA/DNA complex
c. String of As and Us come after (not as strong) → hold together the
RNA and DNA duplex
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i. When polymerase pauses, the As and Us fall apart and the
whole thing dissociates
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4. Rho-dependent
a. Requires protein (rho)
b. Requires DNA sequence that causes a pause in transcription
c. Stretch of DNA upstream of termination site that is devoid of  2o
structure
d. Starts the same but when polymerase pauses, it stays together
(no secondary structure yet)
e. Rho does not pause and binds to transcript so rho catches up →
“bowling ball” → breaks bonds, causes dissociate
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g. Eukaryotic Differences
i. Adds levels of complexity
ii. Euk cells have 3 RNA polymerases
iii. Promoter definition depends on type of polymerase
iv. Many proteins take part in binding of polymerase to DNA
1. Different proteins require different proteins
v. KEY: promoters are recognized by accessory proteins
1. Recruit polymerase
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2. Vs. bacterial system → holoenzyme recognizes promoter
vi. Transcription and Nucleosomes
1. DNA must be accessible to RNA polymerases