Gene Lecture 11 Transcription

11/17/2015
Flow of Genetic Information
Fundamental Genetics
Lecture 9
The Genetic Code

and Transcription
John Donnie A. Ramos, Ph.D.
Dept. of Biological Sciences
College of Science
University of Santo Tomas
The Genetic Code
The Discovery of the Genetic Code
Linear form (mRNA derived from DNA)
Francois Jacob and Jacques Monod

(1961) messenger RNA (mRNA)
Triplet codons (triplets of ribonucleotides coding for 1

amino acid)
Sydney Brenner (1960s) codon in

triplets (minimal use of the 4 mRNA
bases to specificy 20 aa) (43=64)
Unambiguous (1 codon = 1 amino acid only)

Degenerate ( 1 amino acid can be specified by several
codons)
Contains specific start and stop codons
Commaless (no breaks once translation starts until the
stop codon is reached)
Francis Crick frameshift mutations

alters the codons
Mariane Manago and Severo Ochoa polynucleotide phosphorylase
(synthesis of RNA without template)paved the way to the production of
RNA polymeres in cell free-systems
Non-overlapping (single reading frame)

Universal (same ribonucleotide used by all organisms)
The Discovery of the Genetic Code

Marshall Nirenberg and J. Heinrich Matthaei (1661) codons
used cell-free protein synthesizing system and polynucleotide
phosphorylase
RNA Homopolymers (UUUUUU, AAAAAAA, CCCCCC, GGGGG)
UUU (Phenylalanine)
AAA (Lysine)
CCC (Proline)
The Triplet Binding Assay

Developed by M. Nirenberg and P. Leder (1964)
Mimics the in vivo translation of proteins where a mRNA-tRNAribosome complex is formed when all three macromolecules are
allowed to interact.
RNA Mixed Copolymers
1A:5C (1/6 A: 5/6C)
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Repeating Copolymers
Results of Repeating Copolymers
Developed by Gobind Khorana (1960s)

Synthetic long RNAs with repeating sequences
The Universal Genetic Code
Exceptions to the Universal Code
Degeneracy
Wobble
Hypothesis
Start codon
(N-formylmethionine)
Termination
codons
Universal
Viruses
Bacteria
Archaea
Eukaryotes
Transcription
Uses DNA as a template
Catalyzed by RNA polymerase
(holoenzyme of 500 kD)
subunits
Sense strand / template strand

DNA strand used as a template
for transcription
Promoter region DNA sequence
recognized by factor to initiate
transcription (60 bases).
(upstream of a gene)
TATA box (Pribnow box)
TATAAT sequence
Sigma factor (70, 28, 32, 54)
Transcription
RNA polymerase dont need
primers
Elongation in 5 to 3 direction
Rate in E coli: 50 bases/sec at
37C
Termination is a function of rho
() factor hexameric protein
interacting with the end of a
gene
Polycistronic mRNA bacterial
mRNA containing information for
the synthesis of proteins of
related function
Monocystronic mRNA
eukaryotic mRNA containing
information for a single protein.
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Eukaryotic Transcription
Features of eukaryotic transcription different from prokaryotic
transcription:
Transcription inside the nucleus under the direction of 3 different
RNA polymerases
Cis -acting Elements

TATA Box (Goldberg-Hogness Box)
Located 30 bases upstream from the start of
transcription (-30)
Consensus sequence: TATAAAA
Facilitates denaturation of helix because it is ATrich region
CAAT Box
Presence of protein factors (promoters, enhancers, etc.) binding to
the upstream portion of a gene (cis-acting elements) during
initiation step.
Presence of post-transcriptional regulation.
Trans -acting Factors

Transcription factors facilitates template
binding during the initiation of transcription
Example:
TFIID (TATA-binding protein or TBP) binds to
TATA-box
Located 80 bases upstream from the start of

transcription (-80)
Consensus sequence: GGCCAATCT
Influence the efficiency of the promoter
Post-transcriptional Processing
7-methylguanosine cap (7mG)
Protection from nucleases
Role mRNA transport across the
nuclear membrane
3 cleavage site:
AAUAAA
Failure of 3 cleavage results to
absence of poly A tail
Split genes contains intervening

sequences
RNA Splicing
Ribozyme RNA with
catalytic activity
Self-excision process
process of RNA splicing or
intron removal.
Transesterification
interaction between
guanosine and the
transcript.
The Spliceosome
Alternative splicing
Small nuclear ribonucleoproteins
(snRNP or snurps) bonds to GU
or AG sites of introns
2 transesterification processes
Snurps form a loop (lariat) in the
branch point region
Produces isoforms of proteins
2 successive
transesterification processes
11/17/2015
RNA Editing
Substitution editing
changes in the nucleotide bases of a given mRNA
Common in mitochondrial RNA and chloroplast RNA
Example: Apoliprotein B (Apo B) C to U change CAA
to UAA
Insertion / deletion editing

addition or removal of nucleotide sequences
Common in mitochondrial RNA or guide RNA (gRNA)

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11/17/2015

Flow of Genetic Information

The Genetic Code

The Genetic Code

The Discovery of the Genetic Code

Linear form (mRNA derived from DNA)

Francois Jacob and Jacques Monod

Triplet codons (triplets of ribonucleotides coding for 1

Sydney Brenner (1960s) codon in

Unambiguous (1 codon = 1 amino acid only)

Francis Crick frameshift mutations

Non-overlapping (single reading frame)

The Discovery of the Genetic Code

The Triplet Binding Assay

RNA Mixed Copolymers

1A:5C (1/6 A: 5/6C)

Results of Repeating Copolymers

Developed by Gobind Khorana (1960s)

The Universal Genetic Code

Exceptions to the Universal Code

Sense strand / template strand

Cis -acting Elements

Trans -acting Factors

Located 80 bases upstream from the start of

Split genes contains intervening

Insertion / deletion editing

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