ESSENTIAL CELL BIOLOGY
From DNA to Protein: How Cells Read the Genome
• DNA works as a manager delegating tasks to a team of workers
• Gene: a segment of DNA
• RNA copies of DNA are used to make proteins (Central dogma of molecular
biology)
• Transcription: Mechanism by which cells copy DNA into RNA
• Translation: RNA using information to synthesize a protein
• RNA splicing: a process in eukaryotic cells in which segments of an RNA
transcript are removed and the remaining segments stitched back together
before the RNA is translated into a protein
From DNA to RNA
• Production of RNA is the first step of gene expression
• Cells can change/regulate the expression of each of its genes according to its
present needs
1) Portions of DNA Sequence Are Transcribed into RNA
a) RNA: linear polymer made of four different nucleotide subunits linked by
phosphodiester bonds
i) Contains ribonucleotides
ii) Contains uracil instead of thymine
iii) Single-stranded
(1) Can fold into various shapes as a result (much like polypeptides)
(a) Allows for structural, regulatory, or catalytic roles
2) Transcription Produces RNA That Is Complementary to One Strand of DNA
a) RNA polymerase: enzyme that covalently links ribonucleotides to the
growing RNA chain
i) Catalyzes the formation of phosphodiester bonds
ii) Unwinds the DNA helix just ahead to expose a new region of the strand
for copying
iii) Moves in the 5’-to-3’ direction
iv) Powered by ribonucleoside triphosphates (ATP, CTP, UTP, and GTP)
v) Can start an RNA chain without a primer (unlike DNA polymerase)
vi) Makes about 1 mistake per 10 nucleotides (DNA polymerase makes 1 in
4
10 )
7
(1) Not as crucial as RNA is not a permanent storage of genetic
information
b) RNA transcript: the RNA chain produced by transcription
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i) Elongated according to one strand of DNA. The two are
complementary much like DNA replication
c) RNAs are copied from a limited region of DNA. They are much shorter than
DNA
d) RNA does not remain hydrogen-bonded to the DNA template strand. The
RNA chain is displaced so the DNA helix can reform
e) Many RNA copies of the same gene can be made in a short period of time
3) Cells Produce Various Types of RNA
a) Messenger RNAs: carriers of encoded genes from DNA
b) mRNAs code for a single protein in eukaryotes and a few genes in bacteria
c) RNAs can be the final product of some genes as they can serve roles too
i) Ribosomal RNAs: form the structural and catalytic core of the ribosomes
(1) Translates mRNAs into proteins
ii) Transfer RNAs: act as adaptors that select specific amino acids and
hold them in place on a ribosome for their incorporation into protein
iii) MicroRNAs: serve as key regulators of eukaryotic gene expression
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d) Gene expression: the process by which the information encoded in a DNA
sequence is translated into a product that has some effect on a cell or
organism
4) Signals in DNA Tell RNA Polymerase Where to Start and Finish Transcription
a) Transcription start site is recognized differently in bacteria and eukaryotes
i) For bacteria
(1) RNA collides randomly with DNA and binds weakly. Slides along
the length in search of a gene region called a promoter, where it will
bind tightly. Once bound, the RNA opens up the helix and transcribes
until the terminator (stop site) is reached.
(a) Sigma factor: helpers
(2) The strand used for the template is determined by the structure
of the promoter. Promotors are polar and ensures that RNA only
binds in one orientation.
(a) RNA can only synthesize in the 5’-to-3’ direction so the 3’-to-5’
direction is used as the template
5) Initiation of Eukaryotic Gene Transcription is a Complex Process
a) Eukaryotic cells have RNA polymerase I, RNA polymerase II, and RNA
polymerase III
i) I and III transcribe the genes encoding transfer RNA (III), ribosomal
RNA (I), and various other RNAs (III) that play structural and
catalytic roles
ii) II transcribes the vast majority of eukaryotic genes
(1) Focus will be in RNA polymerase II
b) Requires the help of a large set of accessory proteins
i) General transcription factors must assemble at each promotor along
with the polymerase before transcription
c) Mechanisms for initiation are more complex. Single gene is controlled
by a large variety of regulatory DNA sequences
i) Take into account the packing of DNA into nucleosomes and compact
chromatins
6) Eukaryotic RNA Polymerase Requires General Transcription Factors
a) General Transcription Factors: proteins that allow RNA polymerase to begin
transcription
b) TATA box: promotor regions in eukaryotic cells
c) Transcription initiation complex: TFIID and other factors plus RNA
polymerase II
d) RNA polymerase II is phosphorylated before transcription and
dephosphorylated after by enzymes. Only dephosphorylated RNA
polymerase II can initiate RNA synthesis
7) Eukaryotic mRNAs Are Processed in the Nucleus
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a) Bacteria DNA lies in the cytoplasm. Ribosomes immediately attach
to the free 5’ end of synthesized RNA to begin translation
b) Transcription takes place within the nucleus of eukaryotes and
translation takes place on the ribosomes in the cytoplasm
i) mRNAs have to be transported out through small pores in the nuclear
envelope
c) RNA processing
i) Capping
(1) Modifies 5’ end of the RNA transcript
(2) Adds a guanidine and occurs way before transcription finishes
ii) Splicing
iii) Polyadenylation
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(1) Adds a series of repeated adenine nucleotides to the 3’ end (poly-
A-tail) to a newly transcribed mRNA
d) Capping and polyadenylation is used to mark the mRNA and increase
stability
8) In Eukaryotes, Protein-Coding Genes Are Interrupted by Noncoding Sequences
Called Introns
a) Introns: intervening sequences that are long and noncoding within eukaryotic
genes
b) Exons: expressed sequences of genes. Usually shorter than introns
c) Introns and exons apply to both DNA and RNA
9) Introns Are Removed from Pre-mRNAs by RNA Splicing
a) RNA splicing stitches exons together
b) Introns contain a few short nucleotide sequences that act as cues for its
removal
c) RNA splicing is done by snRNPs (small nuclear RNAs and small nuclear
ribonucleoproteins).
i) Forms the core of spliceosome
d) Benefits of Intron-exon
structures:
i) Alternative splicing: transcripts spliced in different ways can produce
different proteins
ii) Intron-exon structures of genes are thought to have sped up the
emergence of new and useful proteins
10) Mature Eukaryotic mRNAs Are Exported from the Nucleus
a) Spliced introns cannot leave the nucleus (dangerous for cell)
i) Introns are degraded within the nucleus and their nucleotides recycled
b) Transport of RNAs out of nucleus is selective: only processed mRNAs are
exported
c) Nuclear pore complexes: acts as gates between nucleus and cytoplasm
d) A set of bound proteins determines whether an mRNA molecule will leave
the nucleus
11) mRNA Molecules Are Eventually Degraded in the Cytosol
a) The lifetime of an mRNA determines the number of proteins it produces
i) Can be used repeatedly
b) Eventually degraded into nucleotides by ribonucleases present in the cytosol
c) Bacteria mRNAs last around 3 minutes (mostly), eukaryotic mRNAs can
last from 30 minutes to 10 hours
12) The Earliest Cells May Have Had Introns in Their Genes
a) Costs of RNA splicing: cell has to maintain a larger genome and has to
discard a large fraction of RNA it synthesizes without ever using it
b) Cells may have shed introns to reproduce more rapidly and efficiently
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i) Evolution of prokaryotes
c) Introns may be the result of an originally parasitic mobile genetic
elements that invaded early eukaryotic ancestors
From RNA to Proteins
1) An mRNA Sequence Is Decoded in Sets of Three Nucleotides
a) Genetic code: the rules by which the nucleotide sequences of a gene
(through mRNA) is translated into the amino acid sequence of a
protein
i) Each group of three consecutive nucleotides in RNA is called a codon
(1) Each codon specifies one amino acid
(2) Some are redundant
b) Reading frames: three different codons depending on where the decoding
process begins
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i) Only one will code for the correct protein
2) tRNA Molecules Match Amino Acids to Codons in mRNA
a) tRNA (transfer RNAs): recognize and bind to a codon at one side and to an
amino acid at another
b) RNAs long enough will fold back on themselves to form a double-
helical structure like DNA: tRNAs are an example
c) Anticodon: a set of three consecutive nucleotides that bind complementary
codon in an mRNA molecule through base pairing
d) Amino acid that matches the codon is bound at the 3’ end of the tRNA
e) Redundancy in codons: there is more than one tRNA for many of the
amino acids or that some tRNA molecules can base-pair with more than
one codon
f) Wobble base-pairing: can tolerate a mismatched codon at the third position
i) Many codons differ only in the third nucleotide
ii) Makes it possible to have fewer kinds of tRNAs
3) Specific Enzymes Couple tRNAs to the Correct Amino Acid
a) Aminoactyl-Trna Synthetases: enzyme that recognizes the correct amino acid
for a tRNA
i) Equal in importance with tRNAs
b) Attachment of amino acids to tRNAs are coupled with ATP hydrolysis
i) Energy is stored to be used to link amino acids covalently to the growing
polypeptide chain
4) The mRNA Message is Decoded by Ribosomes
a) Ribosomes: large complex made from dozens of small proteins and several
RNA molecules called ribosomal RNAs (rRNAs)
i) Site of translation
b) mRNA is pulled through the ribosome as the ribosome translates its
nucleotide sequence into an amino acid sequence using tRNA as adaptors
c) Ribosomes have an mRNA binding site and three tRNA binding sites
i) A site
(1) Charged tRNA enters the A site by base-pairing with the
complementary codon in the mRNA
ii) P site
(1) Amino acid is linked to the peptide chain held by the tRNA
iii) E site
(1) tRNA is ejected
5) The Ribosome is a
Ribozyme
a) rRNAs are responsible for ribosome structure and ability to choreograph
and catalyze protein synthesis
b) Ribosomal proteins are to help fold and stabilize the RNA core
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c) Ribozymes: RNA molecules that possess catalytic activity
i) RNAs may have served as the first catalysts for living cells
6) Specific Codons in mRNA Signal the Ribosome Where to Start and to Stop
Protein Synthesis
a) Translation begins with the codon ‘AUG’
i) Very important as a mistake of one nucleotide can cause a dysfunctional
protein
b) Initiator tRNA: required the initiate translation. Only one capable of binding
to the P site without a large ribosomal subunit
i) Carries the amino acid methionine
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ii) Binds to the 5’ end of the mRNA
iii) Newly formed proteins all have methionine at the N-terminal (the
end of the protein synthesized first)
iv) Methionine is usually removed by a specific protease
c) Translation initiation factors: initiator tRNA and additional proteins
d) Prokaryotes use specific sequences to locate where to begin translation
i) mRNAs can encode for several different proteins unlike eukaryotic
mRNAs
e) Stop codon: signals the end of translation (UAA, UAG, UGA)
i) Release factors: proteins that stop translation by binding to the A site
f) Chaperone proteins: helps fold polypeptides correctly
i) Usually present as polypeptides emerge from the ribosomes
7) Proteins Are Made of Polyribosomes
a) Polyribosomes: binds to each mRNA molecule being translated
i) Many ribosomes working on a single mRNA allows for many more
proteins to be synthesized
ii) Polysomes: short for polyribosomes
iii) Present in both eukaryotes and prokaryotes
(1) Speeds up even more for prokaryotes as mRNA does not need to be
processed
8) Inhibitors of Prokaryotic Protein Synthesis Are Used as Antibiotics
a) Many effective antibiotics are compounds that act by inhibiting bacterial,
but not eukaryotic, RNA and protein synthesis
9) Controlled Protein Breakdown Helps Regulate the Amount of Each Protein in a
Cell
a) Protein population: rate of synthesis and lifespan
i) Proteins can last from years (structural) to seconds (metabolic)
b) Proteolysis: cells possess specialized pathways that enzymatically break
proteins down into their constituent amino acids
i) Proteases: enzymes that degrade proteins
(1) Degrade proteins that aren’t meant to last and those that are damaged
or misfolded
c) Proteasomes: large protein machines for protein breakdown in eukaryotic
cells
i) Contains proteases to prevent them from running rampant in the cell
ii) Acts on proteins that have been marked for destruction through the
attachment of ubiquitin (a protein)
10) There Are Many Steps Between DNA and Protein
a) Post-translational modifications: proteins that require further attention
before they are useful to the cell once they leave the ribosome
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RNA and the Origins of Life
1) Life Requires Autocatalysis
a) RNA world: RNA may have existed before DNA and proteins (can both
store genetic information and catalyze chemical reactions in primitive
cells)
i) Molecular fossils of an earlier world
b) The ability to catalyze reactions has led to the production of more molecules
like themselves
c) RNA molecules can in principle, catalyze their own synthesis
2) RNA Can Both Store Information and Catalyze Chemical Reactions
a) RNA is thought to have played a central role in the origin of life
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b) RNA molecules with their unique folded shapes can serve as catalysts
i) Not as diverse as proteins
3) RNA is Thought to Predate DNA in Evolution
a) Earliest cells may have genetic information stored in RNA instead of DNA
b) Ribose readily forms in the conditions of primitive Earth
i) Deoxyribose is not
c) DNA evolved as it has a more stable backbone as a result of its sugar-
phosphate group and thymine
d) DNA is easier to repair and detect damage
e) It is believed that many of the functions originally performed by RNA
were taken over by DNA and proteins
i) RNA remains the primarily as the intermediary connecting DNA and
proteins
f) Proteins are more diverse than RNA: allows for diversity of structure and
function we see in life today
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