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Molecular Genetics > Lecture 4: RNA molecules and processing > Flashcards

Lecture 4: RNA molecules and processing Flashcards

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1
Q
  • What is a gene?
A
  1. A gene can be defined as a DNA sequence that encodes an RNA molecule

2 - inherited factor that determines a characteristic

3 - a set of DNA nucleotides

Today, a gene is often defined as ‘a DNA sequence that encodes an RNA molecule’.

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2
Q

Colinear meaning:

A

When a continuous sequence of nucleotides in DNA encodes a continuous sequence of amino acids in a
protein, the two are said to be colinear

In EUKARYOTES ,NOT ALL GENES are colinear with the
proteins that they encode.

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3
Q

EXONS AND INTRONS IN EUKARYOTIC GENES…

A

Many eukaryotic genes contain exons and introns.

Both are transcribed into RNA, but introns are later
removed by RNA processing.

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4
Q

In eukaryotic cells, genes are often interrupted by noncoding sequences (introns), which are removed during pre-mRNA processing

EXPLAIN HOW..

A
  1. pre-mRNA processing in
    eukaryotic genes:
    - the 5’ cap is added
    - the 3’ end is cleaved and poly(A) Tail is added
    - introns are removed within the spliceosome
  2. There are four major types of introns, differentiated by how the intron is removed from RNA
  3. Some introns found in rRNA genes and mitochondrial genes are self-splicing.
  4. mRNA has three primary
    regions: a 5’ UTR, a protein coding region, and a 3’ UTR.
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5
Q

In eukaryotic cells, genes are often interrupted by noncoding sequences (introns), which are removed during pre-mRNA processing: STEPS

A
  1. introns, exons and a long 3’ end are all transcribed into pre-mRNA.
  2. A 5’ cap is added
  3. Cleavage at the 3’ end is approximately 10 nucleotides downstream of the consensus sequence.
  4. Polyadenylation at the cleavage site produces the poly(A) tail.
  5. Finally, the introns are removed….
  6. …producing the mature mRNA.

Mature eukaryotic mRNA is produced when pre-mRNA is transcribed and undergoes several types of processing.

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6
Q

Alternative processing results in the production of different
proteins from the same DNA sequence

A

In both processes, the same pre-RNA can be processed in different ways.

A. Alternative splicing uses different combinations of exon.
- either 2 introns are removed to yield one mRNA.
- ..or 2 introns and exon 2 are removed to yield a different mRNA.

B. Multiple 3’ Cleavage sites.
Multiple 3’ cleavage sites use different locations for 3’ cleavage.

  • cleavage may be at 3’ site 1…
  • …or at 3’ site 2.
  • mRNA products pf different lengths are produced after splicing.

EUKARYOTIC CELLS HAVE ALTERNATIVE PATHWAYS FOR PROCESSING pre-mRNA.
A. with alternative splicing, pre-mRNA can be spliced in different ways to produce different mRNAs.

B. with multiple 3 cleavage sites, there are 2 or more potential sites for cleavage and polyadenylation; use of the different sites produces mRNAs of different lengths.

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7
Q

Alternative processing in the calcitonin gene…

A
  1. in thyroid cells, cleavage and polyadenylation take place at the end of exon 4.
  2. …producing an mRNA that contains exons 1,2,3, and 4.
  3. Translation produces the hormone calcitonin.
  4. in brain cells, 3’ cleavage takes place at the end of exon 6.
  5. during splicing, exon 4 is eliminated with the five introns…
  6. …producing an mRNA that contains exons 1,2,3,5 and 6.
  7. Translation yeilds calcitonin-gene-related peptide.

pre-mRNA encoded by the gene for calcitonin undergoes alternative processing.

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8
Q

Synthesis and processing of an mRNA are simultaneous
and highly interlinked…EXPLAIN 3

A

1 * CTD-P binds processing enzymes & factors for capping & splicing RNA

2 * 5’ cap is 7-methylG linked to transcript by 3 phosphates

3 * Cap protects RNA from degradation & is required for
translation

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9
Q

mRNAs may also be
altered by RNA editing…EXPLAIN

A

Individual nucleotides in mRNA may be changed, added, or deleted by RNA editing.

The amino acid sequence
produced by the edited mRNA is not the same as that encoded by DNA.

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10
Q

mRNAs may also be
altered by RNA editing…STEPS

A
  1. The unedited mRNA pairs with guide RNA.
  2. The guide RNA serves as a template for the addition, deletion or alteration of bases.
  3. The mature mRNA is then Released.
  4. Some of the uracil nucleotides (coloured green) were added by RNA editing.

RNA EDITING IS CARRIED OUT BY GUIDE RNAs.
The gRNA has sequences that are partly complementary cleavage and new nucleotides are added, with sequences in the gRNA serving as a template. the ends of the mRNA are then rejoined.

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11
Q

All tRNAs are similar in their secondary structure, a feature that is critical to tRNA function.

EXPLAIN

A
  • All tRNAs are similar in size and have a common secondary structure known as the cloverleaf.
  • Transfer RNAs contain modified bases and are extensively processed after
    transcription in both bacterial and eukaryotic cells.
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12
Q

All tRNAs possess a common secondary structure, the cloverleaf.

The base sequence in the flattened model is for tRNA^Ala

A
  1. This computer-generated space-filing molecular model shows the three-dimensional structure of a tRNA.
  2. Ribbon model emphasizes the internal regions of base pairing.
    - the anticodon comprises 3 bases and interacts with a codon in mRNA.
  3. Flattened cloverleaf model shows pairing between complementary nucleotides.
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13
Q

Ribosomal RNA is processed after transcription in
both bacterial and eukaryotic cells

A
  1. Methyl groups are added to specific bases and to the 2’Carbon aton of some ribose sugars.
  2. The RNA is cleaved into several intermediates…
  3. ..and trimmed
  4. Mature rRNA molecules are the result.
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14
Q

What is the genetic code?

A

The genetic code is a triplet code: three nucleotides
specify a single amino acid

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15
Q

Properties of GENETIC CODE: 5

A
  1. Solving the genetic code required several different approaches, including the use of synthetic mRNAs with random sequences.
  2. The genetic code is nonoverlapping, and universal (almost).
  3. The genetic code is degenerate (meaning that more than one
    codon may specify an amino acid).
  4. The reading frame is set by the initiation codon. The end of the protein-coding sequence of an mRNA is marked by one of three termination codons.
  5. Some tRNAs bind to multiple codons (“wobble”)
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16
Q

The genetic code consists of ….codons which encode
…. amino acids

EXPLAIN ITS COMPONENTS
- 4

A

The genetic code consists of 64 codons which encode
20 amino acids

1 * 3 nucleotides, each nucleotide can have one of four possible bases (A, G, C, or U), so 43 (64) possible codons.

2 * 20 amino acids

3 * Code is degenerate

4 * Many amino acids are
specified by more
than one codon

The gentic code consists of 64 codons. The AA specified by each codon are given in their 3 letter abbreviation. the codons are written 5’ to 3’, as they appear in the mRNA. AUG is an initiation (start) codon as well as the codon for methionine; UAA, UAG and UGA are the termination (stop) codons.

17
Q

The reading frame is set by the initiation (start)
codon: 4 explain

A

1 * Any sequence of nucleotides has three potential reading frames

2 * Each reading frame has different sets of codons and therefore
specifies different amino acid sequences

3 * It is essential for the translation machinery to use the correct
reading frame

4 * The reading frame is set by the initiation codon

18
Q

Some tRNAs bind to multiple codons (“wobble”)

A

Wobble may exits in the pairing of a codon and an anticodon.

The mRNA and tRNA pair in an antiparallel fashion. Pairing at the 1st and 2nd codon positions is in accord with the WATSON-AND-CRICK rules (A with U, G, with C); however, pairing rules are relaxed at the 3rd position of the codon, and G on the anticodon can pair with either U or C on the codon in this example.

19
Q

What is TRANSLATION?
steps?

A

Protein synthesis requires tRNA charging, initiation, elongation, and termination

  • In the process of translation, amino acids are linked together in the order specified by mRNA to create a polypeptide chain.
20
Q

Protein synthesis can be divided into four stages:

A

1 * tRNA charging (tRNAs bind to amino acids)

2 * Initiation (components necessary for translation assemble at
the ribosome)

3 * Elongation (amino acids are joined to the polypeptide chain)

4 * Termination (protein synthesis halts at the termination codon)

21
Q

A number of initiation, elongation, and release factors ..affected by?

A

A number of initiation, elongation, and release factors take part in the process, and energy is supplied by ATP and GTP.

22
Q

The first stage of translation is tRNA charging;

A

the binding of tRNA molecules to their appropriate amino acids

Certain positions on tRNA molecules are recognised by the appropriate aminoacyl-tRNA synthase

An amino acid attaches to the 3’ end of a tRNA.
The carboxyl group (COO-) of the amino acid attaches to the hydroxyl group (OH) of the 2’ or 3’ - carbon atom of the final nucleotide at the 3’ end of the tRNA, in which the base is always adenine.

23
Q

During initiation, the
components necessary
for protein synthesis
assemble

A
    • mRNA
    • Small and large subunits of the ribosome
    • 3 proteins called initiation factors
    • The initiator tRNA with its amino acid (MettRNA )
    • guanosine triphosphate (GTP).
  • small ribosomal subunit binds the
    mRNA at the 5’ end and scans for
    start codon
  • Met-tRNA binds the initiation codon,
    ribosome assembly & initiation factor
    dissociation
24
Q

Elongation; amino acids are joined to create a polypeptide chain

STEPS = 9

A
  1. fMet-tRNA^fMet occupies the P site of the ribosome.
  2. EF-Tu, GTP, and charged tRNA form a complex…
  3. …that enters the A site of the ribosome.

4…After the charges tRNA is placed into the A site, GTP is cleaved to the GDP, and the EF-Tu-GDP complex is released.

  1. EF-Ts regenerates the EF-Tu-GTP complex, which is then ready to combine with another charged tRNA.
  2. A peptide bond forms between the amino acids in the P and A sites, and the tRNA in the p site releases its amino acid.
  3. the ribosome moves down the mRNA to the next codon (translocation) which requires EF-G and GTP.
  4. The tRNA that was in the P site is now in the E site, from which it moves into the cytoplasm.
  5. the tRNA that occupied the A site is now in the P site. The A site is now open and ready to receive another tRNA.
25
Q

Termination; Protein
synthesis ends when the
ribosome translocates to a
termination codon

A
  1. When the ribosome translocates to a stop codon, there is no tRNA with an anticodon that can pair with the codon in the A site.
  2. RF-1 attaches to the A site,…
  3. …and RF-3 forms a complex with GTP nad binds to the ribosome.
  4. the polypeptide is released from the tRNA in the P site.
  5. GTP associated with the RF-3 is hydrolyzed to GDP.
  6. The tRNA, mRNA and release factors released from the ribosome.

TRANSLATION ENDS WHEN A STOP CODON IS ENCOUNTERED. BECAUSE UAG IS THE TERMINATION CODON- RF-1 IS THE RELEASE FACTOR.

26
Q

Study of the 3D structure of the ribosome using X-ray
crystallography helped us understand its function

A

Study of the 3D structure of the ribosome using X-ray
crystallography helped us understand its function

27
Q

Each mRNA molecule may be simultaneously translated by several
ribosomes, producing a structure called a polyribosome.

A

Each mRNA molecule may be simultaneously translated by several
ribosomes, producing a structure called a polyribosome.

28
Q

Quality control: Cells possess mRNA surveillance mechanisms that
eliminate mRNAs with errors that may create problems in translation.

A

Quality control: Cells possess mRNA surveillance mechanisms that
eliminate mRNAs with errors that may create problems in translation.

FOR STALLING AND NONSENSE MUTATION.

29
Q

understanding NONSENSE MUTATION AND HOW ARE THEY FIXED?

A

Nonsense mutations (change of an amino-acid codon to a termination codon):
- translation ends prematurely when the termination codon is encountered
- often arise in eukaryotic transcription when exons are skipped or
improperly spliced

Fixed by: nonsense-mediated mRNA decay (NMD), which results in the rapid elimination of mRNA containing premature termination codons.

30
Q

Understanding STALLING and how to FIX it:

A
  • ribosome stalls on mRNA before translation is terminated

Fixed by: nonstop mRNA decay, which results in the rapid degradation of
abnormal mRNA.

In this mechanism, the codon-free A site of the stalled ribosome is
recognized by a special protein, which binds to the A site and recruits other
proteins, which then degrade the mRNA from its 3’ end.

Molecular Genetics (21 decks)

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