[W9] RNA processing

Question 1

What is RNA processing?

Answer 1

Any post-transcriptional modification to RNA that produces mature, functional RNA.

Question 2

What are the main types of RNA processing?

Answer 2

• tRNA processing
• rRNA processing
• mRNA processing and export
• Nonsense-mediated decay
• Alternative splicing

Question 3

Why is RNA processing important in eukaryotes?

Answer 3

Most genes contain introns; processing removes these and modifies RNA for stability, export, and translation.

Question 4

What terminates tRNA transcription by RNA Pol III?

Answer 4

A poly(U) sequence.

Question 5

How are tRNA introns removed?

Answer 5

Via endonuclease cleavage and successive ligation reactions.

Question 6

What are the key steps in tRNA splicing?

Answer 6

• Endonuclease cleavage
• Phosphorylation of 5′–OH
• Phosphodiesterase opens cyclic phosphate
• RNA ligase joins exons
• Phosphatase removes 2′-phosphate

Question 7

Which polymerase transcribes most rRNAs?

Answer 7

RNA Polymerase I (Pol I).

Question 8

What are ETS and ITS in rRNA genes?

Answer 8

External and internal transcribed spacers – noncoding regions removed during rRNA processing.

Question 9

What are snoRNAs?

Answer 9

Small nucleolar RNAs that guide chemical modifications (e.g. methylation, pseudouridylation) of rRNA.

Question 10

What types of modifications do snoRNAs direct?

Answer 10

• 2′-O-methylation of ribose
• Conversion of uridine to pseudouridine

Question 11

What are the three main processing events for pre-mRNA?

Answer 11

• 5′ capping
• Splicing
• 3′ polyadenylation

Question 12

What is the 5′ cap and its role?

Answer 12

A 7-methylguanosine added via a 5′–5′ triphosphate linkage; stabilizes RNA, aids splicing, export, and translation.

Question 13

What is the polyadenylation signal?

Answer 13

The AAUAAA sequence; signals cleavage and addition of ~200 A residues by poly(A) polymerase.

Question 14

Why is the poly(A) tail important?

Answer 14

Enhances mRNA stability, export from nucleus, and translation initiation.

Question 15

What is splicing?

Answer 15

The removal of introns and joining of exons from pre-mRNA.

Question 16

What is the GU-AG rule?

Answer 16

Most introns begin with GU and end with AG (5′ and 3′ splice sites).

Question 17

What structure is formed during splicing?

Answer 17

A lariat structure, involving a 2′–5′ phosphodiester bond at the branch site A.

Question 18

What are snRNPs?

Answer 18

Small nuclear ribonucleoproteins; complexes of snRNA + proteins that form the spliceosome.

Question 19

Which snRNPs are involved in splicing?

Answer 19

• U1
• U2
• U4
• U5
• U6

Question 20

What are the phases of spliceosome assembly?

Answer 20

• Phase I: Commitment
• Phase II: Assembly (complexes E, A, B1, B2)
• Phase III: Catalysis (complexes C1, C2)

Question 21

What is transesterification in splicing?

Answer 21

A coordinated bond exchange forming the exon–exon junction without requiring ATP.

Question 22

Is splicing coupled with transcription?

Answer 22

Yes — splicing often occurs co-transcriptionally and is integrated with RNA Pol II activity.

Question 23

What is the exon junction complex (EJC)?

Answer 23

A complex deposited at exon–exon boundaries that assists in mRNA transport, localization, and decay.

Question 24

What is NMD?

Answer 24

A surveillance pathway that degrades mRNAs with premature stop codons (PTCs).

Question 25

When does NMD occur?

Answer 25

If the stop codon is upstream of the last exon or >50 nt upstream of the last exon–exon junction.

Question 26

What proteins mediate NMD?

Answer 26

UPF proteins recruit the decapping complex (DCP), which removes the 5′ cap and triggers mRNA degradation.

Question 27

What is alternative splicing?

Answer 27

Production of different mature mRNAs from the same pre-mRNA by varying splice site usage.

Question 28

How common is alternative splicing?

Answer 28

~90% of human genes undergo alternative splicing.

Question 29

What are splicing enhancers and silencers?

Answer 29

* Sequence elements that promote (ESE, ISE) or repress (ESS, ISS) splice site usage.

Question 30

Which proteins bind splicing enhancers and silencers?

Answer 30

* SR proteins → bind ESE * hnRNPs → bind ESS * RBPs → regulate ISE/ISS

Question 31

What regulates alternative splicing?

Answer 31

RNA-binding proteins (RBPs), their expression, location, and interaction with the spliceosome.

Question 32

Can mis-splicing cause disease?

Answer 32

Yes — mutations in splicing elements or RBPs can lead to defective transcripts and disease.

Question 33

Give an example of functional impact of alternative splicing.

Answer 33

Tropomyosin isoforms affect actin binding and motor recruitment in muscle vs non-muscle tissues.

Question 34

What is the transcriptome?

Answer 34

The complete set of RNAs (coding and non-coding) in a cell, tissue, or organism.

Question 35

What is RT-PCR?

Answer 35

Reverse transcription followed by PCR; converts RNA to cDNA for amplification.

Question 36

What is qRT-PCR?

Answer 36

Quantitative real-time PCR; allows sensitive, quantitative detection of gene expression.

Question 37

What is Northern blotting?

Answer 37

Technique for RNA detection using membrane transfer and hybridization with labeled probes.

Question 38

What is RNA-seq?

Answer 38

A high-throughput sequencing method that measures RNA expression across the transcriptome.

Question 39

What are DNA microarrays?

Answer 39

Chips containing DNA probes for genome-wide expression analysis via cDNA hybridization.