NAGE 5 - Small RNAs

Question 1

What is the C value paradox?

Answer 1

The complexity of an organism is not necessarily related to its genome size (c-value)

Question 2

What are non-coding RNAs?

Answer 2

Any RNA not translated into protein.

• Includes house keeping RNAs (rRNA, tRNA and splicesosome snRNAs)
• And regulatory RNAs (miRNA, siRNA, piRNA, long ncRNA)

Question 3

List the types of regulatory ncRNAs and their functions.

Answer 3

• microRNA controls the translation of genes
• siRNA/RNAi is involved in viral defence
• piRNA is important for germ cell production
• long ncRNA (Xist involved in X inactivation)

Question 4

What is lyonisation?

Answer 4

X inactivation, where one or two copies of the X chromosome present is inactivate in females. The inactive X chromosome is present as heterochromatin.

Question 5

What is antisense RNA?

Answer 5

An RNA strand complimentary to mRNA, which binds to the mRNA to form a double stranded sequence, blocking translation.

Question 6

What is RNAi and how does it work?

Answer 6

• RNA interference.
• Double stranded RNA (snRNA or miRNA) is split into duplex fragments with 3’ overhangs by the action of DICER.
• Argonaute-Piwi proteins bind to overhangs, splitting RNA into single strands.
• The RISC complex (RNA-induces silencing complexes) assembles with ATP.
• The RISC complex attaches to complimentary mRNA and cleaves it.

Question 7

How is RNAi different when microRMA or siRNA are used?

Answer 7

• siRNA is made exogenously and cuts the target mRNA at one point.
• MicroRNA is made endogenously and cuts the target mRNA at multiple points

Question 8

What is shRNA?

Answer 8

Small hairpin RNA - artificial microRNA

Question 9

What is the function of RISC complexes?

Answer 9

They recognise and cleave target mRNA molecules which have complimentary sequences to the single stranded guide (siRNA/miRNA)

Question 10

Give an example of an micro RNA.

Answer 10

• Lin-4 is a gene that codes for microRNA, which silences the mRNA which produces Lin-14.
• Therefore, the levels of Lin-4 and Lin-14 are inversely related

Question 11

Describe the process of making microRNA.

Answer 11

• miRNA is transcribed by RNA polymerase II to make long primary pri-miRNA
• Drosha makes pre-miRNA (precursor micro RNA), which may be transported out of the nucleus.
• This is cleaved to make miRNA hairpins

Question 12

Describe the structure of miRNA binding sites.

Answer 12

• miRNA binds to mRNA with two regions of complimentarity with a bulge in the middle. The “seed” region is at the 5’ end and flanked with adenosines.
• There is less stringent complementarity in the 3’ region
• Each miRNA targets several mRNAs.

Question 13

How can miRNA be used in gene therapy?

Answer 13

May be used to treat chronic lymphoid leukaemia. Involves mi15 and mi16.