Small RNAs and the regulation of biological processes

Question 1

What is the C-value paradox?

Answer 1

The size of the genome is not necessarily related to its complexity.

Question 2

What are non-coding RNAs ? Give some examples.

Answer 2

Any RNA that isn’t translated into a protein (i.e. anything except mRNA) EXAMPLES of housekeeping: tRNA, rRNA, siRNA
Regulatory RNA:
microRNA
siRNA
long ncRNA ( Xist, involved in lyonisation i.e. x-chromosome inactivation)

regulatory RNA is to control what mRNA is translated via RNA interference

Question 3

Describe how RNA can be used to block mRNA function.

Answer 3

Antisense RNA can be synthesised which binds to the complementary sense RNA forming double stranded RNA thus preventing it from being translated

Question 4

what is the difference between miRNA and siRNA

Answer 4

siRNA is made exogenously (externally made) and cuts the target mRNA at one point- it can be introduced
microRNA is made endogenously and cuts the target mRNA at multiple points. It is encoded by the genome and regulates gene expression

Question 5

Describe the formation of siRNA.

Answer 5

Double stranded RNA is used to produce siRNA, which inhibits the RNA the dsRNA was produced from.
-You begin with viral dsRNA DICER cuts the dsRNA into smaller pieces (21-25bp) leaving 3’ overhang
AGO proteins then remove the passenger strand only leaving the strand that is anti-sense to the target RNA The RISC complex associates with the single stranded RNA and cleaves the target mRNA when bound with the siRNA
-if it is microRNA, several different RISCs with different sequences will be formed

Question 6

What is Dicer

Answer 6

RNASE III like endonuclease activity

Question 7

What feature of siRNA allows it to be used as a viable therapy?

Answer 7

It is small enough to enter the cell without causing a detrimental cytokine response.

Question 8

What is shRNA

Answer 8

synthesised long piece of RNA which folds back on itself

small hairpin RNA

Question 9

What did the Lin-14/Lin-4 experiment show?

Answer 9

There is genomically encoded miRNA that is involved in gene regulation.

Question 10

Describe the production of miRNA.

Answer 10

-Pri-miRNA is made and exported to cytoplasm as pre-miRNA
-processing allows the pre-miRNA to be passed to RISC complex
-pre-miRNA eventually become miRNA and target RNA to silence.
Apart from cleavage, microRNA can ALSO inhibit target mRNA through base pairing with incomplete complementarity
Some of the microRNA will be transported out of the nucleus in its pre-microRNA form
Forms a RISC complex
RISC complex attaches the microRNA to the target mRNA.
The target mRNA will not be cleaved, but is instead permanently bound to the microRNA, so is functionally useless.

Question 11

What is the classic arrangement of miRNA within the genome and what does it do when it binds to target mRNA?

Answer 11

It occurs as a match-bulge-match arrangement in the genome. One of the matching regions is usually a 3’ UTR and the other is a seed region ( primary binding site) from 5’ end.

Question 12

Give an example of an alternation in miRNA expression causing disease.

Answer 12

Chronic Lymphoid Leukaemia – deletion of part of gene on chromosome 14 leads to loss of miRNA and promotes CLL