miRNA

Question 1

How much of the genome is protein coding?

Answer 1

2%

Question 2

Why were worms used to discover ncRNAs?

Answer 2

Short life cycle and easy to culture.

Question 3

What is the ratio of non coding to total genomic DNA correlated to?

Answer 3

Increasing biological complexity.

Question 4

What percentage of the higher eukaryotic transcriptome does mRNA represent?

Answer 4

Around 4%.

Question 5

How long is miRNA?

Answer 5

21-23 nucleotides.

Question 6

What do they do?

Answer 6

Modulate gene expression by binding to complementary target mRNAs.

Question 7

What was the miRNA seed region?

Answer 7

Nucleotides 2-8.

Question 8

How do we predict the mRNA targets of an miRNA?

Answer 8

miRBase details correctly annotated miRNAs, DIANA predicts miRNAs that target a gene. 60% of the genome contain at least 1 miRNA binding site. Each miRNA can have many target mRNAs. Most mRNAs are target by multiple miRNAs.

Question 9

How do miRNAs exert biological function once bound to mRNA?

Answer 9

Target mRNA destabilisation, repression of translation initiation, post-initiation.

Question 10

Why is there more than one microRNA per target?

Answer 10

The codes for the target are short.

Question 11

How does miRNA function in the nucleus?

Answer 11

Associated with a smaller RISC complex. proposed role in post-transcriptional gene silencing and emerging roles in transcriptional gene silencing/activation.

Question 12

Is the deregulation of miRNAs a feature of human disease?

Answer 12

Yes lots of evidence. Disease associated variants in enzymes and miRNAs.

Question 13

What is motor neurone disease?

Answer 13

A collection of very closely related diseases of which ALS is the most common. Amyotrophic Lateral Sclerosis. Characterized by loss of upper and lower motor neurons. Failure of RNA metabolism is central to disease. Heterogenous disease. Fast and slow progressors.

Question 14

Why are miRNAs being used as biomarkers in ALS?

Answer 14

Good candidates to owing to presence/stability in blood and CSF (cerobospinal fluid). Studies to date are underpowered and lack standardisation.

Question 15

How does an miRNA mimicer function?

Answer 15

dsRNA that binds as though it is miRNA and prevents translation of target mRNA. Occurs when miRNA is downregulated.

Question 16

What is an miRNA inhibitor?

Answer 16

Inhibits miRNA which in turn helps translation to occur. Occurs when miRNA is upregulated.

Question 17

What happened when targeting miRNA dysregulation in mice occurred?

Answer 17

Deletion of miRNA in disease mouse hastened neurodegeneration and vice versa.

Question 18

Which miR-218 is shown to link to ALS?

Answer 18

Normally enriched in human spinal motor neurons but downregulated in ALS. mutant pre-miR-218 doesnt bind to DICER.

Question 19

What are limitations of miRNA as diagnostic tools?

Answer 19

Inherent lack of standardised protocols for collection and handling of samples, subsequent processing and data analysis. Requires large sample size, longitudinal studies tracked against outcomes.

Question 20

What are the barriers to successful miRNA therapies?

Answer 20

Need to define targets of miRNAs. Stability. Targeted delivery.

Question 21

What are the key challenges of miRNA delivery?

Answer 21

Immunostimulatory effects, toxicity due to charge, endosomal escape, targeting to disease site.

Question 22

What are chemical modifications to increase stability and improve targeting to disease site?

Answer 22

2’-o-methyl, LNA, phosphorothioate, peptide nucleic acids.

Question 23

What are some miRNA delivery systems?

Answer 23

Liposomes, dendrimers, PEI particles