Molecular Genetics Lecture 11 - Non-coding RNAs

What is a coding RNA?

mRNA which codes for polypeptides/proteins

What is a non-coding RNA?

transcribed into RNA but not translated into proteins

What are small non-coding RNA (<200 nt)?

microRNA (miRNA), small interfering RNA (siRNA), crRNA

What are long non-coding RNAs (>200 nt)?

IncRNA

What do non-coding RNAs regulate?

gene expression at transcriptional and post-transcriptional levels

What are MicroRNA (miRNA) and small-interfering RNA (siRNA)?

small regulatory RNAs

What is the function of MicroRNA (miRNA) and small-interfering RNA (siRNA)?

regulate the expression and degradation of mRNAs

What is CRISPR RNA (crRNA)?

small regulatory RNA found in prokaryotes that guide endonuclease to foreign DNA and some CRISPR-Cas systems can target RNA

What is the function of CRISPR RNA (crRNA)?

genome defense

What are the functions of small interfering RNA (siRNA)?

genome defense, involved in the suppression of exogenous RNA sequences especially from viruses by RNA interference (RNAi)

What is the function of RNA interference (RNAi)?

tool in gene therapy used to control gene expression

What is the function of MicroRNA (miRNA)?

regulates endogenous gene expression

The human genome contains about _____ miRNA loci

2600

What human diseases do miRNAs implicit?

cancer, diabetes, Alzheimer’s, Parkinson’s

How are miRNA and siRNA used as tools in functional genomic studies?

used to silence genes to understand gene function

What is the name of the enzyme that is at the start of the biosynthetic pathway for anthocyanins?

Chalcone synthase (CHS)

What is the result of this?

regular amount of anthocynanin pigment proteins translated

What is the result of this?

Extra protein translated, overproduction of CHS protein, enhanced purple color

Both sense and antisense sequences of the CHS gene cause ______

reduced pigmentation

Co-suppression is caused by ____ ____

small RNAs

What is the result of this?

sense-antisense duplex forms and prohibits translation

What is the result of this?

co-suppression

What is the major discovery of RNA interference in C. elegans when sense, antisense or double-stranded RNAs that are homologous to the unc-22 gene were introduced into C. elegans?

the silencing of the unc-22 gene causes loss of muscle control

What does ‘unc’ stand for in unc-22 gene?

uncoordinated

What happens when the unc-22 gene is silenced in C. elegans?

causes loss of muscle control

What is the first step of Mechanisms of RNA interference by siRNA?

Generation of siRNA: Dicer endonuclear cleaves dsRNA into 21-23 bp siRNA

What is the second step of Mechanisms of RNA interference by siRNA?

RNA-induced silencing complex (RISC) assembly: passenger strand is removed by Argonaute (Ago) and guide strand of siRNA is loaded into the RISC complex

What is the third step of Mechanisms of RNA interference by siRNA?

Formation of mRNA-siRNA-RISC complex: guide siRNA binds to the target mRNA via complementary base pairing

What is the fourth/last step of Mechanisms of RNA interference by siRNA?

mRNA degradation: Ago protein cleaves mRNA which silences the target gene

What process silences transposons?

RNA inference

What is DNA methylation induced by?

small RNAs

What are the three major steps of silencing transposons?

1. RNAi: activates RNAi pathway, generates siRNA, siRNA degrades mRNA

2. RNA-dependent DNA methylation

3. Histone methylation (H3K9me2) and formation of heterochromatin

4. Transposon is completely silenced

What are the 7 steps of Biogenesis and gene regulation by microRNA (miRNA)?

1. Generation of precursor microRNA (premiRNA): RNA polymerase transcribes miRNA gene —> pri-mRNA

2. Processing of pri-mRNA: Ribonuclease III Drosha cleaves pri-mRNA into 70 bp precursor mRNA (Pre-mRNA)

3. Transport pre-mRNA to a cytoplasms via Ran GTP and Exportin 5 proteins

4. Generation of 22 bp miRNA: miRNA duplex by Dicer enzyme and removal of passenger strand in miRNA duplex by Ago protein

5. Generation of mature guide miRNA and RISC assembly including Ago protein

6. mRNA cleavage: Ago cleaves mRNA if guide miRNA sequences are perfectly complementary to the target

7. Transcriptional repression: imperfect complementary of miRNA to target mRNA —> inhibition of translation