Genetics: Mirna And Sirna

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function of miRNA

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regulate gene expression usually by blocking the translation of selective mRNAs

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function of siRNA

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turn off gene expression by directing degradation of selective mRNAs and establishing compact chromatin structure

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What are miRNAs?

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small noncoding RNA molecules that are *very conserved evolutionarily*

They regulate gene expression at the level of translation either by turnover of messenger RNA or interfering with translation.

They express a transcript that is capped and polyadenylated.

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miRNA synthesis

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miRNA genes are regulated by RNA Polymerase II, which means they are regulated. They give rise to the Primary-miRNA which has a secondary structure with hairpin loops and turns and a 3' overhang.

The Primary-miRNA is then cropped, which gives rise to the Pre-miRNA complex. This is then processed by Dicer, which is an RNase III, along with other proteins, such as Argonaute.

This yields a large protein complex, which binds to the duplex region of the miRNA which removes the complement portion and gives rise to the RNA Induced Silencing Complex (RISC).

The miRNA of the RISC is used as a guide strand to lead the complex to the proper strand it wants to meet. Typically, the RISC binds to the mRNA in the 3' noncoding sequence.

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Argonaute

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wields the miTNA hoping to find a strip of RNA that complements with what it has.

Typically, the miRNA targets the 3' untranslated region.

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miRNA matchmaking

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when matchmaking occurs between the miRNA and its complement, the level of match determines what happens next.

Typically, an incomplete match occurs that simply yields translational inhibition.

However, as a result of RISC binding to the mRNA, P-bodies are targeted.

These P-bodies exist in the cytoplasm of the cell where RNA is typically turned over.

This will either mean:
1. the RNA will be turned over and degraded (starting with the 5' end), or
2. stored and then released


The meeting of the RISC complex with the appropriate mRNA will yield translational repression and ultimately turnover

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P-Bodies

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"Processing Bodies"

When mRNA are bound to miRISC and being repressed they will be segregated into a P-Body in the cytoplasm.

In these P-bodies, they can be degraded or stored for later release and translation.

So this whole mechanism gives you translation repression and RNA turnover.

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Translation Suppression in P-Bodies: Animals vs. Plants

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In plants you get a similar scenario.

The difference comes in when miRISC binds the target mRNA. In plants, the miRNA provides a perfect match to the target mRNA. So of the 19-23 nucleotides, most of them will bind to the mRNA, and any part that doesn't serves as an incision point. An enzyme can come in and cut to create a starting point for degradation.

In animals, only ~7 nucleotides on the miRISC bind to the mRNA, and the complex needs to redirect it to the P-body for eventual degradation.

So in plants, the process is quicker and more dramatic.

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difference between miRNA and siRNA

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siRNA can go through the same RISC pathway as miRNA, but they also have another option- siRNA can cause transcriptional silencing.

The pathway is same up to the binding of Argonaute. At this point, Argonaute can assemble either RISC proteins or RITS (RNA-Induced Transcriptional Silencing) proteins.

These are chromatin binding proteins, and can silence areas of the genome through histone methylation, DNA methylation, and transcriptional repression.

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Delivery of siRNAs

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injecting an siRNA that is complementary to a certain mRNA will result in it being taken up by RISC and can significantly down regulate the mRNA.

The clinical implications here are significant (imagine down regulating specifically targeted cancerous cells)

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siRNA administered via targeted nanoparticles

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Using siRNA for RNA interference.

Researchers built nano particles that contained siRNA and coated them with the protein transferrin, which is a blood plasma protein for iron delivery.

Tumor cells express receptors for this protein abundantly because they need iron to grow, and so they take up these nano particles containing siRNA.

The end result was that the siRNA was found primarily in tumor cells. The siRNA was targeted agains Ribonuclease Reductase, which is a common target of chemotherapy agents. Down regulated the expression of the tumors.

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Genomics of miRNA

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Since miRNAs are RNA Polymerase II transcripts, they have promoter regions, introns, exons, etc.

61% of miRNA genes are found in intronic regions of protein-coding translational units.

18% are found in intronic regions of non-coding translational units (meaning these TUs are coding for RNA, not protein).

20% are found in exonic regions of non-coding TUs.

miRNAs are VERY conserved throughout evolution, meaning it is part of some very important gene regulation system.

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shared principles of Transcription Factors and miRNA action

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Pleiotropy- the ability to act on many different genes.

Combinatorial & Cooperative Activity- at site of initiation, it is not a single transcription factor, but a combination and the results give the final transcriptional result.

Accessibility- find ways to access compact DNA.

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miRNAs as Oncogenes and Tumor Suppressors

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miRNAs can be involved in cancers by directly regulating cell growth or indirectly controlling apoptosis through targeting transcription factors or signaling pathways.