RNA Interference vs. HIV - The Basic Mechanism

The RNA Interference process has an interesting history, and has been used by researchers in several different ways.  Because of the specificity of its gene silencing effects, it is not only used to fight deleterious genes, but also to determine gene function.  This section will show the basic mechanism of RNAi when deployed against HIV mRNA.  For details on the other aspects of RNAi, please see the RESOURCES section of this website. 

Step by Step Process

Situation: HIV Infection

Although in many cases RNAi is used to prevent HIV (before cells are infected with HIV), for the purpose of this explanation, it will be assumed that HIV has already infected the cells in question.  Like a true retrovirus, HIV has released its genome into the cell.  The RNA genome is transcribed into DNA by HIV reverse transcriptase, and has incorporated itself into the cell's own DNA.  The HIV DNA (known as the "provirus")  is producing messenger RNA transcripts of itself.

Step 1: Introduction of siRNA

During this step long double stranded RNA (dsRNA) capable of base pairing with the HIV mRNA is introduced to the cell.  Double stranded RNA is not naturally produced within cells, and can be recognized by the enzyme DICER.  The dsRNA is found and cleaved by DICER into smaller sections called siRNA (Small Interfering RNA).  siRNAs, usually 21-25 nucleotides in length, have 2 nucleotide overhangs at each end and are central to the RNAi process. 

          Current experiments avoid the DICER step, and introduce specifically engineered siRNA directly into the cell.  The siRNAs can be injected, or can be introduced through a vector--lentiviruses are commonly used as siRNA vectors.

Step 2:  Insertion into RISC complex

A double stranded siRNA molecule can insert itself into a protein complex known as RISC (RNA Induced Silencing Complex).  RISC unwinds and discards the complementary strand of the siRNA molecule.  The RISC complex, paired up with the single stranded siRNA, is the unit that interferes with HIV mRNA.

Step 3:  HIV mRNA Cleavage + Degradation

The siRNA inserted into the RISC complex base pairs with the HIV mRNA.  Remember that siRNA molecules (or long dsRNA molecules, depending on the experiment) are artificially synthesized to enable base pairing with the HIV mRNA molecule.  In reality, there are many kinds of HIV mRNAs that vary through the different stages of the virus' life cycle [the HIV InSite provides a detailed account of the mRNAs produced by HIV].

          When base pairing has occurred between the RISC siRNA and the HIV mRNA, RISC cleaves the HIV mRNA, neutralizing its chances of producing HIV associated proteins.  RNAses within the cell identify the remnants of the HIV mRNA as defective.  The final destruction of the mRNA remnants is carried out by these RNAses. 

          RISC and the inserted siRNA molecule can continue to cleave other HIV mRNA molecules.  Thus, one can see how RNA Interference can specifically cleave and neutralize deleterious mRNAs.

The SIRNA Therapeutics RNAi page provides an excellent movie on the RNAi process.  The movie is in MPEG format and is 5.06 MB in size.  Scroll down to the middle of the page, and click on the link to view.

The Nature Publishing Group provides an animated tour of the RNAi process on their RNA Interference Page.  The animation is in AVI format.  It can be watched in segments, or the complete 20.2 MB file can be downloaded.

The Nova Science Now web site provides a simplified and fun movie on the RNAi process.  The 15 minute segment uses an extensive metaphor, and includes information on the initial discovery of RNAi in 1986.  It can be viewed in Windows Media, QuickTime or RealVideo format.