transfected with lacZ dsRNAs only showed degradation in lacZ transcripts and not cyclin E transcripts. Their results led Hannon and his colleagues to suggest RNAi degrades target mRNA through a 'sequence-specific nuclease activity'. They termed the nuclease enzyme RISC. Later Devanand Sarkar and his colleagues Prasanna K. Santhekadur and Byoung Kwon Yoo at the Virginia Commonwealth University elucidated the RISC activity and its molecular mechanism in cancer cells and they identified another new component of the RISC, called AEG-1 [47]. Function in RNA interference Incorporation of siRNA/miRNA The RNase III Dicer is a critical member of RISC that initiates the RNA interference process by producing double-stranded siRNA or single-stranded miRNA. Enzymatic cleavage of dsRNA within the cell produces the short siRNA fragments of 21-23 nucleotides in length with a two-nucleotide 3' overhang. Dicer also processes pre-miRNA, which forms a hairpin loop structure to mimic dsRNA, in a similar fashion. dsRNA fragments are loaded into RISC with each strand having a different fate based on the asymmetry rule phenomenon, the selection of one strand as the guide strand over the other based on thermodynamic stability. The newly generated miRNA or siRNA act as single-stranded guide sequences for RISC to target mRNA for degradation. The strand with the less thermodynamically stable 5' end is selected by the protein Argonaute and integrated into RISC. This strand is known as the guide strand and targets mRNA for degradation. The other strand, known as the passenger strand, is degraded by RISC. Gene regulation Major proteins of RISC, Ago2, SND1, and AEG-1, act as crucial contributors to the gene silencing function of the complex. RISC uses the guide strand of miRNA or siRNA to target complementary 3'-untranslated regions (3'UTR) of mRNA transcripts via Watson-Crick base pairing, allowing it to regulate gene expression of the mRNA transcript in a number of ways. mRNA degradation The most understood function of RISC is degradation of target mRNA which reduces the levels of transcript available to be translated by ribosomes. The endonucleolytic cleavage of the mRNA complementary to the RISC's guide strand by Argonaute protein is the key to RNAi initiation. There are two main requirements for mRNA degradation to take place: a near-perfect complementary match between the guide strand and target mRNA sequence, and, a catalytically active Argonaute protein, called a 'slicer', to cleave the target mRNA. There are two major pathways of mRNA degradation once cleavage has occurred.