Double-stranded RNA (dsRNA) is the trigger of RNAi, triggering the degradation of the complementary single-stranded RNA. After processing by Dicer, the longer dsRNA is degraded into siRNA (small interfering RNA) and effectively locates the target mRNA.

The antisense strand of siRNA guides the synthesis of a nuclear protein called RISC, which is then mediated to cleave the region of the target mRNA molecule that is complementary to the antisense strand of siRNA, thus achieving the function of interfering with the expression of the target gene.

siRNA can be used as a special primer to synthesize dsRNA using the target mRNA as a template under the action of RDRP. The dsRNA can then be degraded into new siRNA and re-enter the cycle.

Dicer: A ribonuclease similar to RNase III, it first degrades dsRNA entering the cell into small fragments, called small interfering RNA or microRNA (miRNA). Dicer also assists in the binding of these small RNA molecules to RISC

RISC (RNA-induced silencing complex): a large multi-subunit protein complex that guides the siRNA or miRNA bound to it to bind to the target and inhibit gene expression

Triggers degradation of mRNA encoded by target genes

Inhibit translation of target mRNA

Causes transcriptional silencing of the target promoter (modifies the chromatin where the target gene is located to silence its transcription)

RNA polymerase II produces the longer primary transcript pri=miRNA

The first step of cleavage is completed by Drosha enzyme. It is cleaved in the nucleus. The cleavage product is pre-miRNA, with two amino acids protruding from the 3' end to facilitate identification before the next step of cleavage.

The second step of cleavage is completed by the Dicer enzyme, which cleaves in the cytoplasm. The product is double-stranded miRNA, which is melted to form mature single-stranded miRNA.

① Formation of small double-stranded fragments after cleavage by Dicer: Dicer's two RNase III domains form an intramolecular dimer, each catalyzing the shearing of one strand, resulting in a double-stranded break.

②Assembly complex: The loading of siRNA requires the help of the double-stranded RNA binding protein R2D2. R2D2 contains two double-stranded RNA binding domains one after the other. Dicer/RAD2/siRNA forms a RISC loading complex

③ Formation of an active silencing complex: R2D2 protein recruits Argonaute protein, protein A interacts with Dicer to generate exchange, and then exchanges with R2D2 to transfer the entire doublet small RNA to protein A, which is degraded to form a functional silencing complex things.

Application in mammals: transfection of exogenous siRNA into cells (transient expression)

Express siRNA in cultured cells or animal models

Research applications: new strategies for reverse genetics and new methods for gene knockout

Therapeutic applications: Inventing new drugs and new strategies for gene therapy

At the transcriptional level, the post-transcriptional level is involved in the regulation of gene expression.

Maintain genome stability

Protecting the genome from exogenous accounting intrusions

Localization of mRNA in eggs and embryos by polarity relative to the cell's native skeleton

Examples: Targeted Ash1 repressor protein controls yeast mating type by silencing H0 gene, localized mRNA initiates muscle differentiation in ascidian embryos

Both cell-cell contact and secreted signaling molecules trigger gene expression in neighboring cells.

Examples: Cell-cell contact activates differential expression in spore-forming bacteria Bacillus subtilis; Notch signaling in the central nervous system of insects controls neuromodulatory switching in the skin

Gradients of secreted signaling molecules guide cells to follow position-specific developmental pathways (the cell's position in the embryo)

Example: Sonic hedgehog morphogen concentration gradient controls the formation of different neurons in vertebrates