Gene Expression

Gene expression occurs in two stages.

1) Transcription  refers to the synthesis of RNA molecule on a DNA template. The process is carried out by DNA-dependent RNA polymerase.

2) Translation: The RNA sequence (mRNA) is converted into amino acid sequence corresponding to a protein.

During the transcription three types of RNA could be produced. All the three RNAs are involved in the translation process.  The RNAs are

a) messenger RNA (mRNA)

b) transfer RNA (tRNA)

c) ribosomal RNA (rRNA)

The strand of DNA that acts as the template during transcription is template strand or antisense strand. The other strand is called the coding strand or sense strand. The sequence of coding DNA strand is the same as that of the mRNA sequence except that it possess T instead of U.

A new class of RNA molecules referred to as ultraconserved non-coding RNAs (UCRs), previously considered “junk”, are now shown to be involved in cancer development. These UCRs might prove useful in disease diagnosis, and determining a patient’s prognosis and treatment. Here is the report.

Read Full Post »

miRNA and siRNA

Both microRNAs (miRNA) and small interfering RNAs (siRNA) play integral roles in the RNAi. Both these molecules play similar roles but differ in their origin . siRNAs are derived from the cleavage of long dsRNA precursors, which could either be produced endogenously or introduced into the cell from outside. When produced endogenously, siRNAs are derived from the cleavage of long dsRNAs which are produced by RNA-dependent RNA polymerases or from transcription of genes or transposable elements. The cleavage of long precursor dsRNAs into siRNA is catalysed by the Dicer endonuclease. One of the two strands of siRNA is finally incorporated into the RISC.

miRNAs, in contrast, are endogenously derived from the cell. They are encoded within the host genome. The miRNA transcripts contain “near-complementary inverted repeats” which fold back on themselves to form hairpin or stem-loop structures. These are processed in the nucleus by the RNase III enzyme Drosha, and a protein called Pasha in Drosophila or DGCR8 in mammals. The processing reaction generates pre-miRNA, which is further cleaved by the cytoplasmic RNase III endonuclease Dicer complex to produce mature miRNA. This is then incorporated into the RISC. The miRNA could silence genes by two different modes: in plants, miRNAs cleave the cognate mRNAs; in animals, miRNAs predominantly inhibit translation by targeting partially complementary sequences in the 3´ UTR (untranslated region) of mRNAs.

Read Full Post »

RISC: RNA-induced Silencing Complex

During the course of gene silencing by RNAi the destruction of target mRNA is catalysed by the siRNA-guided, RNA-induced silencing complex (RISC). RISC, composed of two signature components (siRNA/miRNA and Argonaute), is an endonuclease , which catalyses the cleavage of a single phosphodiester bond on the RNA target. The cleavage reaction requires “Mg²⁺, but not Ca²⁺, and the cleavage product termini suggest a role for Mg²⁺ in catalysis” (Current Biology, 2004, Vol 14: 787-791). Argonaute proteins, reported from RISC complexes of diverse organisms, are the key components of RISC. The Argonaute protein family is highly diverse, and the members contain two domains: a PAZ domain, which is involved in miRNA/siRNA binding, and a PIWI domain, which is related to RNaseH endonucleases and functions in slicer activity.

Read Full Post »

RNAi: Regulating Gene Expression by RNA

RNA interference (RNAi) has emerged as one of the significant mechanisms playing a major role in gene expression regulation. The mechanism relies on diverse small noncoding RNAs, the prominent being siRNAs (small interfering RNAs) and miRNAs (microRNAs), for efficient gene silencing. The RNAi pathway is started by the enzyme Dicer, which is a ribonuclease of the RNase III family. This enzyme cleaves long double-stranded RNA (dsRNA) to short double-stranded fragments of 20–25 base pairs. One of the two strands is incorporated into the RNA-induced silencing complex (RISC). The resulting ribonucleoprotein complex (RISC-RNA) lodges itself on the target mRNA by complementary base pairing (between the mRNA and the single-stranded small RNA bound to RISC). This results into degradation of the target mRNA by the argonaute protein. Argonaute is the catalytic component of the RISC complex. The outcome of the whole process is inhibition of gene expression or gene silencing.

Read Full Post »