Ulb

INSERTION OF 3 U'S

FIGURE 12.24 Editing of Trypanosome mRNA

The guide RNA base pairs along a specific region of the trypanosome mRNA. The extra adenine (A) of the AGA sequence in the guide RNA is used as a template to insert uracil (U) into the mRNA. (Note: some slightly distorted duplex RNA structures allow guanine, as in the AGA sequence shown, to pair with uracil.)

Trypanosomes frequently edit their mRNA by inserting or removing bases.

C to U and U to C editing of mRNA are also found in the mitochondria and chloroplasts of most major groups of plants. Typically from three or four to twenty bases are changed in those transcripts that are edited in plant organelles. In most cases, such editing results in changes in the amino acid sequence of the encoded protein that are necessary for full activity. However, silent editing is occasionally observed. The ultimate in pointlessness appears to be the case of the tobacco chloroplast atpA gene where a CUC codon is edited to CUU in the mRNA. Both codons encode serine. Conceivably, such silent editing might be an adjustment to the differential availability of tRNAs with different anticodons, however there is no evidence for this.

Trypanosomes practice RNA editing relatively often. Moreover, they do not merely modify bases chemically but actually insert or remove them. Some of the primary transcripts of trypanosomes, especially those from the mitochondrial genes, are altered by insertion or removal of multiple uridine nucleotides, one at a time, before the final mRNA is generated (Fig. 12.24). In these cases, the coding sequences found on the DNA have incorrect reading frames. If the trypanosome did not edit its mRNA, the result would be defective, frame-shifted proteins made from out of phase coding sequences.

Multiple insertions of U in trypanosome mRNAs occur at positions specified by short guide RNAs. These are complementary to short stretches of the mRNA but have an extra A. The U residues are inserted into the mRNA opposite the extra A on the guide RNA.

guide RNA Small RNA used to locate sequences on a longer mRNA during RNA editing

FIGURE 12.25 Transport of Eukaryotic mRNA out of the Nucleus

Processed mRNA is free to leave the nucleus whereas primary transcript still attached to snRNA is prevented from leaving.

FIGURE 12.25 Transport of Eukaryotic mRNA out of the Nucleus

Processed mRNA is free to leave the nucleus whereas primary transcript still attached to snRNA is prevented from leaving.

Transport of RNA out of the Nucleus

The nucleus is surrounded by a double membrane. Each nucleus has many pores that allow molecules in or out in a carefully controlled manner. Each nuclear pore is surrounded by a cluster of proteins that control entry and exit, but the details of precisely which molecules are allowed in or out are still murky. We do know that once messenger RNA has received its cap and tail and had its introns spliced out, it is free to exit the nucleus. Binding of the spliceosome to the RNA prevents it from leaving until splicing is finished (Fig. 12.25).

Transport out of the nucleus of large molecules like RNA and proteins requires energy. This is obtained by the hydrolysis of GTP. Protein factors known as exportins and importins control the exit and entry of specific classes of molecules through the nuclear pores. For example exportin-t is specific for export of tRNA.

Messenger RNA is degraded after a relatively short lifetime.

Eukaryotic mRNA must have the tail and cap removed before degradation can proceed.

Degradation of mRNA

Messenger RNA molecules are relatively short-lived and in bacteria the half-life is generally only a couple of minutes. Messenger RNA that is not bound to ribosomes is especially vulnerable to degradation. Bacteria contain multiple ribonucleases. These are involved both in processing of tRNA and rRNA and the degradation of mRNA. These ribonucleases can often substitute for one another at least to some extent and so mutants that have lost only one ribonuclease are usually still viable. Bacterial mRNA is degraded in two stages (Fig. 12.26). First, an endonuclease, usually ribonuclease E, cleaves regions that are unprotected by ribosomes. Next, exonucleases that move in a 3' to 5' direction degrade the fragments. Note that overall degradation moves in a 5' to 3' direction, due to the endonuclease following the ribosomes.

Degradation of mRNA follows a different route in eukaryotes such as yeast. First the poly (A) tail and then the cap must be removed before actual nuclease digestion. Once the poly (A) tail is shortened to 10-20 bases the poly (A)-binding protein (PABP) is released. Only after the PABP has gone can the cap be removed. Once the cap has also gone, an exonuclease, Xrn1, degrades the mRNA in the 5' to 3' direction (Fig. 12.27).

endonuclease A nuclease that cuts a nucleic acid in the middle exonuclease A nuclease that cuts a nucleic acid at the end nuclear pore Pore in nuclear membrane that allows proteins and RNA into and out of the nucleus ribonuclease A nuclease that cuts RNA

Translation

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