Page 60 - Academic Press Encyclopedia of Physical Science and Technology 3rd Molecular Biology

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Encyclopedia of Physical Science and Technology EN006H-655 June 29, 2001 21:21

Gene Expression, Regulation of 509

precursor-RNA synthesized by RNA polymerase II un- approximately double the size of most bacterial genomes
dergoes several posttranscriptional modifications before a and takes days to transcribe. In contrast, exons are typi-
mature mRNA is formed. For example, the transcript is cally short, usually less than 350 nucleotides. This comes

capped at its 5 end, the 3 end is generated by a specific from the fact that splice sites used to define the borders of
cleavage polyadenylation reaction, and intronic sequences the splicing reaction are defined across the exon, not the
are removed by RNA splicing. intron—the so-called exon definition model (see below).
Early after initiation of transcription, when the nascent Some eukaryotic genes are remarkably large. For exam-

RNA chain is 25–30 nucleotides long, the 5 end is modi- ple, the human gene for dystrophin covers approximately
fied by addition of an inverted 7-methylguanosine, the cap 2.4 million base pairs. The RNA polymerase that initi-
nucleotide. The capping enzymes are brought to the tran- ates transcription requires approximately 20 hr to synthe-
scribing polymerase by specific association with the hy- size the full-length precursor-RNA. Subsequently, more
perphosphorylated form of the CTD tail on RNA poly- than 99.5% of the transcript is removed by RNA splicing.
merase II. As mentioned above, the CTD tail becomes Thus, the final mRNA that is transported to the cytoplasm
phosphorylated when RNA polymerase II progress from is only around 14,000 nucleotides. The extreme lengths
the initiation to the elongation phase of RNA synthesis. of eukaryotic genes place a high demand on the stability
Since RNA polymerases I and III do not have a CTD tail, of the transcription complex. Thus, an RNA polymerase
only RNA polymerase II transcripts are capped. The cap that binds to a promoter must stay attached for days with
plays a crucial role in initiation of translation by binding the DNA template to be able to complete synthesis of the
the translational initiation factor eIF4F required for the longest genes.
recruitment of the small subunit of the ribosome to the It is interesting to note that introns in eukaryotic genes
mRNA. The translational start site is then identified by a almost always interrupt the protein-coding portion of the
scanning mechanism where the ribosome usually selects precursor-RNA. Thus, introns are rarely found after the

the first AUG triplet as the start codon for protein synthe- translational stop codon, within the 3 noncoding portion
sis. The selective addition of a cap to RNA polymerase of the mRNA. This organization is significant since the
II transcripts therefore provides a logical explanation to presence of an intron downstream of the translational stop
why this class of RNAs is used for translation. Polyadeny- codon in a reading frame is sensed as a signal that the
lation and RNA splicing are key mechanisms to regulate precursor-RNA has been incorrectly spliced or for other
eukaryotic gene expression and are therefore described in reasons is defective, and will not produce the correct pro-
more detail below. tein after translation in the cytoplasm. Such nuclear tran-
scripts are sent for destruction by a mechanism that is
collectively called the non-sense-mediated mRNA decay
A. Exons and Introns: General Considerations
mechanism.Howthetranslationalreadingframeisreadal-
Virtually all prokaryotic genes are encoded by a colli- ready in the nucleus is not known. The easiest explanation
near DNA sequence: the concept one gene, one mRNA. wouldbethatthereexistsanuclearribosome-likestructure
In contrast, most eukaryotic genes are discontinuous, with that scans the spliced mRNA for a full-length translational
the coding sequences (exons) interrupted by stretches reading frame before the mRNA is transported to the cy-
of noncoding sequences (introns). Introns are present at toplasm. However, this question is controversial and has
the DNA level and in the primary transcription product not been proven.
of the gene (the precursor-RNA), and are removed by
RNA splicing before the mature mRNA is transported to
1. Mechanism of RNA Splice Site Choice During
the cytoplasm. Recent experiments suggest that splicing
Spliceosome Assembly
is necessary for efficient transport of intron-containing
precursor-RNAs. Introns have been found in all types of The sequence elements used to specify the splice sites
eukaryotic RNA—mRNA, rRNA, and tRNA. Because of are remarkably short and degenerate in a eukaryotic
space limitation, only introns in protein-encoding genes precursor-RNA. Thus, short conserved sequence motifs

will be described. at the beginning (5 end) and the end (3 end) of the intron

The number of introns in mRNA-encoding genes varies guide the assembly of a large RNA protein particle, the
considerably among genes. For example, c-jun, histone, spliceosome (Fig. 7), which catalyzes the cleavage and
heat-shock, and the α-interferon genes have no introns, ligation reactions necessary to produce the mature cy-
whereas the gene for dystrophin has more than 70 introns. toplasmic mRNA. The nucleus of eukaryotic cells con-
Also, the size of introns can vary from less than 100 nu- tains several abundant low-molecular-weight RNAs, so-
cleotides to several million nucleotides in length. The ex- called U snRNAs. The U snRNAs derive their name
treme example is the Drosophila Dhc7 gene, which con- from the fact that they were initially characterized as
tains a 3.6 million-nucleotide-long intron. This intron is RNAs rich in uridines. Five of these U snRNAs (U1,