Biology Reference
In-Depth Information
The 3
′
end of eukaryotic RNA is modified by adding 40-200 adenine (A) resi-
dues to a region near the 3
′
end of the transcript to produce the
poly(A) tail
(
Figure 2.5
). The polyadenylation does not simply add the A residues to the end
of the transcript. First, a cleavage occurs between 10 and 30nt downstream of
a specific polyadenylation signal; in insects, this signal is usually AAUAAA and
is found in the 3
′
noncoding region of the RNA. This cleavage results in an
intermediate 3
′
end to which the poly(A) tail is added by the enzyme poly(A)
polymerase. The length of the poly(A) tail may determine how long the mRNA
survives in the cytoplasm before being degraded.
2.7 Splicing Out the Introns
The third modification of the pre-mRNA involves splicing to remove any introns.
Splicing takes place in two steps. Introns have a 5
′
donor and a 3
′
acceptor
end with common consensus sequences (
Figure 2.5
). The 5
′
donor end typically
has the sequence GGUAAGU. After a cut in the donor site, the G at the 5
′
end
forms a loop by attaching to an A nucleotide a short distance upstream from
the pyrimidines near the acceptor splice site. The consensus sequence of the 3
′
acceptor site is AGG. In the final step, a cut is made in the acceptor site, and the
intron is freed. The exons are then joined together. The excised loop (intron) is
released as a lariat-shaped structure and is later degraded (
Figure 2.5
).
RNA splicing occurs in large multicomponent complexes called
spliceosomes
.
Spliceosomes are composed of
>
50 proteins and five types of snRNA molecules
(U1, U2, U4, U5, and U6) (
Reed 2000
). The active catalytic components of the
spliceosome are the snRNAs rather than the proteins, although the proteins are
required and participate in both assembly of the spliceosome and the splicing
reaction. In addition, several proteins play auxiliary roles in splicing and spliceo-
some assembly.
Because many pre-mRNAs contain multiple introns, the splicing machinery
must be able to identify and join the appropriate 5
′
end and 3
′
splice sites to
produce a functional mRNA. The specificity of the splicing operation is deter-
mined by the snRNAs that contain sequences that are complementary to the
splice junctions (
Cooper 2000, Hastings and Krainer 2001
). Introns may have no
function, and synthetic genes lacking introns can function quite well. However,
some introns are important in gene regulation and determine when, or in
what tissue, the gene will be transcribed. For example, sex determination in
Drosophila melanogaster
depends on a cascade of splicing and the pre-mRNA
of the
double-sex
+
gene of the female contains exon 4, whereas males lack
it. Likewise, the splicing of
Sex lethal
+
and
transformer
+
varies by sex (see
Chapter 10).
