Cell Nucleus Biogenesis, Structure and Function
381
2.1
Gene Expression
Eukaryotic gene expression follows the
general principles developed during the
evolution of prokaryotes. Put simply, gene
expression is dictated by DNA sequences
within gene promoters that determine
how the RNA synthetic machinery, RNA
polymerase, is positioned on the gene.
In multicellular eukaryotes, highly com-
plex patterns of gene expression have
evolved
and
correspondingly
complex
mechanisms of gene regulation are seen.
Nevertheless,thesamebasicprincipleun-
derlies the activation of RNA synthesis.
In contrast to prokaryotes, which have
a single RNA polymerase, three different
RNA polymerase complexes perform RNA
synthesis in eukaryotic cells. In most
mammalian cells, RNA polymerase II (pol
II) is the major activity, transcribing all
protein-coding genes to generate patterns
of gene expression that determine cell
type; humans have about 250 distinct
cell types. Synthesis is performed by an
4 MDa holoenzyme containing the pol II
core enzyme and other activities required
during RNA synthesis and processing.
RNA polymerase I (pol I) is dedicated to the
synthesis of the repeated ribosomal RNA
(rRNA) genes, within specialized nuclear
sites – nucleoli – and RNA polymerase III
(pol III), a minor nucleoplasmic activity,
transcribes transfer RNA (tRNA) and 5S
rRNA genes. Small nuclear RNA (snRNA)
and small nucleolar RNA (snoRNA) genes
encode structural RNAs needed for RNA
processing; some are transcribed by pol II
and others by pol III.
A proliferating mammalian cell supports
a continuous rate of RNA synthesis of
2
×
10
8
nucleotides min
1
.Th
isw
i
l
lbe
roughly 40% pre-rRNA, 60% pre-mRNA
(hnRNA) and only 2 to 3% 4 to 5S RNA
(5S and pre-tRNA). Synthetic rates are
3t
o4k
bpm
in
1
for pol I and 1.5 to
2kbp min
1
for pol II/III respectively
and average transcript lengths of
13.5,
15 and
0.1 kb for pol I, pol II, and pol
III. These values correspond to
20 000,
60 000 and
3000 engaged pol I, II, and
III complexes per cell. It is also important
to consider the density of pol complexes on
individual genes. A chromatin spreading
technique called
Miller spreads
can be used
to show that the active rRNA genes of
mammalian cells each support 100 to 120
pol I complexes. In sharp contrast, pol II
transcription units generally support very
few widely dispersed active complexes.
Even when studying adenovirus at the
time of the highest viral transcription, the
active genomes have only four (on average)
associated transcription complexes.
Most
tissues
of
mammalian
origin
express at least 10 000 genes. In rat
liver, for example, there are
10 species
present at
10 000 copies per cell, 500
at
200 copies per cell and
10 000
at
10 copies per cell (33) –
300 000
mRNAs per cell, in all. Note that even
after accounting for mRNA turnover, it is
clear that the majority of genes generate
so few transcripts that they must be
transcriptionally inert for most of the time.
In addition, it is interesting that many
pol II complexes seem to be involved in
synthesis that generates RNA molecules
that turnover in the nucleus. Many of these
are intergenic transcripts that have been
speculated to play a role in maintaining
an active chromatin status across extended
gene domains.
2.1.1
Activating Gene Expression
The critical feature of gene expression
in eukaryotes is that the transcription
machinery is recruited to genes through its
interaction with transcription factors and
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