35 82 The RNA Polymerase Encoded in Phage T7

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Chapter 8 / Major Shifts in Bacterial Transcription
The heat shock response begins in less than 1 min,
which is not enough time for transcription of the rpoH gene
and translation of the mRNA to yield a significant amount
of new s-factor. Instead, two other processes explain the
rapid accumulation of sH. First, the protein itself becomes
stabilized at elevated temperatures. This phenomenon can
be explained as follows: Under normal growth conditions,
sH is destabilized by binding to heat shock proteins, which
cause its destruction. But when the temperature rises, many
other proteins become unfolded and that causes the heat
shock proteins to leave sH alone and attempt to save or
degrade these other, unfolded proteins.
The second effect of high temperature on sH concentration operates at the translation level: High temperature
causes melting of secondary structure in the 59-untranslated
region of the rpoH mRNA, rendering the mRNA more accessible to ribosomes. Miyo Morita and colleagues tested this
hypothesis with mutations in the suspected critical secondary
structure region. They found that the melting temperatures of
the secondary structures in wild-type and mutant mRNAs
correlated with the inducibility of sH synthesis. We will discuss this mechanism in more detail in Chapter 17.
During nitrogen starvation, another s-factor (s54, or
N
s ) directs transcription of genes that encode proteins responsible for nitrogen metabolism. In addition, although
gram-negative bacteria such as E. coli do not sporulate,
they do become relatively resistant to stresses, such as
extreme pH or starvation. The genes that confer stress
resistance are switched on in stationary phase (nonproliferating) E. coli cells by an RNA polymerase bearing the
alternative s-factor, sS or s38. These are all examples of a
fundamental coping mechanism: Bacteria tend to deal
with changes in their environment with global changes in
transcription mediated by shifts in s-factors.
SUMMARY The heat shock response in E. coli is
governed by an alternative s-factor, s32 (sH) which
displaces s70 (sA) and directs the RNA polymerase
to the heat shock gene promoters. The accumulation of sH in response to high temperature is due to
stabilization of sH and enhanced translation of the
mRNA encoding sH. The responses to low nitrogen
and other stresses, such as starvation, depend on
genes recognized by s54 (s N) and s38 (sS), respectively.
the E. coli rsd gene. The name of the gene derives from its
product’s ability to regulate (inhibit) the activity of the major vegetative s, s70, the product of the rpoD gene. Thus,
rsd stands for “regulator of sigma D.
As long as E. coli cells are growing rapidly, most genes
are transcribed by Es70, and no rsd product, Rsd, is made.
However, as we have just seen, when cells are stressed by
such insults as loss of nutrients, high osmolarity, or high
temperature, they stop growing and enter the stationary
phase. At this point, a new set of stress genes is activated
by the new s-factor, sS, which accounts for about onethird of the total amount of RNA polymerase in the cell.
This means that about two-thirds of the s present in the
cell is still s70; nevertheless, expression of genes transcribed by Es70 has fallen by over 10-fold. These observations suggest that something else besides relative
availability of s-factors is influencing gene expression,
and that extra factor appears to be Rsd, which is made as
cells enter stationary phase, then binds to s70 and prevents its association with the core polymerase. Thus, anti-ss
can supplement the s replacement mechanisms by inhibiting the activity of one s in favor of another.
Some anti-ss are subject to control by anti-antis-factors. In sporulating B. subtilis, for example, the antis-factor SpoIIAB binds to and inhibits the activities of
two s-factors required at the onset of sporulation, sF and sG.
But another protein, SpoIIAA, binds to complexes of
SpoIIAB plus sF or sG and releases the s-factors, thus counteracting the effect of the anti-s-factor. Amazingly enough,
the anti-s-factor SpoIIAB can also act as an anti-anti-anti-sfactor by phosphorylating and inactivating SpoIIAA.
SUMMARY Many s-factors are controlled by anti-sfactors that bind to a specific s and block its binding
to the core polymerase. Some of these anti-s-factors
are even controlled by anti-anti-s-factors that bind to
the complexes between a s and an anti-s-factor and
release the s-factor. In at least one case, an anti-sfactor is also an anti-anti-anti-s-factor that phosphorylates and inactivates the cognate anti-anti-s-factor.
8.2
Anti-s-Factors
In addition to the s replacement mechanisms we have just
discussed, bacterial cells have evolved ways of controlling
transcription using anti-s-factors. These proteins do not
compete with a s-factor for binding to a core polymerase.
Instead, they bind directly to a s-factor and inhibit its
function. One example of such an anti-s is the product of
The RNA Polymerase
Encoded in Phage T7
Phage T7 belongs to a class of relatively simple E. coli phages
that also includes T3 and fII. These have a considerably
smaller genome than SPO1 and, therefore, many fewer
genes. In these phages we distinguish three phases of transcription: an early phase called class I, and two late phases
called classes II and III. One of the five class I genes (gene 1)