Figure 1.
Model for sigma factor competition.
(A) Model for sigma factor competition with two types of sigma factors, the housekeeping sigma factor and a generic alternative sigma factor
: the model describes binding of
or
to core RNA polymerase (
) to form holoenzymes (
and
) as well as transcription (promoter binding, transcription initiation and elongation) of the cognate genes and non-specific binding of holoenzymes and core RNAPs to DNA. (B) Core model for holoenzyme formation.
Table 1.
Values adopted in the simulations.
Figure 2.
(A) Number of holoenzymes and
as a function of the copy number of alternative sigma factors. Quantities of all molecular species are expressed as absolute numbers per cell. The gray dashed line represents the onset of the competition, when
. The values of the parameters used in the simulations are summarized in Table 1. (B) Determination of the sigma-core dissociation constants for
and
(see Table 2) by fitting the results of binding assays between cores and sigma factors [30], [39]. The number of core-sigma complexes normalized to the maximal number of holoenzymes,
. Stars show the experimental data and lines are due to the fit. (C) Comparison of model predictions (lines) with an in vitro competition experiment [30] with a fixed amount of core and different equimolar amounts of
and
(stars) in the same conditions as in (B). The plot shows the fraction of sigma factors bound in holoenzymes as a function of the total sigma factor concentration,
.
Table 2.
Fit values.
Figure 3.
(A) Normalized transcription rate (Equation 12) for a σ70-dependent promoter as a function of the number of alternative sigma factors. The numbers of
and cores are fixed. The blue line is for a saturated promoter (with
M) and the cyan line for an unsaturated promoter (with
M). (B) Comparison of model predictions (lines) with an in vitro competition experiment [29] with a fixed amount of core and σH and different amounts of
(stars). The plot shows the transcription rate of a σH-dependent gene (normalized to the maximal value) as a function of the concentration
. (C) The sigma-core and the holoenzyme-promoter dissociation constants (see Table 2) are determined by fitting the results of transcription rate experiments with a fixed amount of cores in the same conditions as in (B) without competition in the presence of a DNA template containing σH- and σ70-driven genes [29], [32]. (D) When a σ70-dependent promoter also binds another type of holoenzyme or overlaps to another promoter,
also acts as a repressor of the σ70-dependent transcription. (E) Normalized transcription rate of a saturated and unsaturated σ70-dependent promoter as a function of the number of
(blue and cyan solid lines with
M and
M, respectively). The dashed line show the corresponding results in the absence of repression by promoter sharing or overlapping.
Figure 4.
Effect of non-specific binding of holoenzymes and cores to DNA.
(A) Formation of holoenzymes in the presence of one type of sigma factor in the absence of DNA (no non-specific binding, dashed line), in the presence of DNA with equal non-specific binding affinities of cores and holoenzymes ( M, dotted line) and with different non-specific binding affinities (
M,
M, solid line). (B) Number of free cytoplasmic holoenzymes
and
(upper row) and total number of holoenzymes (free and non-specifically bound,
, lower row) as functions of the copy number of alternative sigma factors for three different combinations of non-specific binding affinities: in (i) and (ii) all non-specific dissociation constant are equal (
M), in (iii) and (iv) the non-specific dissociation constant for the core is smaller than for the holoenzymes (
M,
M), in (v) and (vi) the non-specific dissociation constant for the
is smaller than for
and core (
M,
M). The dashed lines in all panels shows the reference case without DNA (no non-specific binding).
Figure 5.
Effect of transcript elongation.
(A) Active elongation sequesters core RNAPs for the length of the operon and sigma subunit for some nucleotides. (B) Formation of holoenzymes in the presence of one type of sigma factor without DNA (no specific binding and no transcription with nM, dashed line), in the presence of specific binding (holoenzymes bind to promoter with
M but do not transcribe, case (i)) and in the presence of both specific binding and transcription (case (ii)). The black bars (
) show the case when sigma factor and core unbind as holoenzyme (the binding affinity is described by the equilibrium dissociation constant), the dark blue (
) and the light blue bars (
) when sigma factor separates from core either after promoter unbinding or gene transcription and after 300 nucleotides, respectively (thus, the binding affinity is
). (C) Number of holoenzymes
and
as a function of the copy number of alternative sigma factors in the absence of DNA (case (i)), with transcription of both σ70- and σAlt-dependent genes but with unbinding of sigma factor after 300 nucleotides and core at the end of the operon (case (ii)) and only with the transcription of the σAlt-dependent genes (case (iii)). Values of the parameters are the same as in Figure 5B. (D) Formation of holoenzymes
and
as a function of the copy number of alternative sigma factors without DNA (dashed lines) and transcript elongation (solid lines). (E) Modulation of the effective binding affinities
by sigma factor competition related to the case of Figure 5D. (F) Normalized transcription rate for σ70- and σAlt-dependent promoters as a function of the number of alternative sigma factors, related to the case of Figure 5D (with
nM and
nM).
Figure 6.
(A) During the stringent response RNA polymerases involved in rRNA transcription are quickly released to increase the pool of free cores. (B) Number of holoenzymes and
as a function of the copy number of core RNAPs. The black dashed lines show the number of available RNAPs during the exponential growth state (
) and during the stringent response state. The gray region shows the range of core RNAP for which there is sigma factor competition. (C) Response factor
of the alternative sigma factor-dependent gene transcription (with
M) to an increase of concentration of RNAPs. The blue dashed line shows the maximal sensitivity, that for strong core-sigma binding, is found for
and lies in the competition region. (D) Number of alternative holoenzymes and (E) response factor
related to the σAlt-dependent gene transcription as a function of the number of core RNAPs and alternative sigma factors (with
M). The white line encloses the region of sigma factor competition. The points show possible values of cores and alternative sigma factors for a cell in the exponential growth state and in the stringent state.