1
2
3
4
5
6
Regulation of transcription occurs via specific interaction of both general and
tissue specific transcription factors (TFs) with promoter and/or enhancer
sequences.
The scheme above shows simplified subsequent formation of the complex of TFs
involved in the regulation of transcription. Interaction of general TFIID with the
TATA box induces distortion of the DNA structure (see the next slide).
7
Induction of structural changes upon interaction of TFIID with DNA. This may be
important for the assembly of other TFs involved in the formation of
transcription initiation complex.
This change of confirmation provides a kind of “signature” that is recognized by
other proteins and NA polymerase to recognize the proper binding site. However,
there are also TATA box‐less promoter, where probably other types of
“signatures” occur.
8
The scheme showing the formation of the transcription initiation complex and
the interaction of both positive (open symbols) and negative (solid symbols)
factors.
These proteins bind to the regulatory sequences that might be hundreds or even
thousands of base pairs away from the promoter. These protein interact with
each other and with the RNA polymerase, integrating thus many signals into a
“yes” or “no” response of the basal promoter, i.e. the region adjacent to the TATA
box and recognized by the RNA polymerase.
The individual positive or negative factors are complex and their activity might be
regulated by their phosphorylation status or via their interaction with other
proteins (i.e. momomeric or dimeric) etc..
9
There is a whole family of transcription activating factors (TAFs) that interact with
signalling molecules, e.g. steroid hormones, thyroid hormones or retinoic acid
and in a response to the signal transfer to the nucleus where they regulate
transcription.
One of the type of TAF are leucine zipper or bZIP type TAFs. These TAFs are
dimeric, with leucine‐rich hydrophobic face formed by the Leu that occurs every
7th aa.
That allows the factor to take the proper configuration, which provides the dimer
with the ability to bind DNA via charged aa.
10
An example of the “microprocessor”‐like acting promoter is a promoter of the endo16
gene from the sea urchin.
There have been identified several gene regulatory modules in the endo16 gene that
have positive or negative regulatory role. These modules were identified via formation
of deletion mutants of the transcriptional fusions with reporter gene.
The analysis has revealed that the module A has a positive function and must interact
with its cognate TAFs for transcription to occur.
Module G enhances the expression when the A and B are active.
C, D, E and F are responsible for the specificity of the expression of endo16 during sea
urchin development.
Each of the modules has several protein interaction sites, some of them general, other
unique. Site for the protein SpGCF1 is present in many modules and is probably
responsible for looping of chromatin, allowing thus bringing of distal regulatory modules
close to the basal promoter.
This type of regulation, i.e. based on the different activities of diverse regulatory
sequences is sometimes called combinatorial and is common for development of many
living creatures.
In the combinatorial type of regulations, some modules may act synergistically, some of
them antagonistically, some may have both positive and negative roles (e.g. the module
B, see the figure). This variability allows very precise and responsive regulations towards
changing environmental conditions.
11
An example of the combinatorial gene regulation is the regulation of β‐globin type of hemoglobin chains
of humans.
As discussed in the Lesson 5 (course Bi8940 Developmental biology), the type of hemoglobin produced by
the fetus changes during development. The hemoglobin present in the liver‐produced hemoglobin is
composed of two α‐ and two β‐type chains. The β‐type hemoglobin chains are of several developmental
types, produced by ε, γ1, γ2 and β (in this order). In addition, there is minor adult type of β‐type
hemoglobin, called δ globin.
The genes for the β‐type chains are aligned on the chromosome in the order, in which they are expressed
during development (see the figure).
For the expression of individual cell types is distinctive an upstream regulatory sequence called locus
control region (LCR). LCR is located about 50 kbp away from the most proximal ε gene.
The LCR structure is different in erytrocyte precursor cells in comparison to other cells that could be
demonstrated by the changes in the sensitivity to low concentrations of DNase, suggesting low amount of
nucleosomes bound.
For the expression of the particular genes, the interaction of their regulatory sequences with LCRs is
necessary. Because of LCR can interact only with one regulatory sequence at a time, only one type of
genes for the particular β‐type chain is activated (the first interaction of LCR with ε gene, which is later in
development replaced by the other one, is shown by the double‐headed arrow).
The underlying molecular mechanisms of the specific pattern of the LCR movement from the most
proximal towards the most distal gene cannot be satisfactory explained.
Probably, acetylation of H3 histones might play a role and possibly, other genes outside of the β‐type
chain family are involved in the regulation of LCR activity. That seems to be confirmed by the
identification of other human genes with similar structure, suggesting common regulatory mechanisms
via LCRs.
For
12
13
14
15
16
17
18
19
20
21
22
23
24
25
Microarray expression profiles of 19 fluorescently sorted GFP-marked
lines were analyzed (3–9, 23, 24). The colors associated with each marker
line reflect the developmental stage and cell types sampled. Thirteen
transverse sections were sampled along the root's longitudinal axis (red
lines) (10). CC, companion cells.
26
(A) The majority of enriched GO terms (hierarchically clustered) are associated
with individual cell types (blue bar). A smaller number are present across multiple
cell types (green bar). (fig. S2) (B) GO category enrichment for hair cells
confirms a previous report (15). Enriched cis-elements and an enriched TF family
were also identified. (C) From the top 50% of varying probe sets, 51 dominant
radial patterns were identified. Pattern expression values were mean-normalized
(rows) and log2 transformed to yield relative expression indices for each marker
line (columns). Marker line order is the same for all figures; see table S1 for
marker line abbreviations. (D) Pattern expression peaks were found across one
to five cell types. (E to G) Patterns where expression is enriched in single and
multiple cell types support transcriptional regulation of auxin flux and synthesis. In
all heat maps with probe sets, expression values were mean-normalized and log2
transformed. Expression is false-colored in representations of a root transverse
section, a cut-away of a root tip, and in a lateral root primordium. (E) Auxin
biosynthetic genes (CYP79B2, CYP79B3, SUPERROOT1, and SUPERROOT2)
are transcriptionally enriched in the QC, lateral root primordia, pericycle, and
phloem-pole pericycle (P = 1.99E–11, pattern 5). All AGI identifiers and TAIR
descriptions are found in table S14. (F) Auxin amido-synthases GH3.6 and
GH3.17 that play a role in auxin homeostasis show enriched expression in the
columella, just below the predicted auxin biosynthetic center of the QC (P =
8.82E–4, pattern 13). (G) The expression of the auxin transporter, PINFORMED2,
and auxin transport regulators (PINOID, WAG1) are enriched in the
columella, hair cells, and cortex (P = 1.03E–4, pattern 31).
27
(A) The majority of enriched GO terms (hierarchically clustered) are associated
with individual cell types (blue bar). A smaller number are present across multiple
cell types (green bar). (fig. S2) (B) GO category enrichment for hair cells
confirms a previous report (15). Enriched cis-elements and an enriched TF family
were also identified. (C) From the top 50% of varying probe sets, 51 dominant
radial patterns were identified. Pattern expression values were mean-normalized
(rows) and log2 transformed to yield relative expression indices for each marker
line (columns). Marker line order is the same for all figures; see table S1 for
marker line abbreviations. (D) Pattern expression peaks were found across one
to five cell types. (E to G) Patterns where expression is enriched in single and
multiple cell types support transcriptional regulation of auxin flux and synthesis. In
all heat maps with probe sets, expression values were mean-normalized and log2
transformed. Expression is false-colored in representations of a root transverse
section, a cut-away of a root tip, and in a lateral root primordium. (E) Auxin
biosynthetic genes (CYP79B2, CYP79B3, SUPERROOT1, and SUPERROOT2)
are transcriptionally enriched in the QC, lateral root primordia, pericycle, and
phloem-pole pericycle (P = 1.99E–11, pattern 5). All AGI identifiers and TAIR
descriptions are found in table S14. (F) Auxin amido-synthases GH3.6 and
GH3.17 that play a role in auxin homeostasis show enriched expression in the
columella, just below the predicted auxin biosynthetic center of the QC (P =
8.82E–4, pattern 13). (G) The expression of the auxin transporter, PINFORMED2,
and auxin transport regulators (PINOID, WAG1) are enriched in the
columella, hair cells, and cortex (P = 1.03E–4, pattern 31).
28
Schematic flowchart of the Human Protein Atlas. For each gene, a signature
sequence (PrEST) is defined from the human genome sequence, and following
RT‐PCR, cloning and production of recombinant protein fragments, subsequent
immunization and affinity purification of antisera results inmunospecific
antibodies. The produced antibodies are tested and validated in various
immunoassays. Approved antibodies are used for protein profiling in cells
(immunofluorescence) and tissues (immunohistochemistry) to generate the
images and protein expression data that are presented in the Human Protein
Atlas (Ponten et al., J Int Med, 2011).
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
Excample of an output of transcriptional profiling study using Illumina 
sequencing performed in our lab. Shown is just a tiny fragment of the complete 
list, copmprising about 7K genes revealing differential expression in the studied 
mutant. 
45
46
47