ANJANI KUMAR [Ph.D. Scholar] GENETICS AND PLANT BREEDING BAU, RANCHI

1. Transcription factors as key regulators
of gene expression
M.Sc. Master Seminar
Anjani Kumar
(BAC/M/PBG/003/15-16)
Department of Plant Breeding and Genetics,
Bihar Agricultural College,
BAU, Sabour

2. What is a Transcription Factor (TF)?
Any protein other than RNA Polymerase that
is required for transcription
These are of two types:
General transcription factors Required for the binding of the
RNA pol to the core promoter and its progression to the
elongation stage Are necessary for basal transcription
Regulatory transcription factors Serve to regulate the rate of
transcription of nearby genes They influence the ability of
RNA pol to begin transcription of a particular gene. These
factors recognize cis-regulatory elements located near the
core promoter.

3. The RNA-transcribing enzyme, RNA polymerase II (red), requires general transcription factors (TFII) D, A, B, F, E, and H
(blue), which themselves consist of multiple subunits, to recognize the transcription start site via the TATA box or related
sequences in the core promoter. The sum of these factors, known as the pre-initiation complex (PIC), is required for basal
transcription.
Transcription factors (green) bind to specific DNA sequences (red) via their DNA-binding domain (DBD) and modulate the
rate of transcription via their transactivation domain(s) (TAD).
[Source: http://cro.sagepub.com/content/15/5/282/F1.expansion.html]
A simplistic view of regulatory mechanisms of gene
transcription
DNA binding domain – DBD
Binds specific sequence of
base pairs
Transcriptional activation
domain – TAD
Interacts with basal TF
directly with RNA pol II
Protein-protein interaction
domain – PPID
Interaction with other
transcription factors

4. Source: PlantTFDB @ CBI, PKU ,10/25/2016
Major TF family and genes reported in plants….

5. Source: http://voices.nationalgeographic.com/2009/03/23/corn_domesticated_8700_years_ago/
Teosinte branched1
(Tb1) transcription
factor in
domestication of
maize
When Tb1 is expressed, it represses the outgrowth of lateral branches; maize plants carrying loss of function
alleles produce numerous lateral branches (tillers). During the domestication of teosinte to produce maize,
an allele was selected that altered the regulation of Tb1, increasing its expression in primary auxiliary
meristems (Doebley et al., 1997)
Why these TFs are so important....???

6. The semidwarf varieties of wheat are short
due to a mutation in at least one of two
Reduced height-1 loci (Rht-B1 and Rht-D1]
[Karen Century et al., 2008]
Peng et al. (1999) elegantly demonstrated
that Rht-B1 and Rht-D1 were orthologs of
the Arabidopsis GIBBERELLIN INSENSITIVE
(GAI) gene, a member of the GRAS family
of TFs, which function as transcriptional
repressors of growth.
Major yield gains achieved by conventional plant breeders,
ALSO had role of TFs
Source: http://www.newhallmill.org.uk/wht-rht.htm
Fig. The picture shows the wheat variety "Mercia" with a control and
four lines with different Rht genes.
Rht-0, with no reduced-height gene, is used as the control for height
comparison.
Rht-1 and Rht-2 typically produce semi-dwarf plants, two-thirds the
height of the control.
Rht-3 and Rht-12 typically produce dwarf plants, one-third the height of
the control.

7. • Two TFs have been identified as playing a major role in reducing grain
shattering in domesticated rice plants.
• One of these was isolated as a quantitative trait locus (QTL) in a cross
between a shattering-type ‘Indica’ cultivar and a non-shattering type
‘Japonica’ (Konishi et al., 2006).
• This gene, qSH1, encodes a BEL1-type homeodomain protein that is
orthologous to Arabidopsis REPLUMLESS (RPL), which is involved in the
formation of an abscission zone in the Arabidopsis silique.
• The other TF affecting this trait is shattering4 (sh4), allelic to sha1 (Li et al.,
2006; Lin et al., 2007). SH4 is a member of the trihelix family of plant-specific
TFs and was isolated as a major QTL for shattering in a cross between O.
sativa and Oryza rufipogon.
Role of TF in Rice shattering

8. Review of literature suggests that TFs plays different regulatory role in
plants (stress tolerance, defense, metabolite biosynthesis etc.
T.F spp. Function Reference
AtMYB096 Arabidopsis thaliana Drought tolerance (ABA and
JA–mediated)
Seo et al. 2009
OsMYB55 Oryza sativa Heat stress tolerance El-kereamy et al.
2012
ScMYBAS1 Saccharum officinarum Drought and salt tolerance Prabu and Theertha
2011, Prabu and
Prasad 2012
AtMYB011/AtMYB
012/ AtMYB111
Arabidopsis thaliana Phenylpropanoid pathway/
Flavonol biosynthesis
Stracke et al. 2007
AtMYB44 Arabidopsis thaliana Plant defense response
against aphid
Liu et al. 2010
AtMYB15 Arabidopsis thaliana cold stress tolerance Agarwal et al. 2006
GmMYB Glycine max salt, drought and/or cold
stress
Liao et al. 2008

9. T.F spp. Function Reference
WRKY57
OsWRKY11
OsWRKY13
A . thaliana
Oryza sativa
Oryza sativa
drought tolerance
drought and heat tolerance
bacterial blight Xanthomonas oryzae pv
oryzae and the fungal blast
Magnaportha grisea
Lindemose,Søren
et al.2013
Qiu et al., 2007,
2008
NTL6, PR1, PR2, PR5
Arabidopsis thaliana
Positiveregulatorofpathogenresistance
against P. syringae
Seo et al., 2010
DREB Oryza sativa drought & salt Dubouzet et al.
2003
CBF Brassica napus, Triticum
aestivum Lycopersicon
Esculentum
tolerance to freezing and
drought/low-temperature exposure
Jaglo et al., 2001).
WD40 Zea mays (maize) Regulation of anthocyanin pathway in
seed aleurone and scutellum
Carey et al. (2004)
WD40 Vitis vinifera (grape) Contributes to the accumulation of
anthocyanins
Matus et al. (2010)
OsNAC6, Oryza sativa Slightly increased tolerance to rice blast
disease
Nakashima et al.,
2007
bZIP A . thaliana Drought,salt,cold Choi et al.2000
AP2/EREBP Arabidopsis and rice drought tolerance Karaba et al., 2007
Continue….

10. How these TFs works in plants….
(Front. Plant Sci., 09 February 2016 | http://dx.doi.org/10.3389/fpls.2016.00067)

11. (Int. J. Mol. Sci. 2013, 14(4), 7515-7541; doi:10.3390/ijms14047515)

12. Regulation of Flavonoid biosynthesis by TFs
(Shutian Li et al. 2014)
Figure . The biosynthetic pathway for flavonols, anthocyanins, and PAs in Arabidopsis. This pathway starts with the general phenylpropanoid metabolism
and subsequent steps are catalyzed by a series of structural enzymes leading to the biosynthesis of 3 final end products, including flavonols,
anthocyanins, and PAs. The early biosynthetic genes (EBGs) are activated by 3 functionally redundant R2R3-MYB proteins (MYB11, MYB12, and
MYB111), whereas the expression of the late biosynthetic genes (LBGs) requires the transcriptional activation activity of the R2R3-MYB/bHLH/WD40
(MBW) complex. Enzymes are denoted in uppercase and corresponding genetic loci are indicated in italic lowercase letters. PAL, phenylalanine
ammonia-lyase; C4H, cinnamate 4-hydroxylase; 4CL, 4-coumarate:CoA ligase; CHS, chalcone synthase; CHI, chalcone isomerase; F3H, flavanone 3-
hydroxylase; F3′H, flavanone 3′-hydroxylase; DFR, dihydroflavonol 4-reductase; LDOX, leucoanthocyanidin dioxygenase; ANR, anthocyanidin reductase;
tt, transparent testa; ban, banyuls.
TFs
TFs

13. Fig. Phenotype of strawberry plants transformed with 35S:FvMYB10,FvMYB10 RNAi, and wild-type controls. Growth
and pigmentation of strawberry plants transformed with 35S:FvMYB10, FvMYB10 RNAi, and wild-type controls (A). Detailed phenotype of
strawberry leaves,flowers and fruit of 35S:FvMYB10,FvMYB10 RNAi, and wild-typecontrols.Linesof35S:FvMYB10 had pigmented leaves,
petioles, stigmas and petals, and mature fruit had darkred/purple skin and red flesh. The mature fruit of FvMYB10 RNAi lines had white
skin and white flesh, and the only pigmented tissue was the petioles (B).
(Kui Lin-Wang et al.2014 )
MYB
FOR ENHANCED
PIGMENTATION
Application
of TFs

14. Fig. Freezing tolerance of cbf2 mutant plants. Three-week-old WT and cbf2 plants grown under long-day
photoperiods at 20°C were exposed to different freezing temperatures for 6 h. Freezing tolerance was
estimated as the percentage of plants surviving each specific temperature after 7 days of recovery under
unstressed conditions. (A) Tolerance of nonacclimated plants. (B) Representative nonacclimated WT
and cbf2 plants 7 days after being exposed to -6°C for 6 h. (C) Tolerance of cold-acclimated (7 days at
4°C) plants. (D) Representative cold-acclimated WT and cbf2 plants 7 days after being exposed to -10°C
for 6 h. In A and C, data are expressed as means of three independent experiments with 50 plants each.
Bars indicate SE Fernando Novillo et al.2004,
CBF for freezing
tolerance

15. Figure 4. Stress-tolerance assays of AaDREB1 overexpressing transgenic Arabidopsis. Ten-day-old
seedlings of AaDREB1 transgenic lines (T17, T122, T196) and the empty vector control plants (WT) were
treated with either 100 mM NaCl for 12 days, 10% PEG for 10 days, or exposed to 4 °C for 20 h, then
grown under normal growth conditions for 3 days. Scale bar represents 1.5 cm
(Zong et al.2016)
DREB for
salt,
drought and
cold

16. Overexpression of AtDREB1 Improved Salt Tolerance of Transgenic Rice
Plants
Fig. Salt tolerance analyses of wild type plants and‑ ‑ AtDREB1 transgenic rice. (A) The phenotype of
two week old‑ ‑ AtDREB1 transgenic lines and wild type rice plants grown in the glasshouse under normal‑
growth conditions before being transferred to 150 mM NaCl for 16 days.
Int. J. Mol. Sci. 2016, 17(4), 611; doi:10.3390/ijms17040611
(hp://dx.doi.org/10.3390/ijms17040611
Jun Mei Zong et al.2016‑

17. Improved drought resistance of SNAC1-overexpressing transgenic rice at reproductive stage. (a)
Overexpression contruct (Upper) and RNA gel blot analysis of SNAC1 in transgenic plants and
the WT (Lower). (b) Southern blot analysis of transgenic plants using hygromycin resistance
gene as a probe. (c) Appearance of one positive (S19) and one negative (S18) transgenic
families in the field with severe drought stress. (d) Cosegregation of SNAC1-overexpressing
(RNA gel blot analysis) with the improved drought tolerance in the T1 family of S19. SS(%), seed-
setting rate
Hu et al. PNAS 2006;103:12987-12992
Improved drought resistance of SNAC1-overexpressing transgenic rice at reproductive stage.
NAC for drought
resistance

18. Prospect of Application of TF in Crop Improvement
1. IN CROP DOMESTICATION AND BREEDING
Because of their nature as master switches for major regulatory networks and their prior
role in the domestication of many crop species, TFs are predicted to be among the
best and safest candidate loci for engineering these traits.
2. ENHANCING NUTRIENT USE EFFICIENCY
An example of the successful engineering of enhanced nitrogen uptake using a TF was
reported by Yanagisawa et al. (2004), the overexpression of maize Dof1 gene, which was
known to be involved in organic acid metabolism, to create transgenic Arabidopsis plants
that showed increases in free amino acid content and total nitrogen uptake, as well as
improved growth under low nitrogen conditions.
3. IMPROVEMENT IN YIELD POTENTIAL
There are a number of approaches that might be taken to boost intrinsic yield,
including increasing photosynthetic capacity, modifying plant architecture,
controlling disease and pest, and enhancing the plant’s rate of growth

19. •Though, TFs are a useful candidate for qualitative and quantitative
trait improvement in plants.
•TF technologies often require optimization, either to reduce
unwanted side effects such as growth retardation or to enhance the
desired trait to the level at which it is of commercial value.
•Optimization require a tissue-specific, developmental, or inducible
promoters rather than the usual constitutive promoters, to limit
expression of the transgene to the appropriate tissues or
environmental condition
•Lengthy and costly process of developing a new commercial
genetically MODIFIED CROP
•Securing approvals from regulatory authorities
Challenges while using TFs in agricultural product
development

20. Conclusion
• This is well proven that TFs acts as master switches for
major regulatory networks.
• They regulate the co-ordinated expression of several genes
in a multi-genic pathways, thereby a single TF may be
sufficient for engineering the target traits.
• Taken together, various roles played by TFs in plants, these
could be potential source of candidate genes for modifying
complex traits in crops.
• TF-based strategies appear to be very promising and could
play a major role in development of genetically engineered
crops.

21. References
Konishi S, Izawa T, Lin SY, Ebana K, Fukuta Y, Sasaki T, Yano M (2006). An SNP caused loss of see
shattering during rice domestication. Science 312: 1392–1396
Doebley J, Stec A, Hubbard L (1997) The evolution of apical dominance in
maize. Nature 386: 485–488
Peng J, Richards DE, Hartley NM, Murphy GP, Devos KM, Flintham JE, Beales J, Fish LJ,
Worland AJ, Pelica F, et al (1999) ‘Green revolution’ genes encode mutant gibberellin response
modulators. Nature 400: 256–261
Lin Z, Griffith ME, Li X, Zhu Z, Tan L, Fu Y, Zhang W, Wang X, Xie D, Sun C (2007) Origin of
seed shattering in rice (Oryza sativa L.). Planta 226: 11–20
Li C, Zhou A, Sang T (2006) Rice domestication by reducing shattering. Science 311: 1936–
1939
Shutian Li (2013) Transcriptional control of flavonoid biosynthesis. 10.4161/psb.27522
KuiLin-Wang1, TonyK.McGhie2, MindyWang1, YuhuiLiu3, BenjaminWarren1, RoyStorey4,
RichardV.Espley1 and AndrewC.Allan1,5 (2014) Engineering the anthocyanin regulatory
complex of strawberry (Fragaria vesca) Plant science doi: 10.3389/fpls.2014.00651

22. KuiLin-Wang1, TonyK.McGhie2, MindyWang1, YuhuiLiu3, BenjaminWarren1, RoyStorey4,
RichardV.Espley1 and AndrewC.Allan1,5 (2014) Engineering the anthocyanin regulatory
complex of strawberry (Fragaria vesca) Plant science doi: 10.3389/fpls.2014.00651
Fernando Novillo, Jose´ M. Alonso, Joseph R. Ecker, and Julio Salinas (2003) CBF2DREB1C is
a negative regulator of CBF1DREB1B and CBF3DREB1A expression and plays a central role in
stress tolerance in Arabidopsis. Pnas doi10.1073pnas.0303029101
Honghong Hu, Mingqiu Dai, Jialing Yao, Benze Xiao, Xianghua Li , Qifa Zhang, and Lizhong
Xiong (2006) Overexpressing a NAM, ATAF, and CUC (NAC) transcription factor enhances
drought resistance and salt tolerance in rice. pnas.0604882103
Jun Mei Zong , Xiao Wei Li , Yuan Hang Zhou, Fa Wei Wang, Nan Wang, Yuan Yuan Dong,‑ ‑ ‑ ‑ ‑
Yan Xi Yuan, Huan Chen, Xiu Ming Liu, Na Yao and Hai Yan Li (2016) The‑ ‑ ‑ AaDREB1
Transcription Factor from the Cold Tolerant Plant‑ Adonis amurensis Enhances Abiotic Stress
Tolerance in Transgenic Plant. Int. J. Mol. Sci. 17(4), 611; doi:10.3390/ijms17040611
Yanagisawa S, Akiyama A, Kisaka H, Uchimiya H, Miwa T (2004) Metabolic engineering with
Dof1 transcription factor in plants: improved nitrogen assimilation and growth under low
nitrogen conditions. Proc Natl Acad Sci USA 101: 7833–7838

23. Thanks