Auxin metabolism and signaling

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Auxin metabolism and signaling
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Introduction
Plant physiology (Taiz Zeiger) ch.19 p425
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A kinds of auxin and structure
Natural auxin
Synthetic auxin
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Functions of Auxin

• Stimulates cell elongation
• Stimulates cell division in the cambium and,
  in combination with cytokinins in tissue culture
• Stimulates differentiation of phloem and xylem
  
• Stimulates root initiation on stem cuttings
  and lateral root development in tissue culture
• Mediates the tropistic response of bending in
  response to gravity and light
• The auxin supply from the apical bud
  suppresses growth of lateral buds
• Delays leaf senescence
• Can inhibit or promote (via ethylene
  stimulation) leaf and fruit abscission
• Can induce fruit setting and growth in some
  plants
• Delays fruit ripening
• Promotes flowering in Bromeliads
• Stimulates growth of flower parts
• Promotes (via ethylene production) femaleness
  in dioecious flowers
• Stimulates the production of ethylene at high
  concentrations

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Auxin metabolism
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IAA biosynthesis
(IGP)
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Tryptophan-dependent pathway
Plant physiology (Taiz Zeiger) ch.19 p428
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• YUCCA
• flavin monooxygenase
• cytochrome 450
• CYP79B2,CYP79B3,
• CYP83B1

Current Opinion in Plant Biology 2001, 4234240
It appears likely that several pathways are
present, perhaps in different cells,
developmental stages, or as regulated by
environmental inputs.
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Tryptophan-independent pathway
orp mutants completely lack tryptophan synthase
ß-activity due to mutations two unlinked loci,
orp1 and orp2
The Plant Cell, Vol. 4, 711-719, June 1992
When grown on media containing 302H2O or
15Nanthranilate,
tryptophan IAA
WT o o
mutant x o
In orp seedling, de novo IAA biosynthesis occurs
without tryptophan as precursor.
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All three mutants were synthesizing IAA de novo.
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Plant physiology (Taiz Zeiger) ch19. p430
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IAA conjugates in plant

• IAA has been found to be conjugated to simple
  sugars, cyclitols, high molecular weight
  polysaccharides.
• The carbohydrate component of glycoproteins via
  an ester or anhydride linkage.
• IAA can also be conjugated by amide linkage to
  single amino acids, peptides or proteins.

Plant physiology (Taiz Zeiger) ch19. p431
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• Conjugates are thought to be involved in a
  variety of hormonally related processes such as
• Transport of IAA within the plant,
• The storage and subsequent reuse of IAA
• Protection of IAA from enzymatic destruction
• As components of a homeostatic mechanism for the
  control of IAA levels
• As an entry route into the subsequent catabolism
  of IAA

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IAA degradation
Plant physiology (Taiz Zeiger) ch19. p431
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Auxin Signaling
Journal of Cell Science 2006 (119, pp. 1199-1202)
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1. Auxin-responsive genes 1) Small Auxin-Up
RNAs(SAURs) - SAURs were initially
identified in soybean as rapidly accumulating
transcripts following a short auxin
treatment - SAURs genes have been
idintified in many other plants, including 77
predicted SAURs in Arabidopsis
genome. - Several SAUR transcripts are
highly unstable due to a conserved
downstream element (DST) found in the
3untranslated region.
THE JOURNAL OF BIOLOGICAL CHEMISTRY Vol. 278, No.
26, Issue of June 27, pp. 2393623943, 2003
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Tissue-specific expression of zmSAUR2
THE JOURNAL OF BIOLOGICAL CHEMISTRY Vol. 278, No.
26, Issue of June 27, pp. 2393623943, 2003
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1. Auxin-responsive genes 2) GH3 gene
- Arabidopsis contains 19 GH3 genes, several of
which are auxin inducible. - some GH3
family members encodes a novel class of
acyl-adenylate-forming enzyme
capable of acting as IAA-amino synthases in the
conjugation of IAA to amino acids.
ex) over expressing GH3.6 exhibit resistance
to applied IAA, while seedlings containing
mutation in GH3.5 or GH3.17 exhibit increased
sensitivity.
The Plant Cell, Vol. 17, 616627
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1. Auxin-responsive genes 3)
AUXIN/INDOLE-3-ACETIC ACID(Aux/IAA) -
Third large class of auxin inducible
transcripts. - In Arabidopsis contains
at 29-member gene family - Aux/IAA
induction occurs within minutes of auxin
treatment and do not require new
protein synthesis. - encode 2035KDa
proteins, localized to the nucleus and negatively
acting transcription factor (repress
auxin response) - Aux/IAA protein share
four highly conserved motif termed domain
I,II,III,IV
- Domain I putative transcriptional
repression motif - Domain II
constitutes a degron that
targets the Aux/IAA-protein for
degradation by the ubiquitin-26S
proteasome proteolytic pathway -
Domain III and IV dimerization with other
Aux/IAA-protein and with Auxin
Response Factor(ARFs)
Developmental Cell, Vol. 1, 595604, November,
2001
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2. Auxin response factors 1) ARF-mediated
transcriptional regulation - ARF protein
share conserved N-terminal DNA binding domain
that mediate binding to the AuxRE
sequence(TGTCTC) - middle region (MR) of
ARF highly divergent (serine-rich,
glutamine-rich) ?
serine-rich ARFs repress auxin responsive gene
exrpession ? glutamine-rich ARFs
act as transcriptional activator
Developmental Cell, Vol. 1, 595604, November,
2001
The Plant Cell, Vol. 15, 533543
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Middle region (MR) of ARF the loss of function
mutations in Aux/IAA 1) serine-rich ARFs ?
ETTIN/ARF3 lack the C-terminal dimerization
motif, gynoecium patterning defect
in flowers by auxin transport
inhibitor NPA ? HOOKLESS SUPPRESSOR
1(HSS1/ARF2)-ethylene and light regulate
protein level of this
serine-rich ARF to mediate differential cell
elongation in the hypocotyl ? arf2 mutant
several phenotype, increased size of several
organs, agravitropic stem,
delayed senescence and abscission.
? arf10/arf16 double mutant exhibit
agravitropic root growth, due to aberrant root
cap
cell differentiation 2) glutamine-rich
ARFs ? MONOPTEROUS (MP/ARF5) defect in
embryo patterining with severe allels lacking
all
basal structure ?
NON-PHOTOTROPIC HYPOCOTYL 4(NPH4/ARF7)
diminished tropic responses in the
shoot
due to reduced auxin-mediated cell expansion
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ltarf2 mutant (serine-rich ARFs)gt several
phenotype, increased size of several organs,
agravitropic stem, delayed senescence and
abscission. The Plant Journal (2005) 43,
2946
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2. Auxin response factors 2) Domain of ARF
protein - ARF protein contain a
C-terminal dimerization domain highly related to
motif III and IV of Aux/IAA protein.
- C-term mediate homo and heterodimer
formation between ARF protein and
heterodimerization with Aux/IAA
protein - ARF-ARF dimerization
facilitate the binding of these transcription
factor to
their target site, ARF dimerization
is not essential for
AuxRE binding.

The Plant Cell, S81S95, Supplement 2002
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2. Auxin response factors 3) Domain of ARF
protein
Model for Aux/IAA Action
Aux/IAAs form a variety of dimers with both ARFs
and other Aux/IAAs. The equilibrium of these
dimerization events has the potential to alter
gene expression from TGTCNC-containing AuxREs in
auxin responsive promoters. Auxin regulates
both the degradation of Aux/IAA proteins and the
transcription of Aux/IAA genes. Given that the
diverse patterns of degradation and expression
with the large Aux/IAA family would lead to
considerable temporal variation in the relative
abundance of different Aux/IAAs, the interactions
are likely to be very complex.
The Plant Cell, S81S95, Supplement 2002
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3. Regulation of auxin response by SCFTIR1
ubiquitin-ligase - Ubiquitination model. E3
enzyme or Ubiquitin-ligase, catalyze conjugation
of ubiquition to substrate protein.
onece ubiquitinylated, these substrates are
targeted for proteolysis by the 26S proteasome.
1) SCF-type ubiquitin-ligase - multi-subunit
enzyme skp1, cullin1,F-box protein, small
ring domain protein (RBX1, ROC1, HRT1) -
CUL1 scaffold protein, binding RBX1(its C-
terminal domain) and SKP1(its N-terminal
domain) - F-box protein bind to
heter-trimeric core, interaction between
their F-box domain and SKP1 2) TIR1 700 F-box
protein encoded in the Arabidops - mutation in
TIR1 auxin-response defects including
auxin-resistant root growth and reduced
later root envelopment 3) SCF subunit mutation
do not effect auxin signal, but pleiotropic
phenotype ? SCFTIR1 ubiquitin-ligase complex
positive regulator auxin response
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3. Regulation of auxin response by SCFTIR1
ubiquitin-ligase - Reduction in CUL1 levels
results in severe shoot meristem defects
The EMBO Journal Vol. 22 No. 13 pp. 3314, 2003
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Regulation of auxin response by the SCFTIR1
ubiquitin-ligase
a) Aux/IAA-proteins are stable and dimerize with
ARF transcriptional activators, repressing
the activation of auxin-inducible
genes. b) Upon an auxin stimulus - the TIR1
F-protein recruits the Aux/IAA repressors to
the SCFTIR1 complex - Aux/IAA protein
degradation by 26S proteasome - decline of
Aux/IAA level - derepresses the Q-rich ARF,
activate auxin-inducible gene ?
Synthesis of nascent Aux/IAA-protein
negative feedback loop
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4. Regulation of SCFTIR1 ubiquitin-ligase
( A model for SCFTIR1 regulation by the cyclic
RUB modification of CUL1)
The CSN complex can cleave the RUB modifier
from CUL1, thus facilitating CAND1 binding to
CUL1 and SCF disassembly. Conjugation of
RUB to CUL1 by the AXR1-ECR1 and RCE1 enzymes
frees CUL1 from CAND1, promoting re-assembly of
the active complex. Genetic and biochemical
studies have shown that all of the components
depicted in this figure are required for optimal
SCFTIR1 activity in vivo
CSN COP9 signalosome ECR1 E1 Like Conjugating
enzyme Related 1 RUB Related to Ubiquitin RCE1
RUB1 Conjugating Enzyme 1 CAND1 Cullin
Associated and Nedd8 Dissociated 1 AXR Auxin
Resistant
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5. Identification of an auxin receptor
Classical studies provide support for both
plasma membrane and intercellular sites of auxin
perception 1) ABP1(Auxin Binding Protein1)
binds biologically active auxin with high
affinity consistent with the presence of a
C-terminal KDEL endoplasmic retention motif, the
majority of ABP1 is ER localized. A small
fraction escapes to the cell surface, and
several lines of evidence implicate plasma
membrane associated ABP1 in early auxin
mediated electrophysiologic responses.

• - Arabidopsis abp1 null mutants
• early embryo-lethal phenotype, indicating that
  ABP1 is an essential gene
• However, the molecular activity of ABP1 remains
  undetermined, and it does not
• appear to be involved in the SCFTIR1-mediated
  signaling pathway

Plant Molecular Biology 49 339348, 2002.
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6. Conclusions and future perspective