plant growth harmones

7/29/2019 plant growth harmones

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The growth hormone auxin

NH

COOH


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Discovery of auxin


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Auxin: the growth hormone

water for 18 hourswater for 18 hours +IAA for 18 hours+IAA for 18 hours


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N

H

COOH


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The roles of auxin in plant growth and development

Control + IAA

Root development

Vascular initiation and patterning

Flower development

Embryogenesis

Tropism


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Auxin appears to be necessary and sufficient for

plant organogenesis

Control + IAA


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Auxin functions are regulated at

multiple levels

1. Auxin homeostasis (biosynthesis, conjugation,and degradation)

2. Auxin polar transport

3. Auxin signal transduction


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Approaches used for dissecting

mechanisms of auxin actions

1. Biochemical approaches: characterization ofauxin inducible genes

AUX/IAA genes

Auxin Response Element (AuxREs)

Plants treated with auxin RNA isolation

Plants treated with water RNA isolation


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Transcription is rapidly induced by auxin

Independent of protein synthesis - primaryresponse

Induced by cycloheximide

Unstable nuclear proteins

Large gene family (29 in Arabidopsis)

The AUX/IAA genes

AUX/IAA proteins probably serve as negative regulators


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degradation proteosome mediated

auxin promotes degradation

Domain II is important for stability

omain structures of AUX/IAA proteins

I II III IV


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The Auxin Response Elements

1. Analysis of the promoter regions of auxin inducible genes

2. Identify the conserved elements: TGTCTC

1. Construct an auxin reporter

2. Identify the transcription factors binding to the AuxREs


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Auxin reporter DR5

Cotyledons

hypocotyl

Root meristem

Auxin response

elements

GFP Gene

DR5-GFP auxin reporter


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Transcription factors bind to AuxREs

Cotyledons

hypocotyl

Root meristem

Use AuxREs as a bait for yeast one hybrid screen

Auxin response factors: 22 members in Arabidopsis


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Approaches used for dissecting

mechanisms of auxin actions

1. Biochemical approaches: characterization of

auxin inducible genesAUX/IAA genes and ARFs (auxin response factors)

2. Genetic approaches: Identification andcharacterization of mutants resistant to exogenous auxin

and auxin polar transport inhibitors.

axrmutants, tirmutants


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Auxin resistance mutant screens

+

auxin

Two types of mutants:

1) Auxin uptake: aux1, axr4

2) Auxin response: axr1, axr2, axr3, axr5, axr6, tir1

-

auxin


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Recessive: aux1, axr1,, axr4, axr5, axr6, msg1, tir1

Dominant: axr2, axr3, msg2, iaa28,

Auxin resistance mutant screens


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axr3-1 axr3-3 Col-0

AXR3/IAA17 VVGWPPVR

axr3-1 L

axr3-3 G

Dominant axr mutants contain mutations

in the domain II in Aux/IAA proteins


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A plausible auxin signaling mechansim

1. Aux / IAA proteins are short lived and function as

negative regulators

2. Mutations in Aux /IAA confer dominant auxinresistance; probably increase the half-life of Aux

/IAA proteins

3. Auxin signaling may depend on the degradation of

Aux / IAA proteins


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Auxin signaling mechanisms

ARF

Aux / IAA

ARF

Aux / IAA

auxin

Removal of Aux /IAA

ARF / ARF Transcription


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Predictions from the model

ARF

Aux / IAA

auxin

Removal of Aux /IAA


1. Inactivation of ARF should cause the same phenotypes as

those induced by stabilized Aux/IAA

2. Inactivation of Aux/IAA should lead to auxin-

overproduction phenotypes

3. Overexpression of ARF may suppress dominant Aux/IAA

mutants


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Arabidopsis pattern mutants

Basal mutant

(mp, bdl)

BDL encodes an Auxin / Indole-3-Acetic Acid protein (IAA12)

MP encodes an auxin response factor (ARF5)


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ARF

Aux / IAA

auxin

Removal of Aux /IAA


How is Aux/IAA degraded?

Ubiquitin-mediated protein degradation machinery?


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An auxin signal transduction pathway

Low [auxin]

High [auxin]


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Previous genetic studies

1) The responses to excess exogenous

auxin

2) Root elongation as a primary physiological

readout

3) Known auxin mutantspin1 andpidare not

auxin resistent


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Polar auxin transport


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Carrier mediated auxin transport

Influx:

a. passive

b. AUX1 permease

Efflux:

Polarity set upby efflux carries

if they are

located only at

the base


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PIN proteins as auxin efflux carriers


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PIN proteins are polarly localized


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Current hypothesis for plant organogenesis

Auxin gradients determine the formation of lateral organs

Auxin peak


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Auxin functions are regulated at multiple

levels

1. Auxin biosynthesis

2. Auxin polar transport

3. Auxin signal transduction


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Auxin biosynthesis

1. Necessary for determining the physiological roles

of auxin

2. Necessary for understanding auxin movementsand dynamics

3. Provides genetic foundations for dissecting the

mechanisms of auxin in plant development


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Proposed tryptophan dependent

auxin biosynthetic pathways

TrpTrpiaaM

Hydrolase

Nitrilase

N

H

NH2

COOH

N

H

NH2

N

H

NH2

O

NH

O

COOH

N

H

H

O

N

H

NOH

N

H

N

N

H

OH

O


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Auxin overproduction yucca1-D mutant

WT yucca1-DWT yucca1-D

Zhao et al. (2001) Science


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YUCCA encodes a flavin-monooxygenase

involved in auxin synthesis

Trp IOX Auxin

NitrilaseNitrilaseYUCCAYUCCA

OOH

NH2

N

H

NH2

N

H

HN

N

H

OH N

N

H

OH

N

H

CN OH

N

H

O

Zhao et al. (2001) Science

Overexpression of YUCCA genes causes


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Overexpression ofYUCCA genes causes

auxin over-production


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Flower defects in yuc1yuc4 double mutants


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Disruption of shoot-root axis by

yuc1yuc4yuc10yuc11

WT

yuc1

yuc4

yuc10

yuc11

C i th


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Can exogenous auxin rescue the yucca

mutant phenotypes?

1. Auxin transport

2. Auxin gradient

3. The right dosage

P d ti f i i it b th b t i l


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Production of auxin in situ by the bacterial

auxin biosynthesis gene iaaM

iaaM iaaHN

H

NH2

COOH

N

H

NH2

O

N

H

OH

O

When expressed in plants, iaaMconverts tryptophan to indole-3-

acetamide, which is hydrolyzed by

non-specific hydrolases in plants


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Complementation ofyuc mutants by expressing the

iaaM gene under the control of aYUCCA promoter

A YUCpromoter iaaM gene

Transform yucmutants

Can the iaaM gene rescue yuc phenotypes?

The c1 c4 do ble m tant is resc ed b the


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The yuc1yuc4 double mutant is rescued by the

iaaM gene under the YUC1 promoter


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Proposed IAA biosynthetic pathway in plants

Sugawara S. et.al. PNAS 2009;106:5430-5435

2009 by National Academy of Sciences

YUC genes have very restricted expression


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YUCgenes have very restricted expression

domains

YUC4 in situ

YUC4 in situ


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Previous genetic studies

1) The responses to excess exogenous

auxin

2) Root elongation as a primary physiological

readout

3) Known auxin mutantspin1 andpidare not

auxin resistent

Genetic screens for yuc1yuc4


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Genetic screens foryuc1yuc4

enhancers

yuc1yuc4

The npy1 (naked pins in yuc )


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The npy1 (naked pins in yuc )

mutant is an yuc1yuc4 enhancer

wt yuc1yuc4 npy1yuc1yuc4 npy1yuc1yuc4

Analogous mechanisms between phototropic


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Analogous mechanisms between phototropic

response and auxin-regulated organogenesis

PID NPY1

ARF5 / MP?

Developmental

signals

PHOT1 NPH3

ARF7 / NPH4?

OrganogenesisPhototropic responses

Blue light

Auxin transport

(PIN3)?

Auxin transport

(PIN1)?

YUCCA

auxin

Formation of pins is a hallmark for defects in


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Formation of pins is a hallmark for defects in

auxin pathways

wt pin1 pid mp

Pin-like maize mutants


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ba1 spi1 Bif1 bif2

BARREN STALK1 (BA1) encodes a bHLH transcription factor

SPARSE INFLORESCECNE1 (SPI1) encodes a YUC-like auxin biosynthesis

enzyme

BARREN INFLORESCENCE2(BIF2) encodes a protein kinase involved in

auxin transport and signaling. One of its targets for phosphorylation is BA1.

Pin like maize mutants

wt

plant growth harmones

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