Recent Advances in Understanding of Senescence in Flower

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Soobedar Yadav Roll. No. 10135 Discipline of Horticulture IARI, New Delhi

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Flower senescence
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Flower senescence
death of flower

is the irreversible, terminal phase of

developmental processes that lead to cellular break down and (Van Doon, 2005)

Why study flower senescence

Understanding senescence mechanism? Extending vase-life of flower by delaying senescence.

Based on ethylene sensitivity flower senescence can be divided into two groups
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1. Ethylene - sensitive flowers : Ethylene Rose Carnation Orchid Petunia

2. Ethylene-insensitive flowers: Abscisic acid (ABA) and Proteases

Alestomeria Gerbera Gladiolus Iris sp. Lilium sp. Daffodils Tulipa sp.

Flower senescence
End of flower opening

or cutting of flower

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High rate of respiration

Vascular occlusion ABA production

Water stress

Increased generation of superoxide radicals (O-2)

Increase in oxidative stress

Decrease polar lipid content Peroxidation of lipid Loss in membrane integrity Flower Senescence

Arora et al, 2005

Theories of Senescence
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1. DNA Damage theories

Accumulated damage to DNA inhibits cell functions and division. Decline in DNA repair capacity -Gene expression inhibited. Damage to mitochondrial DNA Decrease in energy production

2. Built-in Breakdown theories

Genetically programmed process, Senescence associated genes express after a fixed life span of cells. Expression of genes coding for hydrolytic enzymes. Formation of DNA synthesis inhibiting proteins.

3. Telomere Theory
Gradual shortening of Telomeres with each division / Reduction in telomerase activity. Division stops when telomere becomes very short. Replicative cell senescence.

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4. Free radical damage theory

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Damage by highly reactive free radicals like OH, O2, H2O2 , etc.

Disrupts cell and nuclear membranes.

Alters membrane permeability. Damage DNA / Proteins.

Antioxidants reduce free radical damage.

5. Harmonal theory
Ethylene ABA Jasmonate Auxin

Cellular and molecular modes of ethylene action

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Pollination

Stigma

Style

Ovary

Petals

C2H2

Ethylene Receptors

Signal Transduction

Altered Gene Expression

Senescence

Changing focus of flower senescence research......

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Woltering E. J. 2010

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Yang cycle and ethylene biosynthesis pathway

ACC; 1-aminocyclopropane-1-carboxylic acid; GACC; 1-(glutamyl)-ACC; KMBA; 2-keto-4methylthiobytyric acid; MACC; 1-(malonyl)-ACC; MTA; 5'-methylthioadenosine; MTR; 5'methylthioribose; MTR-1-P; 5'-methylthioribose-1-phosphate; SAM; S-adenosylmethionine.

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Overview of added or altered genes in the ethylene biosynthesis pathway

Abbreviations: ACC; 1-aminocyclopropane-1-carboxylic acid; MTA; 5'-methylthioadenosine; SAM; S-adenosylmethionine

Tug of War between polyamines and ethylene

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Pandey et al., 2000

Polyamines , Ethylene and Senescence

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Common

precursor SAM

Competitive

demand for a limited pool

PAs delay

senescence by inhibiting ACC synthesis

Senescence Regulated gene

Senescence down regulated gene-Their Expression decrease
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during senescence i.e. Photosynthetic associated gene Senescence Associated gene(SAGs) Their expression decrease

during senescence
Group A (Class I) Protease

Ribonuclease
Lipase ACC synthase ACC oxidase Group B (Class II) Glutamine synthase (Convert NH+4 to Glutamine)

Programme cell death

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Programme cell death is a process where individual cell activate as intrinsic senescence programme
PCD is the death of individual cell types, and senescence for whole organs

In two floral cell types

Apoptotic- ( Tapetum and Pollen-tube) Autophagous (Petals and Tepal there is stronger evidence)

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Programmed Cell Death in Floral Organs: How and Why do Flowers Die?

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Rogers et al, Annals of Botany (2006) 97: 309315

Programmed cell death begins earlier in the mesophyll cells of tulip petals than in the epidermal cells
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H. Mochizuki-Kawai et al. / Postharvest Biol. and Tech. (2013) 79 :912

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From models to ornamentals: how is flower senescence regulated?

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Rogers H.J. Plant Mol Biol (2013) 82:563574

RNAi in Flower Senescence.

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RNAi is a biological process in which RNA molecules inhibit gene expression, typically by causing the destruction of specific mRNA molecules

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RNA gene silencing studies of flower senescence

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Chen et al (2004). Plant Molecular Biology 55: 521530,

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(Florigene Ltd.)

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Antioxidants Free radical

Action of Antioxidant

Effect of 5-sulfosalicylic acid on antioxidant activity in relation to vase life of Gladiolus cut flowers
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Ezhilmathi K. et al ., Plant Growth Regul (2007) 51:99108

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Total Soluble Protein & Protease Activity

3.5 3.0 (mg g-1 F.W)

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2.5
2.0 1.5 1.0 0.5 0.0 Stage 1 Stage 2 Stage 3 Stage 4 Stage 5 Stage 70.00 60.00 50.00 40.00 30.00 20.00 10.00 0.00

Control 5 SSA

N- Ethylmaleimide
Aprotenin Leupeptin E -64

Protein

( g-1 F W.)

Control
5 SSA N- Ethylmaleimide Aprotenin Leupeptin Stage Stage Stage Stage Stage 1 2 3 4 5 Stage E -64

Protease

Soobedar et al., 2013

Mineral nutrient remobilization during corolla senescence in ethylene-sensitive and -insensitive flowers

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Jones M.L. 2013

Role of Nitric oxide in delaying senescence of cut rose flowers

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12 hr

24 hr

48 hr

W. B. Liao et al. / Scientia Horticulturae 155 (2013) 3038

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2-(4-Carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (CPTIO)

Cyclodextrin-based Nanosponges improve 1-MCP efficacy in extending the postharvest quality of cut flowers

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Anemone coronaria L. Ranunculus asiaticus L. Helianthus annuus L. Rosa hybrida L. Paeonia lactiflora L. Papaver nudicaule L

L. Seglie et al. / Scientia Horticulturae 159 (2013) 162165

Vase life of cut rose cultivars Avalanche and Fiesta as affected by Nano-Silver and S-carvone treatments
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Z. Nazemi Rafi, A. Ramezanian / South African Journal of Bot. 86 (2013) 6872

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CASE STUDY : 1

Materials and Methods

The 3.2 kb fragment etr 1-1
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was cloned from Arabidopsis to

pTA7001 vector This construct was transformed into Agrobacterium tumefaciens strain LBA4404 using electroporation PCR amplification was performed to further confirm the destination vector integrated with etr1-1 gene.

The bacterial culture for inoculation of explants was centrifuged

and was diluted to 1:200 (MS) medium Infected explants were placed on co-cultivation medium at 25uC for

23 days.
Transcripts of Arabidopsis etr1-1 gene was substantially induced in flowers with 30 mM dexamethasone (DEX).

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To determine /compared gene expression changes of petals with DEX to those without DEX at 24 h and 48 h by microarray

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Transcripts of Arabidopsis etr1-1 gene was substantially induced in flowers with 30 mM dexamethasone (DEX)

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After 24 hr. Gene ontology

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After 48 hr. Gene ontology

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Inference..
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The results further confirmed that flower longevity awarded by the etr1-1 gene is related to the level of its expression E7H and E9G lines, which exhibited ethylene insensitivity only when the inducer was applied The vase life of E9G flowers was considerably longer than that of E7H flowers either with or without DEX The vase life of flowers was extended by an average of over 610 days, depending on the level of etr1-1 gene expression.

CASE STUDY : 2
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B.C. In et al. / Postharvest Biology and Technology 86 (2013) 272279

Materials and methods

Carnation plants (Dianthus caryophyllus L.cv. Moonstone)

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The flowers were subse-quently treated singly with

1. 2. 3. 4. Control distilled water 5 L L1ethylene, 100 nL L11-MCP for 6 h Repeated 1-MCP treatments every other day

Ethylene gas measure to gas chromatograph Total RNA was isolated from the receptacle (R), petal claw (PC), and petal blade (PB) by using Trizol method Total RNA was quantified with Nano Drop DN-1000spectrophotometer To determine the expression patterns of ethylene biosynthesis and receptor genes during flower senescence, a RT-PCR analysis of the transcript levels was performed using gene-specific primers Primers was designed for DcACS1,DcACO1, DcETR1, DcERS1 and DcERS2 genes and synthesized by Integrated DNA Technologies (Coralville, IA).

Objective
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Determined the relationship between the mRNA levels of ethylene biosynthesis and ethylene receptor genes Determined the degree of ethylene sensitivity of carnation flowers after ethylene and 1-MCP treatments

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Inference..
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Initial block in ethylene perception with 1-MCP, carnation flowers regain ethylene sensitivity and resume endogenous ethylene biosynthesis over time for single treatment

Successive treatments with 1-MCP completely inhibited ethylene responses

In this study, the mRNA levels of DcETR1 and DcERS1 decreased during flower opening and increased at onset of flower senescence.

Indian Agricultural Research Institute, New Delhi

CASE STUDY : 3

Objectives To determine, first, whether antisense BoACS1 (broccoli ACC synthase) and BoACO1 (broccoli ACC Oxidase) genes would confer reductions in ethylene biosynthesis in transgenic plants To determined the ethylene reductions were associated with delayed flower senescence
Gene transformation
Vector pCS2 or pCS5

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Inference.
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Some of transgenic plants were characterized by reduced

expression

of

endogenous

genes,

reduced

ethylene

biosynthesis, and delayed flower senescence The antisense BoACO1 transformation was evidently more effective than the antisense BoACS1 transformation in reducing ethylene biosynthesis This implies that transformation with antisense ACC Oxidase is more effective in lowering ethylene mediated physiological activities

Road ahead
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Understanding the molecular mechanism of the flower senescence Develop cheap commercial nano formulations incorporating to vase for decrease the senescence

Function, expression, regulation and interaction of the structural genes and regulatory genes

Changing flower senescence by modification of ethylene and polyamine biosynthetic pathway

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