Aspects of the hormonal physiology of fruit development in Vitis vinifera L

ASIEíJTS 0F TtrE EORtiOlï/iI, PEYSIOLOGT 0F FRIIIT DEVEIOP¡IET{T

I]f

UITTS VTII-ITER.ê, ].,.

 thesis subnitteå

by

John tr* Consid.ine B,îg:r.Sci.(Melb), IÐ4.

f0 the Ihiversity of lklelaide for the d.egree of

MÀSTSR OXI AGRICI]TTTIRÂL SCIEIVCE

Dept. of Plant Pþsiology,

University of .A,ôelaide 1969

WAITE (ì. INSTITUTE

rt.-to

LIBRARY

.r{..-l

(nHtr

oF

l_ r_ t_

Page T.ABLE OF ^CO}ITENTS

List of figures List of tables l,ist of plates

Summary

Statement

I. General- Introd.uction

(¡)

(+)

II, ^{ô ju}

v

ix

x xi.

](Y

1

(r ₎ 1

(z)

Structr:re and development of the fruit

lhe relationship between plant grouth regulators

and- fruit set and ùevelopment

(i) Gibbereltins

(ii ) ^Growth retard.ants

the relationship between end-ogenous grov'rth regulators

and. fruit set and- d.evelopment

(i) End.ogenous gibberellin-like compounds

(:-i) End.ogenous abscisic acid--l-ike ^conpounds The relationship between the grora.th retardant

CCC and- end-ogenous gibberellin-like and abscisic acid.-like compounds

t

int

t

^on

a:ld. CCC on fruit clrrster clevelonment

+,4

o

9

(r ₎ (z) (¡)

(+)

(r)

fntrod-uction

Materials and- methods Resul-ts

(i ) ^Dorad-i11o (ii) Muscat

(iii _{) sul-tana} (iv) Cape Cunant (-.) Zani,e Gurrant Discussion

Conclusions

10

11

12 12 13 16 16 18

21

22 25 53

^a

l-v Table of Contents (contínued.)

IfI. Gibberellin-like and. abscisic acid--like ^ggmpegn(þ

i n rrnqne nr¡¡ni os and. ¡nthoz.a orÀ tho affon* nf *ho

aDDlice,tion of ^CCC

Page

64 6+

65 75 B6

92

103 104 120

125

138

146

153

(r ₎ (z) (¡)

(q)

Introduction

Materials and. method.s

Results

Discussion and conclusions

vr.

ïV.

General Discussion

V.

Acknowled.Er¡ents

.Appendlces

A.

Experiments on extraction and. assay techniques

3.

Colwnn chromatography stud.ies

C.

Experiments related_

to

the gas liquid_

chronatography

of

plant hormonos

D.

Extr¡eriments ¡elated.

to

the d_etection

of

growüh substances on chromatogra,ms

E.

Anatomical stuclies

of

ornrle d.evelopment in

Cape Cr:nant

frI. Siblioeralhv

LTST ^CF" FIGURES

Figures from section II on the intoraction of ^GÀ and- tCC on

reprocluctive fruit cluster cle-,relopment

1¡

Pagc 2T 28 20 2A

3O

31 31 32

Figurc 2.1 tr'igi.me 2.2

Figure 2"1 Figr-lre 2.

{

Figure 2,!

Fi6ure 2.6 tr'igure

2.I

^rr

Muscat 1966-7

Ca.pe

1,a66-7

Cape

1966-7

Zante

1966-7

DoraC'i1l-o 1967-8

-

^{ìtumber of}

It

seeded berri-es ^per cluster - ^Number of seeclLess berries per

cluster.

- ^{Total number} of berries pel cfuster

lt It

3O

X'igure Figure Figure

Figure ï'igure Figure Figtr-re Fi6ure Figu-re Figure Figure

2" 16 2"17 2.18 2"19 2"20 2"21 2"2?

2.23

^Tigure 2.11 ^rr

Figu-re 2. 12 ^tr Figure 2"13 iluscat Figurc 2. 14 ^It Figì;re 2. 15 ^u

- rt seeùle¡ss Jrerries

- ^{ï'resh weigi:t} of seeded berries - Length/wiatn ratio of seedeù

berries 32

- ^Rachis length. 33

- CLustarroight ³³

- ^j'ú-unber of seocled- berries ¡er

cluster' 3:l-

- l'Tutr-ucl of seecll-ess berries per

oluster ^31- - ^{Total numT:e::} of berries ^per

cl-uster ³⁵ - ^{?ercent seooLess} berries 3i - ^{Fresh weight} of seeileci berries 36

- ^{Fre,sh weight} of seeôIess berries 35

- ^Length of ^scedr-ed. berrics ₃₇ - ^rtrid-th of seecietL bcrrics 3T

- ^Lonp;th of seedless berries ₃₈ - ^l,'7idth of seed.less berries ₃₈ - Length/wiatfr ratio seed.eù

berries ₃₉

2"8 2"9 2,10

il

Û

ll

lt

if n

ft n n

rl

?t

lt il il

lt il n lt

il il il ll tt

il il tt

Fi^gare 2"2Q ^lluscat 1967-8

-

Length/wiatn

ratio

^seeC.less

vi

Page

46 lr7

¿t .4'l_I

48

Figure tr'igure Figure Figure Figure Figure Figure Figure Figure F.igure Figure Figure Figure

2.25 2.26 2"27 2.28 2"29 2" 30 2"31

éo )Z

2" 33

2.34 2.35 2"36 2.37

il il

Suitana

il

11

il It rt n rì

cape

il il

il It It

ìl il il il il il ll

|l

It

berries .- _{Rachis length}

- ^{Ciuster vreight}

- ^Nr:¡nber of berries Per cluster - ^{Fresh weight} of ^berries

- ^Length of ^berries - ^ItÏíd.th of benies

- Length/vrid.th ratio of ^berries - Refraction of es¡rressed juice - ^{Rachis length}

- ^{Cluster weight}

- ^{Total number} of berries - ^{Percent seedless berries} - ^{Fresh vreight} of ^seedless

bouies - Length/'*i,ftn of seeùl-ess

berries - ^{Raohis length}

- ^{Oluster vreight}

- ^Number of seeùl-ess bcrries - ^{Fresh weight} of seodlcss

berries

39 40 li0 +r11

41 42 4z 43 43

/1 /.

-r_ r Ll¿[

45

.:i')4tr,

A(,

Figure 2"38

Figure 2.39

Figure 2"{0 Figure 2.41 Fíguee 2"{2

il

1t

il

Zante

lt

rt

il

il

Figure 2"{l tt rr - Lengthfividth ratio of ^seodl-ess berries ₄₉ îi.gure 2"44 It rr - Refraction of expressed- juice ₅₀ Figure 2.{l rr rr - ^Ped-icei length 50

Figr:.re 2,{6 !' n - ^Rachis length _,1

figure 2"17 rilI - ^Cluster weight 51

Figures from section fII on the analysis of enùogenous growth substances in cleveloping flowers anå fruit

Figuire 3"1 ^{Flow d,iagrarn} of extraction tecJ:nique ⁶⁹ Figure J.2 Chromatography of extracts 80

Figure 3.3 Chromatography of anther extracts 81

Figure 3"4 ^0ape Currantc ^Changes in activity of ^endogenous regulators with time in the presence a;nd absence

of CCC ^BZ

I'igure 3.5 Sultana¡ ^Changes in ¡rctivity of en<logenous

regulators lvith tine in the presenco and absence cf ^CCC

Fi¿¡re 1.6 ÐoraC.illo: ^Chanrìes in actir¡itÍ of ^endogenous regulators with ^time ín the ^presence and

, absonce of ^CCC.

Figure 3.7 ^Changes in the concen'bration of prono'ber ÏfI

vv'ith stage of development in untreatod flowers and. berries

!''igr:re 1.8 ^Changes in the concentration of the /r3-A-Iike r:cmpound with stage of ctevelopment in

untreated flovrers and berri-es

Figr:re 3.9 The changes in the ratio of the concont¡ation of the l.B¿!-like compound. a.nd. prorooter III ^r,rrith stage of d.eveÌopment in unt¡eated- flowe¡s ant berri_es

Figures from .tippend.ix Ä.

!'igure À"1 Partition a:ral-ysÍss pre-anthesis ^Cape Ourrant

Figure Â"2 Anion ercchange chrornatogra¡hy: þost-anthesis Doraùillo and. GA,

Figure A.l Ânion exchangc chromatography: a¡ricot extract,

and- .t13¡l nlus ^GA,

X'igure A",z[ Test of chromatograp]r¡,r ned-ia for suitability in thc bioassay of plant ^horrnones

Figures from ^{Âppenùis 3}

Figure 3.1 Silica gel partition chromatcgraphy of apricot extract.

-\J:La

Page

ó)

1Gl

1vî

83

B4

85

B5

110

118

123

tr'igures from .&ppenri-ix ^C

tr''igure

C.1

GLC

of

plant ho¡mones v¡ith _QFj as ^the stationary ^phase

Figrre

C"2

^GT.C

of

plant hormones

ïrith

SI ₃₀ as the

stationary _Bhese

Figure

C"l

^GIll

of

plant hormonesl improved. recor¡ery

of G\ at

higher temperatures

Figure

C.{

^GIÆ

of

plant bormones¡ thc influence of temperature progrer,rnning

Figure

C"l

lssay

for

i-nhibj-tors

in

post-anthesis Cape

Currant e¡tracts

I'igure

C.6

GLC

of

the

inhibitive

^zone

cf

^{C¿ps Currant} extraet

that

correspond.ed-

to

tbe Rf of authentic (+) ^f,.n¿

vlii

Page

133

133

134

135

136

137

l_lc

îabIe 2"1 Tabl-e 2"2

Table 2"3

Table 3.1

Table 3.2

Table 3"J

Tab]e -4.1

fabl-e ^.4,.2 Table D.1

LTST OT' TÄBIES

Sr:.rnmary of results ^f cr 1966-7 ^antì- 1967-8 The effect of stage of development on the response of berry weÍ-ght to the application

of

^{2oppm c¿(t967-8)}

T'he effect of ^{GA applierl} at anthesis ^{ancl CCC} applieù pre-anthesis ^on the length/hid-lh ratio of berries(tg6Z-8)

Cape Currant¡ lhe effeet of ^{CCC on cluster}

ôevelopment at maturity, ^{on rrines} that ^ì¡¡ere

sampleù in the analysis of the effect of

CCC on enC.ogenous growth regrrlators Sultanac ^fbe effect of ^{CCC on cluster}

d.evelopment at maturity, ^{on vines} that were sampled- in the analysis ^c¡f the effect of

CCC on encì-cgenous growth regulators

[']re 1eve1 of the liB¡i-l-ike compound present in

ÉIrape e-ttracts anC the effect of ^CCC applied- iwo weeks pre-anthesis (averaged- over the three cultivars )

Surnrnary of chromatographic ùa.ta for silica ^gel thin layer ^plates

Ttre effcct of silica geI ^{G on} the refraction of glucose sol-utions

The detection of a number of physiologically important conporrnds on pnper vrith chromo-

genic reagents

Page

5"

59

76 6o

(t

7T

112

116

1 lrl

x

tïsî

^{0F Pt¿,18S} Page

Section III

Plate 3.1 ^One replicate of Sultana extracts after thin- Iayer chromatography

.tlopenclix Â

P1ate 4.1 Chromatograms

of

^pl-ant hormones

(*nr/+/1, IM, t.

^ABA &t¡lì.A) on

silica

^{ge1 G}

in

solvents of composition f¡om

left to rightg

_l0s!0:1,

CHCI] eEt"Lc.

;

^HCOOH; _l0s1Ce1; Bjt1Je19 BOsZ0rl;

and. JO:30¡1 ^{CHC13; ETOE:HCOOE} Appendix 3

Plate 3" 1 Chromatography of fractions eluted. from a silica gel column with rnixtures of chloroform to

methanol (see text for d-etail)

Aonend-i¡r E

?1ate E" 1

7z

114

121

150

150

150

151 151

151

151

152

Plate 8.2 Plate E"l Plate E. .{

Plate _E")

?Iate E,6 Plate _{E" J}

Plate E"B

L.S. d-eveloping ovule, 2-3 weeks pre-anthesis-

inegaspo"e mother cell

L-S. d-eveloping ovule t ^2-3 weeks pre-anthosj_s- d-iviC-ing megaspore rnother ce1l

L-S. a;ether, 2-3 v¡eeks l¡re-anthesis ^showing

po11en tetrad.s

T.S. of an ovule at anthesis showing the absence

of the embryo sac

L.S. of a¡r o¡nr1e at anthesis without an embryo sac

T.S. of an owufe at anthesis with a fully

developed. embryo sac

L. S" of the micropylar end- of a fully C.eveloped_

ovule at anthesis demonstrating the ^presence of an egþ nucleus

L.S. of an ovary sampled. approximately 2 weeks

after anthesis

T.S. of a¡ ovary sampled- approwimately 2 v¡eeks

after anthesis, d.emonstrating the expansion of the nucellus without a concomita¡rt d-evelopment

of the integuments

Plate E"l

152

xl- ÄSPEOTS 0F TIm IIOR]úON.¿I-L PIITSIOL,OGY CF F}ìUIî ÐElEf,OPLtúlI'I

rN VTTIS ^./TNIFERI.

SLlflü.¿rÌ,Y

[\ro concplementar¡r stuùÍes v¡ere

unC.ertaken of the horrnonal physiolog;y of set anå

clevelopment of tire fruit of Vitis ¡j-]!ljier,a, wi-th special reference to gibberellin pronoters antr inhibitors" In

one approach the cffects of applications of factorial combinaticns of ^GX. and- CCC were rneasurecl on ^-,¡ari-ous parameters of cluster developmont. Time of application of ^{each comporrnd} was also exa¡ninetl. In the seooncl-

apprcach, estimati-ons cf entogenous G.Á-like ¡¡nd X3A-1ike compor:ncls .¡¡cre _maùe in ext¡acts of flowers anl fruits

ù:rin6ç the critical periorl-s of, anthcsis ancl early pericarp

clevelopment.

Pre-anthesis applications of ^GÀ

increasecl fruit-set in the stenospermocarpic Sulta^na, rlecreasecl sot in tJ:c parthcirccarpic lapo ancì hacf nc¡

effect on three othet cultivars. Anthesis

applicaticns increasecl the set of seed_less benies in Doradillo, had no effect on I,{uscat, tape cr Zanter md c-ecreaseiì- set iir sultana. Post-anflresis .applications increaseù the set of seeåless berries on cultivars other than the parthenocarpic. Pre-anthesis ap;rlications of

CCC increaseå set thou¿çh not ali,vays signifioantly. ^post- anthesis applications r¡¡ere without effect on set. t----..--..-

xra

n

Àppl-ications cf ^G.tl, prior to anthesis were the most effective in reducing seecl- .levelopmentg post-

anthesis applications ^lvere '¡rithcut effect ^on Doracr.illo

but reciucec'r seeÒ d-evelopnent in l,[uscat. Intermec]-iate resuJ.is ï\rere generally obtained from anthesis applicaticns. ^{I'Vith one} exception ^(Muscat pre-arithesis 196Ç7) ^CCil ciiù not alter the

proportion of seeåecl anir. seeôl-ess berries set"

In general both ccmpounds exerteù

signifioant, usualì-y oirposite, effects on pericarp d-evelopment" ïlhere seeà growth was unaffecterl- by ^G,A.

application, this compoun.,l hacl no influence olt the final fresh weight of the berry. Ilowever, if seed developrnent was imperfect then GA ha,L a ^profound. effect on the final

size of the roemy. The magnitucle of this effect ùepenC-ecì-

upon the cul-tivar ancl time of applicatione those berries in which seecl clevelopment vras incomplete sho¡,'¡eil greatest response to ^G.rt applied at set, but vrhere no seed- growth occurred at all then response ï\¡as greatest to a"nthesi-s

applioations. ^CCC, irrespective of tíme of application, significantly recr:ucerl berry size in all cultivars except those rn¡hich vrere parthenocarpi_c.

G-4. significa^ntly affecteù the shape

of bemies, especially wh.en appliecÌ at anlhesis. ^The

clegree of the response vras unrel_ated to seell d_evelopment,

although in the ^seeded- cultivar I'nluscat the change in

shape \¡/as unaccompanied by any change in berry vreight.

Cape was the most responsivo, while l\fuscat, SuJtana ancl

Zante repre less so. DoracliLlo _was r:naffecteC_. CCC

significa.ntl-y recì.uced. the renglh/-,tlid-th ratio of ^a1l_ berries with a norrnal_ ratio greater than about 1"1"

XI11

GÀ often significantly increased

the length cf the rachis anct pedicel but only when

applied- pre-anthesis. There was a trencl, sometimes

significant, for ^CCC to retard- the elongation of the

rachis a¡rd the ^ped-icel.

Cluster vreight was affecteci by

the treatments in a manner similar to berry weight"

Pre-anthesis applications of ^GÀ to seeclec) cultivars resultecl in '.lecreaseù yieì-d- because a greater proportion of berries had- inhibitecl seecl growth a¡:rL were smalIer.

Later applications of GA, notably in Doraùi11o, increased.

yielcl because of retuceù inhibition of ^seed growth and_

greater set a¡icì clevelopment of seec"less berries" -Anthesis

applications of ^G.A. were most effective Ín increasing yield- in zante and- cape, while post-anthesis applications

ïuere more effective in Sultana.

fn tho exami_nation of enC.ogenous growth

regulators in the fruit of Ðoratli11o, sultana anc]- caper

qualitative d-ifferences in GA-rike compounds were noteti- betlveen the parthenocarpic cultivar, Cape, ar:cL the

other two cultivarsi the ùifference between Sul_tana a.reô

Dorad-illo _appearec to be only quantitative" significant ⁱ

changes in ooncentration of GA-rike substances during eorly ^íLl

development occumecì q¿ly _1n the seedeù cul_tivar Doraclil1o. ^l

No significant response to ^CCC was noteù. ^,,

xLv

Jtn ¿3.4-1ike ccmpound. was present ì_n

all extrasts examj.ne'ù and. sigr:ifica::t changes occr:med in its concent¡ation. îhere ^'was a trencl in all three

cuLtivars (significant in DoraCrillo, P- 0.0)) for the

leve1 of the inh-lbitor, which was generally highest at anthesis, to ùecline nole slowÌy after treatment with

ccc.

These results and the results of othe¡s a¡e usecl to formulate a general b.¡4lothesis of the control of fruit-set and. pericarp C.evelopment in Vitis vinifera.

-

ï\'

This thesis contains no material r:vhich

has been accepted for the avrard of any other ^d-egree or ^di-plorna i-n any u:riversity ano, to the best of my belief, no material which has previously been published- or .¿lr¡itten bJ' another person has been included, except v¡hen d-ue reference is ^made in the text of this thesis"