The distribution of Ricinodendron rautanenii trees is
confined to a fairly distinct band across southern
Africa. This stretches from South West Africa in the
west, through Botswana and Zimbabwe, to Mozambique in
the east. These plants are a potential source of both
timber and food. In this respect, the fruits and seeds
of this species are highly nutritious and the latter,
by virtue of their high lipid content, represent an
excellent source of oils for both domestic and industrial
uses. For these reasons consideration is being given
to cultivating this species on a commercial scale. One
problem, however, is that the seeds of these plants are
dormant and in the first part of this study an attempt
was made to establish the cause of this dormancy and how
it could be overcome.
A thorough investigation of all the possible causes of
dormancy revealed that ethylene was the only naturally
occurring stimulus that could relieve this condition.
Exogenously applied gibberellin (GA3 ) and ethrel were
found to be equally as effective as ethylene, but such
treatments cannot be regarded as natural. It was also
found that all the dormancy breaking treatments were only
effective once the endocarp had been removed. This indicated
that R. rautanenii seeds had a combined coat
imposed-physiological dormancy where ethylene was only
able to stimulate germination once the endocarp had been removed. Scarification treatments showed that the
endocarp most probably had its effect by restricting
embryo enlargement since this structure did not inhibit
water uptake or gaseous exchange. Once the ger-
mination requirements of these seeds had been established
a more detailed investigation was carried out to determine
their general sensitivity to ethylene, as well
as the actual role this gas played in breaking dormancy.
Ricinodendron rautanenii seeds were found to exhibit a
high degree of sensitivity to ethylene. The threshold
concentration at which a response was obtained was
approximately 10¯³ microlitres per litre and this is
the lowest yet recorded for any species prior to any
additional seed treatments. At concentrations above
this, the response was saturated indicating that the
seeds are well adapted to the ethylene concentrations
most likely to occur in the field. In addition to
this, manketti seeds also responded to ethylene after
only very brief exposures to the gas and optimum germination
was recorded after 30 minutes incubation in
an ethylene saturated atmosphere. The temperature
range over which an optimum response to this phytohormone
was obtained was found to be between 25 and
35°C.
One of the most striking features regarding the sensitivity
of these seeds was the apparent ability of dry
and partially imbibed seeds to perceive the dormancy breaking stimulus. Furthermore, once treated, the
seeds retained the dormancy breaking effect of ethylene
even when subjected to almost complete re-dehydration.
In this instance, 50 per cent germination was recorded
for ethylene treated seeds which had lost approximately
97 per cent of their moisture content between dormancy
breaking and re-incubation. It was thus concluded
that, not only could Ricinodendron rautanenii seeds
respond to very low ethylene concentrations but could
probably also retain the effects of this gas during
adverse environmental conditions.
The effects of imbibition and dormancy breaking were
followed separately at the ultrastructural and biochemical
level. The ultrastructure of dry embryonic
axes of these seeds was characterized by massive stores
of food reserves in the form of lipid and protein.
Upon imbibition the number and size of spherosomes decreased
and protein and globoid hydrolysis was clearly
evident. Polysomes and microbodies (including mitochondria)
were also visible prior to dormancy breaking
but there was no evidence of any endoplasmic reticulum
or dictyosomes. Imbibition also resulted in the expans
ion of the nuclei and there were indications of an
increase in the granular content of the associated
nucleoli. The number of nucleolar vacuoles, however,
remained unchanged. These features indicated that
nuclear activity had commenced albeit limited. The
ultrastructure ·of untreated seeds which were maintained in the imbibed state for an extended period of time
(six days) was also examined. Cells of the embryonic
axes of these seeds showed no further changes with regard
to their nuclei and protein hydrolysis appeared
to have ceased. At this time spherosomes resembled
those in freshly imbibed tissue in terms of their size
and numbers, suggesting that the lipid reserves had been
resynthesized.
No immediate ultrastructural changes were observed after
ethylene treatments. However, 24 and 48 hours after
dormancy breaking further expansion of the nuclei was
noted. At the same time the nucleolar vacuoles disappeared
and the granular content of this region increased
markedly. This suggested that an increase in
the synthesis of various RNA fractions was taking place.
Vigorous protein hydrolysis was also observed after the
ethylene treatment whereas spherosome numbers increased.
Three days after the dormancy breaking treatment, the
first signs of germination were visible. Externally
this was characterized by a splitting of the testa in the
region of the radicle. At this time, endoplasmic
reticulum and dictyosomes were still not visible but from
this point onwards the ultrastructural changes observed
were typical of those recorded during the germination of
other species. Thus, no single ultrastructural feature
could be associated with the breaking of dormancy and the
most notable changes which occurred during this period took place in the nucleus.
Biochemical changes occurring during imbibition resulted
in an overall decrease in the levels of extractable food
reserves present in the embryonic axes. During this
period, lipid levels were found to decrease by 44 per
cent, protein levels by 12 per cent, sucrose levels by
68 per cent and glucose, fructose and starch levels by
100 per cent. These levels were found to return to
their original values when seeds were incubated under
moist conditions in the absence of ethylene for extended
periods of time. Ethylene treatments, on the other hand
caused a further, marked decrease in sucrose levels,
whereas protein and lipid levels increased. Hydrolysis
of the endosperm reserves commenced three days after the
application of ethylene and this was characterized by a
decrease in lipid levels and an overall increase in
soluble carbohydrates. The timing of this event
suggested that the endosperm was not involved in the actual
process of dormancy breaking.
The importance of protein synthesis in dormancy breaking
was also investigated. It was found that seeds incubated
with a protein synthesis inhibitor, cycloheximide,
failed to germinate, confirming the view that protein
synthesis is an essential pre-requisite for germination.
Inhibition of RNA synthesis with actinomycin D, on the
other hand, did not prevent germination. This suggested
that the materials necessary for early protein synthesis were already present in the dry seeds. Actual measurements
of protein synthesis showed that this process
took place in the embryonic axis, cotyledons and endosperm
of seeds imbibed for as little as two hours.
Protein synthetic abilities increased considerably in
most instances after 48 hours imbibition but then decreased
upon application of ethylene. At the same time,
however, a marked increase in the uptake of 14C-Ieucine
was noted in ethylene treated axes. This may indirectly
reflect an effect of ethylene on membrane permeability.
Protein synthesis in cycloheximide and actinomycin D
treated embryonic axes was also measured. No consistent
trends were evident but it was found that after ethylene
treatments, protein synthesis was generally lowest in
those seeds which were destined to germinate. In
addition, these seeds also exhibited the greatest uptake
of [14]C-Ieucine.
Ricinodendron rautanenii seeds incubated with compounds
known to stimulate the pentose phosphate pathway failed
to germinate. This indicated that dormancy in this
species was probably not the result of a block in alternate
respiration.
The possible involvement of endogenous phytohormones in
the overall process of dormancy breaking was also in-
vestigated. In this regard, the role of gibberellic
acid appeared to be enigmatic. This is based on the
observation that applied gibberellins could stimulate germination whereas inhibitors of endogenous gibberellin
synthesis applied to ethylene treated seeds had no effect.
It was concluded from this that the effects of ethylene
are not mediated via an enhancement of endogenous
gibberellin synthesis.
A preliminary investigation carried out on the endogenous
cytokinins showed that this hormone was absent from dry
and imbibed seeds. A transient increase in zeatin levels
was observed 24 hours after the ethylene treatment. A
similar transient increase was noted in non-induced seeds
maintained under moist conditions for six days. In this
latter instance, however, the peak co-chromatographed
with the biologically less active cytokinin, zeatin
glucoside.
A basal level of endogenous ethylene production was recorded
in all imbibed Ricinodendron rautanenii seeds.
Ethrel, ethylene and gibberellin treatments caused an
initial, transient increase in this ethylene production
after which no further significant changes were recorded.
It is suggested that dormancy breaking in this species is
not related to enhanced endogenous ethylene synthesis.
The results of the biochemical and ultrastructural studies
are discussed in relation to what is known regarding
features associated with dormancy and its removal and on
the known effects of ethylene on seed tissues. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 1982.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:ukzn/oai:http://researchspace.ukzn.ac.za:10413/11152 |
| Date | January 1982 |
| Creators | Keegan, Arthur Brian. |
| Contributors | Van Staden, Johannes. |
| Source Sets | South African National ETD Portal |
| Language | en_ZA |
| Detected Language | English |
| Type | Thesis |
Page generated in 0.0345 seconds