ndylium), embryos of seeds held at 15°C elongate for approximately 6 weeks, then stop growing when
approximately half their full size [45]. Although the initial rate of growth is slower in seeds held at 2°C,
elongation of the embryo continues logarithmically for 9 weeks. Parallel changes occur in the endosperm,
but in reverse (i.e., the endosperm of seeds held at 2°C is consumed by the embryo, while that in seeds
held at 15°C is not). If lack of suitable nutrients were responsible for the failure of embryos to develop at
15°C, one would expect that growth of excised embryos in vitro at 15°C could be stimulated by supply-
ing appropriate nutrients. Stokes [93] observed that arginine and glycine concentrations in the endosperm
were higher in seeds held at 2°C than in those held at 20°C. When embryos cultured in vitro at 20°C were
supplied with glucose plus various sources of nitrogen, arginine and glycine were the most effective
amino acids in supporting growth, although KNO 3 was the best source of nitrogen. From these and other
data, Stokes [93] concluded that exposure to 2°C stimulated embryo growth by increasing the quantities
of arginine and glycine available to the embryo.
A similar situation occurs in both black ash (Fraxinus nigra) [94] and European ash (F. excelsior)
[95,96], except that chilling is not essential for embryo enlargement but is required for germination once
embryos have reached full size. Stokes [92] provides other examples of seeds with similar requirements.
Axes from dormant hazel embryos will grow in vitro when supplied with inorganic salts and sucrose [97],
suggesting that failure of the intact embryo to germinate is due to inability to mobilize nutrients from the
cotyledons [98,99]. Application of GA 3 both breaks embryo dormancy and permits mobilization of re-
serves, suggesting that gibberellin biosynthesis following chilling has a similar effect (see later).
- Protein Metabolism
A group of proteins termed “late-embryogenesis-abundant” (Lea) proteins accumulates as seeds mature
and become dehydrated (see Ref. 87). These appear to bind water, thereby protecting macromolecules
such as nucleic acids (?) from dehydration and resultant denaturation. Leaproteins disappear during ger-
mination.
Several facts, summarized by Quatrano [100], suggest that such proteins play a role in dormancy: (1)
embryos of viviparous mutants do not synthesize these proteins if cultured on a medium containing ABA;
(2) dehydration of immature embryos induces the production of the proteins, possibly by stimulating the
synthesis of ABA; and (3) treating mature seeds with ABA prevents both germination and the loss of Lea
proteins.
Most studies of Leaproteins have involved species whose seeds either are nondormant or have a
shallow dormancy, and no studies are known involving species with deeply dormant seeds. Therefore, the
connection between such proteins and dormancy remains tenuous. ABA blocks germination while in-
ducing or maintaining the synthesis of Leaproteins, but these two responses may be unrelated.
Protein metabolism has also been implicated as a factor in the breaking of dormancy. As already
noted, holding Heracleum sphondyliumseeds at 2°C permits the hydrolysis of reserve proteins and their
transfer to the embryo, whereas holding them at 20°C does not [45]. In apple embryos, however, hydrol-
ysis of reserve proteins occurs at both 5 and 20°C [101]. Furthermore, no proteolysis is observed in seeds
held in the fruit at 0°C, although this treatment also breaks embryo dormancy. Similarly, Chen and Varner
[102] reported that dormant and nondormant seeds of wild oats (Avena fatuaL.) synthesize protein at sim-
ilar rates.
Lewak et al. [103] suggested that an insufficient supply of amino acids may prevent germination in
dormant apple seeds. Protease activity increases with chilling, reaching a maximum after 7 weeks, then
declines to the level observed in nonchilled seeds. The authors suggested that germination is dependent
on a supply of amino acids released by hydrolysis of proteins. However, they presented no data on the ef-
fects of amino acids on germination of dormant embryos.
Subsequent work (see later) emphasized the effects of dormancy-breaking treatments on the con-
centrations of specific proteins or polypeptides. The rationale for much of this work is that regardless of
what substances control induction or breaking of dormancy, enzymes (proteins) must be synthesized be-
fore such compounds can be produced. Therefore, changes in protein content should precede changes in
other compounds, be they carbohydrates or hormones. Protein analysis involves electrophoretic separa-
tion of extracted proteins, together with the use of radiolabeled amino acids as markers for newly syn-
thesized polypeptides. Although no significant changes were observed in total soluble protein content of
pear [104] or apple embryos [105] during chilling, Eichholtz et al. [105] observed an increase in the con-
centrations of four peptides in the embryonic axes of apple embryos held at 5°C. No changes were evi-
DORMANCY: MANIFESTATIONS AND CAUSES 173