The production of two kinds of spores differing in structure and function, the smaller one producing the male gametophyte and larger one producing the female gametophyte, is termed heterospory.
For the first time distinct heterospory (structural as well as functional) is evident in pteridophytes; but out of several hundred forms in the group, only 9 genera (Selaginella, Isoetes, Stylites, Marsilea, Pilularia, Regnellidium, Salvinia, Azolla, and Platyzoma) are heterosporous.
Heterospory has been of considerable interest because of its bearing on the evolution of seed. It brings in a new type of gametophyte, endosporic, which is a biological advancement over exosporous gametophyte of homosporous forms.
Female gametophyte developing within large megaspore at the expense of reserve food is independent of external environment and is a better place for the initiation of embryo. Contrary to it, free-living (exosporic) green prothalli of homosporous forms are dependent on external conditions and have to be self- supportive in food supply.
How far pteridophytes approach seed habit or fall short of it is a matter in itself but our understanding of the phenomenon of heterospory is far from complete. Heterospory can be ascribed to result from expression of one sex and repression of other in particular regions. Plants with sex chromosomes have a direct cytological basis for sex expression but none of heterosporous pteridophytes are reported to have sex chromosomes.
Incipient Heterosporous Form:
An account of heterospory should begin with the form showing incipient heterospory. Equisetum is a homosporous form but produces two types of gametophytes; smaller ones are male and larger ones female.
The latter become hermaphrodite if fertilization is delayed. Nonetheless, in a population the proportion of male and female gametophytes is influenced by environmental conditions. Sex determination, therefore, seems to occur during vegetative phase of gametophyte.
Developmental Events and Heterospory:
In the absence of sex determination process, in heterosporous pteridophytes the developmental events are the best guide for an understanding of the phenomenon of heterospory. It has been pointed out that differences between microspores and megaspores are there not only in terms of size and sex-expression but these spores are to be viewed in terms of food reserves, organelles, nuclear shape and wall construction.
A perusal of the forms showing heterospory indicates that the most significant events in the determination process are:
(a) Time of sex determination.
(b) Number of archesporial cells in a sporangium; whether the archesporial cells directly differentiate as sporocytes or divide mitotically thereby increasing the number of sporocytes.
(c) Number of sporocytes initiating and completing meiosis.
(d) Number of spores attaining maturity.
In Selaginella, the micro- and megasporangia are similar until the sporocyte stage. All the sporocytes in a microsporangium undergo meiosis and produce large number of microspores. In a megasporangium most of the sporocytes degenerate leaving only a few to undergo meiosis and in most of the species only one sporocyte survives to form spores. After meiosis further difference may occur in spores.
In most species four functional megaspores result, but in some, the four spores are of different dimensions. The tetrad may result into two large and two small spores (S. stenophylla) only one large and three small spores (S. molliceps) or as an extreme a single large, functional megaspore with no trace of degenerates (S. erythrops). In a collection of Selaginella sp. the author has, observed presence of larger spores in microsporangia (Rashid, previously unpublished).
Determining processes leading to heterospory are operative at different stages of development and no generalization is possible. Among lower pteridophytes, in Selaginella the determining process is operative at premeiotic as well as postmeiotic stages. By contrast, in Isoetes it is operative only at premeiotic stage. Among higher pteridophytes, in Marsilea the control is postmeiotic but in Salviniaceae it is premeiotic as well as postmeiotic.
Microsporogenesis versus Megasporogenesis: An Appraisal:
Microsporogenesis is comparable to sporogenesis in homosporous forms. Almost all of sporocytes complete their development and produce innumerable microspores. The microspores are, however, very small structures with very little food reserves and their chloroplasts are incapable of further development or synthesis.
A microspore is, therefore, dependent in its growth on a small reserve, confined to a small space, its cytoplasm is also poor in colloidal macromolecules. This suggestive limitation of food supply and insubstantial cytoplasm leads to maleness in direct contrast to femaleness of megaspore.
In megasporogenesis there is extensive degeneration of sporocytes as well as of megaspores formed. There are more than one ways of forming megaspores. In Selaginella there is competition between megasporocytes and a variable number of megaspores are formed.
Establishment of Femaleness:
Femaleness in a megaspore can be described to be present ab initio. This is evidenced by rapid differentiation of female gametophyte and female sex organ.
Cellular and Subcellular Events in Differentiation of Heterospory:
Some studies Horner & Beltz, 1970; Pettitt, 1970, 1971, 1974, 1976, 1977 provide an insight into the ways in which determining processes leading to heterospory are operative.
Early studies on cellular differentiation of heterospory are indicative of the following significant differences in two types of sporangia of Selaginella pilifera:
(In this species microsporangia and megasporangia develop opposite to each other):
(a) The megasporangial primordium is slightly larger than microsporangial primordium,
(b) During development, all sporocytes except one in megasporangium show loss of RNA stainability, whereas the entire sporogenous tissue in a microsporangium shows uniform staining,
(c) All microsporocytes prior to meiosis are surrounded by distinct callose wall, whereas in a megasporangium only the functional megasporocyte shows callose wall.
The presence of callose wall around the developing sporocyte is suggested to isolate it from surrounding cells and facilitate it in its new developmental pathway. Nevertheless, the underlying cellular events leading to early expression of spore type and sex determination are still an enigma.
Studies on megasporocyte degeneration in Selaginella spp. provide answers to some of the questions such as:
(a) Why some megasporocytes degenerate?
(b) Of the surviving megasporocytes which one is able to complete meiosis and form megaspores?
(c) Of the megaspores formed which is the functional?
In the megasporangium of S. sulcata approximately equal number of megasporocytes of two kinds are discernible viable and non-viable. These can be distinguished on the basis of their size and ultrastructure. The viable ones are about 15 microns or more in diameter and are characterized at the prophase by the aggregation of mitochondria plastids at the centre of cell.
In addition, their nuclei contain intranuclear vesicles. The non-viable ones are about seven to ten microns in diameter and are characterized by the presence of lysosomes (implicated in cell death) and degenerate by a process corresponding to cellular autophagy.
Presence of lysosomes can explain degeneration of some of the sporocytes but does not account for final selection; in which only one of the megasporocytes is able to complete meiosis and form megaspores. Nonetheless, the intranuclear vesicles in viable megasporocytes of S. sulcata are similar to certain of the cytoplasmic vesicles implicated in autophagy.
These vesicles might be affecting nuclear degeneration. If this phenomenon is operative in all, except one of the megasporocytes, it can be understood as to why only one sporocyte is successful in meiosis. Along with the megasporocyte which completes meiosis and forms megaspore tetrads the others which fail also persist in the sporangium.
On these persisting megasporocytes an exine is deposited but it is neither massive nor structurally complex like the wall which surrounds the megaspore. The persistent megasporocytes gradually degenerate as the spores of megaspore tetrad enlarge.
In efforts at resolution of developmental mechanism controlling heterospory there is evidence for pinocytosis during microsporogenesis in Selaginella. The microspores take up material of maternal origin. The injested material is degraded within the cell by vacuolar system, which allows some of the material to pass into the cytoplasm. The significance of this finding will be apparent on knowing the comparable information on megaspore side.
The mature megasporangium in S. sulcata contains a single tetrad of megaspores and the spores in this tetrad are of two types. Two of the first kind are 350-400 microns in diameter and appear to be functional. Other two of the second kind are 200-300 microns in diameter and appear to be abortive.
It is possible that viable megaspores are the ones which have inherited the mitochondria or plastids from the mother cell. These propositions could be confirmed by densitometric measurement of the megasporocyte cytoplasm which was found to be polarized. At cytokinesis the cytoplasm was partitioned into four equal cells but not equal in composition, thus suggesting an innate constitutional differences between functional and non-functional megaspores.
Environmental Factors and Heterospory:
A perusal of developmental aspects of heterospory in different forms indicates that determination of spore types is operative at different stages of development. Nutritional and environmental factors influence the pattern of development. Quite early it was found that spore development in Marsilea can be manipulated by environmental factors.
Under favourable nutritive conditions, spores in developing microsporangia aborted and the survivors enlarged. The enlargement was proportional to degeneration and spores enlarged up to 16 times their normal size. Rarely single spore survived and in its cytological features; such as vacuolation, nuclear shape and position to the cell it resembled a megaspore.
In plants grown in unfavourable nutritive conditions, such as chilling, microspores were found in sporangia which, by their position at the tip of sorus, would contain megaspores. However, these altered spores were not made to germinate and it is, therefore, not certain whether alteration was mere structural or functional.
More significant are studies on Selaginella which indicate that the formation of megasporangia in this plant is under ethylene control. Clumps of S. wallacei when sprayed with 2-chloroethylphosphonic acid (ethepon), an ethylene releasing compound, at 7.65 and 76.5 mg/L favoured the formation of megasporangia and controls sprayed with water, produced high proportion of microsporangia.
Treatment of S. palescens with ethepon (38 mg/L) caused the production of megasporangia in the microsporangiate files of strobilli. In Selaginella the distinguishing feature between micro- and megasporangia is the number of sporocytes; there are many more microsporocytes in a microsporangium than the number of megasporocytes in a megasporangium.
Ethylene is known to retard cell division and bring about cell degeneration. It is suggested that ethylene acts by reducing the number of sporocytes in Selaginella by inhibiting cell division in sporogenous tissue and induces degeneration of sporocytes, thus favouring the formation of megaspores.