The Development of Endosperm in Grasses

The grass seed or caryopsis originates from a monocarpellary ovary with a single ovule and contains the main storage tissue, the endosperm. For most grass crop species (i.e. cereals), the value of the crop is largely determined by the endosperm, both in quantitative and qualitative terms. The endosperm is the result of the fertilization of two polar nuclei in the central cell of the embryo sac by one sperm cell nucleus, which generates a triploid (3n, 3C) nucleus, whereas the diploid (2n, 2C) embryo originates from fertilization of the egg cell by the second sperm cell nucleus. The main function of the endosperm is to provide nutrients to the developing and, later, germinating embryo. In contrast to many species, including Arabidopsis (Arabidopsis thaliana), the grass endosperm is a persistent seed structure. It is the foremost source of calories for human and livestock nutrition and provides the raw material for countless manufactured foods, goods, and biofuels. In spite of the importance of the grass endosperm, its development has not been thoroughly investigated in many crop species, much less in noncrop species. There is considerable uniformity in the development of the endosperm among grasses, especially during its early stages (Weatherwax, 1930; Bennett et al., 1975). And although deviations are known, these generally involve secondary aspects of development. It is generally true that the endosperm of most grasses is starchy and dry at maturity, which of course is a valuable trait, but there are exceptions. For example, in a survey of 169 grass genera (over 25% of total genera in the family), 30 were found to possess species with liquid or soft endosperms at maturity, and the viscous state of the endosperm can be retained for several decades. Nine additional genera were found to have semisolid endosperms (Terrell, 1971). Among grasses, endosperm development is by far best characterized in maize (Zea mays) for historical, economic, anatomical, and genetic reasons; therefore, we will primarily refer to knowledge obtained from this cereal as a paradigm for grass endosperm development. Wherever appropriate and possible, differences between maize and other grass species will be highlighted and discussed. Here, we provide an overview of the phases of endosperm development, including the unique features of genetic, molecular, and cell regulatory mechanisms. The reader interested in an in-depth discussion of different aspects of endosperm development in grasses is referred to several previous works (Kiesselbach, 1949; Bennett et al., 1975; Kowles and Phillips, 1988; Lopes and Larkins, 1993; Olsen et al., 1999; Becraft, 2001; Larkins et al., 2001; Olsen, 2001, 2004, 2007; Sabelli et al., 2005b, 2007). The analysis of mutants has provided substantial knowledge of the regulation of endosperm development, and the relevant literature is extensive. Rather than reviewing every mutation, we will discuss selected mutations that in our opinion provide crucial insight. There are a number of publications that contain a wealth of information about mutations affecting endosperm development in cereals (Jarvi and Eslick, 1975; Nelson, 1980; Neuffer and Sheridan, 1980; Satoh and Omura, 1981; Bosnes et al., 1987; Kowles et al., 1992; Scanlon et al., 1994; Kurata et al., 2005; Dolfini et al., 2007). Phases in endosperm development. Although this figure refers to maize, it is a good example of endosperm development in other grasses as well. A, Double fertilization, syncytium formation, and cellularization of the endosperm occur within 3 to 4 DAP. The pollen tube and sperm nuclei are shown in yellow, polar nuclei in the central cell of the female gametophyte and endosperm nuclei are shown in red, and the egg cell nucleus and embryo nuclei are shown in green. Outlines of the multicellular endosperm and embryo are drawn in red and green, respectively. Modified from Kiesselbach (1999) with permission. B, From about 4 to 20 DAP, the endosperm undergoes a phase of mitotic cell proliferation, followed (from around 8–10 DAP) by endoreduplication, as shown by flow-cytometric profiles obtained from 7-, 11-, and 19-DAP endosperms (red), and by PCD (starting around 16 DAP). The endoreduplication phase and part of the cell division phase coincide with a dramatic growth of the endosperm and the accumulation of storage compounds. The dynamics of key parameters during mid endosperm development, such as fresh weight (red line), nuclei number (blue line), mitotic index (brown line), and average DNA content (C value; green line), are shown at bottom. They are loosely based on the work of Engelen-Eigles et al. (2001) and only illustrate trends. Al, Aleurone; CSEn, central starchy endosperm; Em, embryo; En, endosperm; Nu, nucellus; Pe, pericarp; Pl, placentochalaza; SAl, subaleurone layer; TC, transfer cells. This panel is reproduced in part from Larkins et al. (2001) and Sabelli et al. (2005b), with permission. In many species, including maize (Kiesselbach, 1949), wheat (Triticum aestivum; Bennett et al., 1975), Job's tears (Coix lacryma-jobi; Weatherwax, 1930), Koda millet (Paspalum scrobiculatum; Narayanaswami, 1954), and Chinese lovegrass (Eragrostis unioloides; Deshpande, 1976), syngamy (the fusion of one sperm nucleus with the egg cell nucleus) and fusion between a sperm nucleus and two polar nuclei to create the endosperm are simultaneous. It is intriguing that the mitotic activity of the triploid primary endosperm nucleus is very fast, whereas this process in the zygote undergoes a noticeable hiatus. Recent findings in Arabidopsis indicate that proliferation of the central cell requires a positive signal from the fertilized egg cell (Nowack et al., 2006), implying that the polar nuclei are “primed” for faster cell cycle activity. Whether a similar mechanism operates in the grass family is not known. The presence of supernumerary polar nuclei prior to fertilization has been documented in Koda millet and sugarcane (Saccharum officinarum; Narayanaswami, 1954) and in the indeterminate gametophyte maize mutant (Lin, 1978), suggesting a latent pathway with the potential to lead to early cell cycle activity and premature endosperm proliferation, which are otherwise normally repressed. Grass endosperm follows a frequently encountered mode of endosperm development, the nuclear (or coenocytic) type (Lopes and Larkins, 1993; Olsen, 2004), in which the initial triploid nucleus undergoes several rounds of often synchronous division in the absence of cell wall formation and cytokinesis, resulting in the formation of a syncytium. Fertilization of the polar nuclei results in the primary endosperm nucleus, which within hours begins to rapidly divide. As previously noted, cell division in the zygote is invariably slower. For example, by the time the zygote divides for the first time there are four to eight endosperm nuclei in maize (Randolph, 1936), up to 24 endosperm nuclei in Koda millet (Narayanaswami, 1954), and a large number of endosperm nuclei in Chinese lovegrass (Deshpande, 1976). When the embryonic cells do start to proliferate, they divide at a slower rate than the endosperm nuclei (comparable to the rate of cell division of meristematic cells), most likely because proliferation of the endosperm nuclei does not involve the synthesis of cytoplasm, cell membranes, and cell walls. The observation that a decline in the rate of endosperm proliferation (to approximately equal that of embryonic cells) is concomitant with cellularization of the syncytium lends support to this interpretation (Bennett et al., 1975). Indeed, the cell cycle of the coenocytic endosperm typically lacks the formation of interzonal phragmoplasts between daughter nuclei, reinforcing the view that the program responsible for creating some parts of the cytoskeletal apparatus found in somatic cells is suppressed. Thus, coenocytic endosperm development could be viewed as an evolutionary strategy, through repression of the program controlling cytokinesis and cell wall formation, to rapidly populate the large, preformed cytoplasm of the central cell and attain a greater basal cell number to support the growth of this tissue and prepare it to nurture growth of the embryo, especially during the period in which specific cells dedicated to nutrient uptake by the endosperm have not yet differentiated. The maternally derived nucellus and antipodal cells probably support coenocytic endosperm growth with amino acids, nucleotides, and carbohydrates (Bennett et al., 1975; Radchuk et al., For about endosperm nuclei synchronous with to cell cycle are in which nuclei from the to the embryo to the to between and in (Bennett et al., 1975). In maize, nuclear proliferation in the syncytium generates up to nuclei, during the first 3 DAP. In several and species, in of nuclei have been (Bennett et al., 1975). nuclei the of the embryo sac and, as a result of of the central at the of the primary endosperm in there is a for about the initial period of proliferation, which with a dramatic of the to prepare for the cellularization of the first of It is not this of mitotic activity is in other grasses, such as maize and in which cellularization rapidly analysis of maize endosperm development mutations at the that the first division of the primary endosperm nucleus two endosperm and a of cell proliferation from the to the of the tissue of that the main genetic controlling the rate of coenocytic endosperm development can be on specific (Bennett et al., 1975). The of antipodal cells of the embryo sac by mitotic division during growth of the endosperm and to at endosperm stages (Randolph, Kowles and Phillips, In the coenocytic endosperm undergoes cellularization by the formation of and an process that from the of the endosperm the central et al., 1994; Olsen et al., 1999; Olsen, are by that from the nuclear resulting in approximately nuclei in one the central cell wall formation is through the of at of and of an This is followed by of the cell resulting in which are an cell wall that nucleus. of the cell the nuclei divide and which is followed by Thus, the of is the central with an of The process of cellularization the central cell is with which in cereals is around 3 to DAP. this process the formation of a a cytoskeletal that typically the of the cell wall during the somatic cell cycle Among grasses, endosperm development is best in cell the cereal transfer starchy endosperm and cells. cell of the cereal endosperm, the and cellularization is transfer cells. cells have cell wall and which nutrient and amino uptake by the endosperm. cells have been in some in several grass species in et al., these cells are frequently found at the of the endosperm, within the caryopsis among species et al., In and transfer cells are the and in maize, they are the They at such as at the between tissue and the endosperm. with other the of these cells probably has a of et al., cells typically have a cytoplasm that is in are during of the transfer of the maize a that results in a transfer cell and endosperm et al., 2007). The has been to transfer cell in and its of accumulation has been as a for in a specific of the coenocytic endosperm that will transfer cells et al., cell during a of development, as shown by the of the maize which has an basal of transfer cells et al., in the central cell of the maize embryo sac are for of the transfer cell et al., of maize are in transfer cell and et al., suggesting a in the from potential and to be by a which is the in the basal of the coenocytic endosperm et al., Recent in with maize endosperm have previous that development of the basal transfer cell requires a from tissue et al., cells a generally one and to or several of cells that the endosperm in the transfer cell In maize, the between and from the of endosperm which to and and cells have and other cytoskeletal of meristematic is to be and the first division of the endosperm et al., the first in cell among cereals and to the number of in different species. For example, in the first of cell is the accumulation of and cytoplasm DAP). In maize, it follows a period of cell from the endosperm and is by of cell division from to which results in a of cells most of the starchy endosperm. Although formation both and cell only the to its 20 (Kiesselbach, cells to the are and than the starchy endosperm cells and are referred to as subaleurone cells. growth of the is to endosperm growth The mutants in maize, with cell division in the and development of starchy endosperm, support this view et al., cells are normally but in they endoreduplication and are of and in and and and in of cells to be from that of transfer as shown by the maize which lacks but a of transfer cells et al., The cytoplasm of cells is and because of with cells contain which a of to the maize The is the only tissue at endosperm maturity, a specific program that it from et al., seed in to from the embryo, a program that results in the synthesis of a of and which of endosperm cell and of and in the endosperm for uptake by the embryo. In several such as and the is and Olsen, Although the have been et al., et al., et al., the regulation of these has not been Recent results a key for some signal first be or by at the cell and by to the of both on the cell is by through or et al., 2007). The that the of cells is based on and that the of starchy endosperm cells and cells is not as the two cell can during development, which based on genetic and The analysis of mutants that cell and cell are two et al., on the of several in different a for cell and mutants affecting the have been in both maize and but information is of developing maize endosperm cell and the cereal endosperm is such a an of its development the of and nutrient and the regulation of and et al., In cell cell endoreduplication, and are with of and suggesting that the cell and accumulation phase is with activity and is starchy endosperm cells to suggesting that cereal endosperm cells are to synthesis at of of in to could be part of such an to accumulation et al., The from the cell division phase the storage phase of endosperm development is by of et al., et al., 2005; et al., 2007; et al., and to be by et al., and the of et al., activity to be during the early with cell proliferation, as shown by the analysis of cell wall in maize et al., and et al., Thus, a in the caryopsis is with endosperm cell proliferation, whereas a in with the the accumulation likely for the of and and the of to be a a to and in from endosperm and mutants that otherwise not on et al., because is for the of that are by including be in to and could by both and the of and The is because it to be specific to cereals and is in the In and maize, is with the development of tissue and the transfer cell et al., 2005; et al., and an in and with the the storage phase et al., by cell similar to It has been that cereals are the main source of in livestock and are the source in storage are responsible for the and of the from endosperm which are for and and the of other The storage in cereals are and in or and in although additional and Larkins, 1999; and are found in which are and and in in and in et al., and found in which maize and the in and and Larkins, 1999; et al., are in and and are generally in amino acids, in the amino and They amino from the of and in to of total endosperm in the genera and but for only to of endosperm in and in which most storage of In endosperm primarily storage that maize and and and in the of the In wheat and grasses, these are to large storage In maize and other as as the are retained within the through an mechanism and Larkins, 1999; and Larkins, The and storage in maize and are in different of and et al., and et al., and results in of on the and et al., The of within the by specific between these and mutations that the of and lead to the et al., et al., et al., et al., 2007). Although there are generally are and Larkins, et al., Sabelli and et al., and of in the endosperm, is primarily at the and the generally during and of endosperm development, to specific et al., 2001; et al., and 2007; and regulatory for first in and the later, were found in and maize et al., et al., This typically contains two the and a and 2007; et al., The with a the et al., whereas the is by such as in maize et al., which a in the of maize et al., The importance of in cereals is by the that at the a in the of maize from et al., in the which in and et al., and the wheat weight which has only with the et al., Among the the has been best characterized in maize et al., 2007). It several cell that the embryo (i.e. at around 4 DAP). As the embryo the and by early to mid endosperm development (i.e. around there are only of the at the of the endosperm. cells of the endosperm cellularization phase (Kiesselbach, 1949; Kiesselbach and and are in and with a on several cells are to be and in the embryo with primarily through an et al., 2007). Indeed, an is in the maize which in the et al., potential for the from and at the of the from at two in the and which have et al., for a of in between embryo and endosperm from the which similar to Arabidopsis and 2001; et al., The an in the embryonic in maize et al., 2007). that cells with similar to maize are in wheat and although on these are The lacks of maize et al., 2007). different of cell occur during endosperm one is which results in a the second is to cell which most cells the endosperm; and the is endoreduplication, which rounds of DNA or cytokinesis, resulting in cells. As information about the regulation of nuclear proliferation and the cellularization is and primarily whereas the two of cell have been characterized in some in phase of mitotic cell division cellularization of the endosperm and is largely responsible for the of endosperm cells. This period to in the central endosperm but approximately 20 to in the and subaleurone and Phillips, division to be in the cereal endosperm. typically occur in a first at the of the endosperm and in the central have been with to the in of the nuclei, and cells and are with cell that cell division and Phillips, The mitotic index around to and the period from to DAP, the endosperm rapidly to the seed This growth to be with cell division and as as endoreduplication the of nuclei and Phillips, From approximately to DAP, maize endosperm cells and from a mitotic to an endoreduplication cell in which and rounds of DNA synthesis and cytokinesis and Phillips, Larkins et al., 2001; Sabelli and Larkins, of the of the a in nuclear is in tissue with the nuclei and at the of the endosperm and nuclei in the central DNA nuclear and cell are and Phillips, 1988; et al., The endoreduplication cycle results in loosely and Phillips, which at the and and in cells is likely to an in the of the grass endosperm, as shown by the analysis of in which dramatic in with development of the caryopsis and Although the of endosperm nuclei is to be in most species, in wheat (Triticum it to which could result in the repression of et al., during endosperm development in cereals et al., and 1992; et al., and is with nuclear and cell the growth of the and the synthesis and accumulation of storage such as and storage Although several have been for endoreduplication in the endosperm, including a mechanism to provide to support cell and tissue growth cell and the of by the embryo during and Larkins, one of these The of different cell cycle have been such as of and and of which crucial but in cell cycle regulation et al., 2001; Sabelli et al., 2005b, 2007; and can be as or and are for the and in activity at to in maize, concomitant with the of endoreduplication, and the view that the from the mitotic to the endoreduplication cell cycle of mitotic and of and Larkins, et al., et al., et al., 2005; et al., Thus, of activity to be for the from a mitotic to an endoreduplication cell cycle during endosperm development. in endosperm development. are a family of that primarily cells from phase by the activity of which is for the of many be unique in that at two in maize and et al., Sabelli and Larkins, the of and in endosperm development are not Although early that by with other in cells et al., have shown that the of during the endoreduplication phase of endosperm development, suggesting that at some activity be et al., is by suggesting a between and and its is with mitotic activity than endoreduplication, which differences between these two et al., Sabelli and Larkins, during endoreduplication the view that activity is retained during this phase of development. that the of and the activity of key cell cycle both the cell cycle and the development of the endosperm on and which have been et al., 2005b, 2007). PCD an in cereal endosperm development, and it is to nutrient and uptake by the embryo at et al., 2007). PCD in maize starchy endosperm at around 16 in two the central starchy endosperm cells and cells the PCD from these two which that by approximately the of the endosperm is and In a similar process although in its and with endosperm cells but the PCD by 30 and Although PCD in some of the of PCD in such as DNA and nuclear the in are have not been there is for a by a of with activity et al., et al., 2007). in the and of PCD in the endosperm. both in the caryopsis and specific are with PCD et al., In PCD as shown by and PCD in maize in which the pathway is and from starchy endosperm, PCD in cells is by than In both starchy endosperm and to or PCD et al., 2007). analysis the of as as the and in endosperm PCD and during seed development et al., is about seed and are in and the by and regulation and and 2005; et al., 2007). most information the of the embryo and than the starchy endosperm. and are in these in cereals et al., et al., 2007; et al., but is far from of the in the of premature in cereals has highlighted the by the of et al., as shown by the repression of in the by the et al., from the is for endosperm and seed development Indeed, analysis that in the endosperm is to cell proliferation with endoreduplication and cell et al., et al., In an often results in or development of the transfer cell et al., of the of in endosperm development of the endosperm is to be largely genetic there is that this is the et al., Kowles et al., et al., endosperm development is the result of genetic and which are only to be in The activity of derived from the two is in the endosperm by in this is the only tissue in in which is to In maize, several are such as and et al., et al., et al., and et al., et al., et al., 2004), and et al., and are because they which are part of large through such as and et al., of other in Arabidopsis to development of the endosperm fertilization, but maize mutant has been of and is in maize, suggesting of function during endosperm development et al., et al., 2007). Although the regulation of in grasses is in its of between cell cycle regulation and endosperm and development is in Arabidopsis et al., and similar in grasses as well. Although substantial has been in cell proliferation, and the that these as as key aspects such as cell cell endoreduplication, and the accumulation of storage are dramatic and such as cellularization of the syncytium. This that and rapidly as a result of the of regulatory The of cellularization in many nuclear suggesting that a among the nuclear of many cells to the to be the endosperm do cells they to and in endoreduplication and cell growth of the endosperm with its endoreduplication the and accumulation of storage in and is it for these are and with cell proliferation, cell and the accumulation of storage is endosperm development in grasses by to the that the of different and This the that the endosperm is far from an and tissue with and the endosperm is a tissue with cell and it undergoes many of the encountered during the development of and including cell proliferation, cell and The of grasses to on the of these The grasses that have been and were for the development of human they will the and and its of the responsible for the (from a human endosperm of grasses the of which through and is and could the of cereal species to provide for the