The influence of tetrad shape and intersporal callose wall formation on pollen aperture pattern ontogeny in two eudicot species

Abstract

Background and aims: In flowering plants, microsporogenesis is accompanied by various types of cytoplasmic partitioning (cytokinesis). Patterns of male cytokinesis are suspected to play a role in the diversity of aperture patterns found in pollen grains of angiosperms. The relationships between intersporal wall formation, tetrad shape and pollen aperture pattern ontogeny are studied.

Methods: A comparative analysis of meiosis and aperture distribution was performed within tetrads in two triporate eudicot species with contrasting aperture arrangements within their tetrads [Epilobium roseum (Onagraceae) and Paranomus reflexus (Proteaceae)].

Key results and conclusions: Intersporal wall formation is a two-step process in both species. Cytokinesis is first achieved by the formation of naked centripetal cell plates. These naked cell plates are then covered by additional thick, localized callose deposits that differ in location between the two species. Apertures are finally formed in areas in which additional callose is deposited on the cell plates. The recorded variation in tetrad shape is correlated with variations in aperture pattern, demonstrating the role of cell partitioning in aperture pattern ontogeny.

Fig. 1.

Simultaneous cytokinesis and aperture arrangements within tetrads: (A) microsporocytes enclosed within a thick callose wall before meiosis; (B) and (C) simultaneous cytokinesis in which both nuclear divisions occur before cytokinesis; (D) microspores are separated through the formation of intersporal callose walls. N, Nucleus, cy, cytoplasm; ca, callose. (E) Arrangement of apertures within tetrads: in Fischer's, apertures meet in six pairs; in Garside's, apertures meet in four groups of three.

Fig. 2.

Intersporal wall formation, tetrad shapes and aperture arrangements within tetrads of Epilobium roseum: (A) simultaneous cytokinesis with centripetal cell-plate formation; (B) tetrahedral tetrad soon after the completion of the cell plates (the asterisk indicates the last points of contact between the cytoplasm of the newly formed microspores); (C) tetrahedral tetrad during additional callose deposition with additional callose in the centre (arrows indicate places where additional callose has not been deposited; (D) Lower (a), middle (b) and upper (c) focus on a tetrahedral tetrad with thick callose deposits in the centre (arrows) after the completion of intersporal wall formation; (E) tetrahedral tetrad (a) during pollen maturation and (b) shown as a schematic representation of pore arrangement (black dots) and pollen grains (large grey discs); (F) tetragonal tetrad during additional callose deposition with additional callose in the centre (arrows indicate places where additional callose has not been deposited); (G) Lower (a), middle (b) and upper (c) focus on a tetragonal tetrad with thick callose deposits in the centre (arrows) after the completion of intersporal wall formation; (H) tetragonal tetrad (a) during pollen maturation and (b) shown as a schematic representations of pore arrangement (black dots) and pollen grains (large grey discs); (I) rhomboidal tetrad soon after the completion of cell-plate formation (asterisks indicate the last points of contact between the cytoplasms of the newly formed microspores); (J) rhomboidal tetrad during additional callose deposition with additional callose in the centre (arrows indicate places where additional callose has not been deposited); (K) lower (a), middle (b) and upper (c) focus on a rhomboidal tetrad at the late tetrad stage – at the lower focus, callose deposits are thicker, while they are narrower at the higher focus (arrows); (L) rhomboidal tetrad (a) during pollen maturation and (b) in a schematic representation of pore arrangement (black dots) and pollen grains (large grey discs). The stain used in (A)–(E,a), (F)–(H,a) and (I)–(L,a) is anilin blue. Scale bars = 10 µm.

Fig. 3.

Intersporal wall formation, tetrad shape, aperture arrangement within tetrads and pollen grains of Paranomus reflexus: (A) simultaneous cytokinesis with centripetal cell-plate formation (the asterisk indicates the last points of contact between the cytoplasm of the newly formed microspores); (B) tetrahedral tetrad soon after the completion of the cell plates; (C) lower (a) and upper (b) focus on an intermediate tetrad (see text) with unequal cleavage walls – notice that in the upper focus, the top external microspore appears to extend towards the bottom external microspore, resulting in a tetrad which is not strictly rhomboidal; (D) tetragonal tetrad with cell plates embedded in additional thick callose deposits (arrows indicate places where callose has not been deposited); (E) tetragonal tetrad after completion of the intersporal wall (a), shown as a schematic representation in top view in (b) (the aperture position was inferred from the pollen grain shape, and the position of the pollen grains within the tetrad, see Discussion) – the apertures (black areas) are located at the extremities of the elongated microspores (grey); (F) lower (a) and upper (b) focus on a tetrahedral tetrad with cell plates partially covered with additional callose deposits; (G) tetrahedral tetrad with additional thick callose deposits (arrows indicate places where callose has not been deposited); (H) lower (a) and upper (b) focus on a tetrahedral tetrad after the completion of the intersporal wall formation – callose has been deposited in the places where cell plates intersect in threes (arrow). (I) Tetrahedral tetrad after the completion of intersporal wall formation shown in lower (a), middle (b) and upper (c) focus – microspores are triangular and their vertices meet in four groups of three in the places where three cell plates intersect – and in a schematic representation (d) in which the pores (black areas) are placed at the vertex of the triangular microspores (grey) and arranged in four groups of three pores. (J,a–c) An intermediate tetrad after the completion of intersporal formation shown in lower (a), middle (b) and upper (c) focus – in the lower view the vertices of four microspores meet to form a single group, while in the upper view the microspore vertices meet in two groups of three. The microspores are separated by heavy additional callose deposits that decrease in thickness towards the intersections of the intersporal walls. (J,d) Schematic representation composed of two kinds of pollen grains (grey): elongated ones along the sides (left and right), and triangular ones at the top and bottom. Pores (black areas) are arranged in a single group of four at the bottom and two groups of three at the top. (K–M) Developing pollen grains – the pores appear as red areas at the vertices of the pollen grain: (K) triporate triangular pollen; (L) elongate diporate pollen grain; (M) quadrangular tetraporate pollen. (N) Atypical tetrad with a tetraporate pollen grain. Transmission light microscopy was used in (A)–(D), (F)–(H) and (J,a–c) with anilin blue staining. Light microscopy was used in (Ea), (I,a–c) and (K)–(N) with congo red staining. Scale bars = 10 µm.