80 LAB 07: ANGIOSPERM SYNAPOMORPHIES Introduction

LAB 07: ANGIOSPERM SYNAPOMORPHIES Introduction:

Phylogenetic relationships in the flowering plants (Cronk 2009, Fig. 1.8)

Diversity in flower morphology: A complete flower consists of sepals, petals, stamens and pistil (of the gynoecium, with one or more carpels). In many species, flowers are incomplete, lacking one or more of the four types of floral organs. Virtually all wind-pollinated flowers, for example, are unisexual and commonly lack perianth parts (petals and sepals).

Incomplete flowers. A, Pistillate flower of Salix (willow) containing only a pistil subtended by a bract, staminate flower consists only of stamens; B, Saururus cernuus (lizard’s tail) lacking both petals and sepals; C, Caltha palustris (marsh marigold) lacking petals.

The diversity of floral form often correlates tightly with pollination vectors. The suite of characters that constitute adaptations to a particular pollinator are known as pollination syndromes.

Examples of diversity in flower form and pollinating vectors. A, Salvia, pollinated by bumblebees; B, Lilium with hawkmoth pollinator; C, Arum, an inflorescence pollinated by carrion flies; D, trumpet-vine with hummingbird; E, Kigelia with bat; F, Vallisneria, a rare case of water pollination in which water conveys the entire male flowers to the female flowers.

Laboratory Exercise: Angiosperm synapomorphies I. The flower as a synapomorphy and a key innovation: Dissect at least 4 flowers, one from each lineage listed. Work on damp paper towel, using the dissecting scope and making cross/long sections of pistils. Fill in the Table below.

Lineage Basal Monocot Basal Eudicot Core Eudicot (ANITA clade) (Caryophyllid Name of Flower Or Magnolid Asterid or Rosid) Are all flower parts present? (complete or incomplete)

Symmetry: bilateral or radial

. Calyx: color, number of sepals, fused or not?

Corolla: color, number of petals, fused or not?

Stamens: color, number, fusion of filaments or anthers? fusion to petals?

Pistils: simple or compound. If compound, number of carpels (by externally visible compartmentalization of ovary; number of styles or stigmas; number of internal compartments seen in cross-section) Ovary position inferior or superior

Ovules: approximate number per pistil *In ovary long section/prepared slides.

Nectaries: absent or present, location and shape. (Eudicots: often a yellow disk at base of ovary; Monocots: within ovary, with surface exit pores)

II. The flower in evolutionary context: Observe the flowers provided, the material is organized evolutionarily, following the current understanding of angiosperm relationships as depicted in the phylogenetic tree below. Write in an example and summarize floral features on the phylogeny below for each lineage or grade.

1.) Summarize 3-4 floral characteristics that you have observed in your dissections and in lab demos that are ancestral, as observed in the ANITA grade or Magnolid dicots.

2.) Summarize 3-4 floral characteristics that you have observed in your dissections and in lab demos that are derived, as observed in the core eudicots.

III. The flower as a reproductive unit: Pollination syndromes

1-Observe plants demonstrating different types of floral specializations to pollinators. Although most gymnosperms are wind-pollinated (some Gnetophytes and Cycads being exceptions), wind-pollination in angiosperms is a derived condition. The oldest known angiosperm pollen is of the insect type. But where ecological circumstances have favored wind-pollination, such as high wind velocity and a paucity of animal pollinators, a number of angiosperm taxa have shifted from animal to wind pollination. Animal pollination is by far the most common and clearly represents an enormous increase in efficiency over the wind pollination that predominates in gymnosperms. Angiosperms are pollinated by many kinds of animals (bees, butterflies, moths, flies, birds, bats, rodents, marsupials, etc.). The wide variety of shapes, colors and “rewards” (nectar, pollen, wax, oils, scents) are part of the “pollination syndrome”, a suite of adaptations to attract a particular kind of animal to transfer pollen among different flowers. Specialists in pollination biology can usually tell what kind of animal pollinates a particular flower from its morphology. For example, big sturdy red/orange flowers with little odor are often bird- pollinated; blue/yellow flowers with nectar guides (stripes or dots on the petals, often visible only in UV light) are usually bee pollinated; blue, yellow or red flowers massed in a flat- topped inflorescence (a “landing platform”) are often butterfly pollinated; nectaries at the base of a long corolla tube are often associated with long-tongued insects such as hawkmoths or butterflies, or if the flowers are red - by hummingbirds; night-blooming flowers of white/dull color with a strong odor may be moth or bat pollinated; flowers that have foul or spicy odors, often with mottled red colors are usually beetle or carrion fly-pollinated (they mimic rotten flesh). Analyze the Table below and fill in an example for each syndrome in the last column, among the flowers available in the lab.

FLORAL CHARACTERISTICS Time of Pollinator Color Scent Corolla Reward Example Flowering Blue, yellow, Bilateral landing Nectar and /or Bee Fresh, strong Day purple platform pollen Landing platform; Bright; Butterfly Fresh, weak Day sometimes nectar Nectar only often red spurs Dissected; White or Night or Moth Sweet, strong sometimes nectar Nectar only pale dusk spurs Nectar and /or Fly (reward) Light Faint Day Radial, shallow pollen Brownish, Fly (carrion) Rotten, strong Day or night Enclosed or open None purplish Often green Nectar and /or Beetle Various, strong Day or night Enclosed or open or white pollen

Bright; Tubular or pendant; Birds None Day Nectar only often red ovary often inferior Showy flower or Nectar and /or Bat Whitish Musky, strong Night inflorescence pollen Non-flying Unscented to Robust ,exerted Copius nectar Dull-colored Night mammals variously strong styles and stamens and/or pollen

Modified from Judd et al 2002

2-Match flower scents in the scent demonstration cups to pollination syndromes in the table. Not all scents will be obvious (nor pleasant!), have fun!

ANGIOSPERM REPRODUCTION Background In the majority of angiosperm species, the ovule has two integuments. In certain derived families where only one integument is present (e.g. Asteraceae, Orchidaceae), the single integument is interpreted as a loss of one integument or fusion of two integuments. Within the ovule, the nucellus (megasporangium) of angiosperms may be considerably reduced in comparison to gymnosperms. In many taxa it consists of little more than a cell layer over the megaspore mother cell. In other taxa, there is a more substantial nucellus, but even in that case it is largely destroyed as the female gametophyte develops. Female gametophyte. In most angiosperms and gymnosperms, meiosis produces a linear array of potential megaspores. The megaspore wall of angiosperms is unique among seed plants in being thin and lacking sporopollenin. The sequence of nuclear divisions, abortions and migrations that produce the mature female gametophyte can follow over ten different developmental pathways. About 80% of the angiosperm female gametophytes follow a particular sequence of events known as the Polygonum type (a derived type, see lecture notes for ancestral gametophyte development, e.g. in basal angiosperms). The megaspore nucleus undergoes mitosis and the two nuclei migrate to opposite ends of the cell. Two subsequent mitoses produce a total of four nuclei at each end of the cell. Two nuclei, one from each group of four move toward the center of the cell and are not involved in subsequent cellularization. The other six nuclei, three at each end of the cell, develop walls. The trio of cells at the micropylar end consists of an egg flanked by two synergids. There is no hint of archegonia. The three cells at the chalazal end (near the ovule stalk) are known as antipodals and have no known function. The two nuclei that are not walled off are called polar nuclei. They lie near each other in the original megaspore cytoplasm forming the central cell. Thus, the mature female gametophyte, or embryo sac, consists of 7 cells and 8 nuclei. The female gametophyte develops very quickly and is complete before pollination occurs.

Development of a Polygonum type angiosperm ovule beginning with the formation of the integuments and single megasporocyte (A,B), continuing through the formation of megaspores (C-E), and concluding with the successive stages in development of the embryo sac (F-J). (From Gifford and Foster, 1998)

chalazal end

Male gametophyte. In 70% of angiosperm species, when pollen is released from the anther it consists of just 2 cells, the generative cell and the tube cell. When the generative cell divides, the products differentiate directly as sperm. In 30% of angiosperm species, the sperm are already present when the pollen is released, 3-celled pollen represents the most derived condition. The pollen wall of angiosperms has unique structural features that make it recognizable in the fossil record. In almost all species the outermost wall (exine) consists of columns of fused sporopollenin granules. Many gymnosperms are monosulcate. A sulcus is a slit or furrow in the exine at the distal pole of the pollen grain (the one furthest from the point of contact between microspores in a tetrad). The oldest angiosperm pollen is monosulcate also. In more derived angiosperms, pollen grains are characterized by multiple apertures (e.g. three in Eudicots) with a variety of shapes and positions. Pollen is in a dessicated condition when shed from anthers. It hydrates after being deposited on a compatible stigma. About 75% of angiosperm species are self-incompatible, due to interactions between proteins in the pollen wall and in the stigmatic cells. In self-incompatible species pollen grains fail to germinate or stop growing shortly after germination. Self-incompatibility may have evolved in angiosperms following the evolution of bisexual flowers. Bisexuality involves the risk of self-pollination leading to low genetic variability and sometimes to inbreeding depression. Self-pollination is favored in some species - such as those that are weedy or long distance migrators. Fertilization. In a compatible reaction, pollen grains germinate on the stigma and the tubes grow down through the style to the ovule. Angiosperm pollen tubes grow about two millimeters per hour, a rate about 2,000 times faster than that seen in most gymnosperms. In flowering plants only a few hours elapse between pollination and fertilization. Pollen tubes grow into the ovary and find their way to the ovules guided by signals from the synergid cells. Before the tubes reach the embryo sac (female gametophyte) one of the two synergids flanking the egg degenerates. It has been shown in several species that pollen tubes penetrate the embryo sac via the degenerating synergid. The two sperm within a pollen tube are discharged into the synergid. Although the egg and synergid are mostly walled cells, they lack walls at their chalazal ends. It is assumed that the sperm can move through the cell membrane at that end. One moves into the egg and fuses with the egg nucleus and the other moves into the central cell and there fuses with the two polar nuclei to produce the endosperm, a triploid tissue that nourishes the developing embryo. Double fertilization in angiosperms should be contrasted with that of the two Gnetophytes Ephedra and Gnetum. In both genera, one fertilization event produces supernumerary embryos that ultimately die. Nourishment of the surviving embryo is carried out by cells of the female gametophyte. In angiosperms, the female gametophyte consists of just seven cells (8 nuclei). Nourishment of the embryo in most flowering plant species is carried out by the endosperm. If fertilization fails in an angiosperm ovule, no food supply for the embryo is produced - an obvious efficiency. In Cycads and Ginkgo, the cells of the female gametophyte are fully stocked with food whether or not fertilization occurs - an obvious inefficiency. In conifers, an intermediate situation prevails. The cellular female gametophyte develops, but does not accumulate a full complement of reserve material if fertilization does not occur. Fruits. It is possible that a shift from wind to animal-pollination constituted the selection pressure that led to carpel evolution. Many seeds are good to eat and there may have been strong selection to shield ovules from visiting pollinators. As the seeds within it mature, so does the ovary wall, creating a fruit- a structure unique to the flowering plants. Fruits may function as intact dispersal units: fleshy ones often being eaten by animals that defecate the seeds at some distance, dry fruits being wind-dispersed; or fruits may split open and the seeds themselves dispersed by a variety of means.

Laboratory Exercise: angiosperm reproduction

I. Male Gametophyte (Pollen) and Pollen Tube

1. Prepare Tradescantia pollen for in vivo germination. Place pollen in a drop of pollen germination medium on a slide, add a coverslip and place in a humid chamber (petri dish lined with damp kimwipe) to avoid evaporation. Observe after about 1 hour. 2. Microsporogenesis- Observe slides of sequential development of pollen in Lilium anthers, beginning with the sporogenous tissue and ending with mature pollen (early anthers, pollen mother cells, first and second division, mature anthers and pollen tetrad).

Lilium - Microsporogenesis: A. Microsporocyte, B. End of Meiosis I. C. Tetrad of Microspores.

3. Observe slides of germinating Lilium pollen on stigma.

II. Female Gametophyte (Embryo Sac) There are 11 recognized types of female gametophyte development! They can be grouped into three major categories, depending on the number of megaspores that participate in the formation of the embryo sac: monosporic, bisporic and tetrasporic. Review the typical Polygonum type in the introductory notes (monosporic, follow the diagrams), then observe slides of the Lilium type (tetrasporic). Study the following sequence in the slides provided. Draw and label the stages in the space provided: a) megasporocyte, Identify integuments and nucellus.

b) mature embryo sac: three antipodal cells, two synergid cells, one egg nucleus and one central cell,

c) fertilization, look for evidence of pollen tube intrusion into embryo sac, d) young embryo with endosperm. What is the ploidy level of the endosperm?______e) mature seed.

III. The Fruit: Use the attached key to identify different types of fruits available in lab.

Key To Fruit Types

1a. Fruit derived from several ovaries of one or more flowers

2a. Fruit arising from the several ovaries of as many flowers (examples: pineapple, mulberry)

MULTIPLE FRUIT

2b. Fruit arising from the coalescence of several ripened ovaries of one flower (example: raspberry, blackberry)

AGGREGATE FRUIT

1b. Fruit derived from a single ovary (simple or compound)

3a. Fruit fleshy or juicy when ripe

4a. Ovary wall of fruit (or pericarp) entirely or in part fleshy

5a. Fruit indehiscent

6a. Ovary wall entirely fleshy (examples: tomato, cranberry, grape, currant, banana, melon [pepo], and citrus fruit [hesperidium])

BERRY

6b. Ovary wall of three distinct layers, the inner one bony (endocarp), the middle fleshy (mesocarp), and the outer "skin- like" (exocarp) (examples: peach, plum, cherry)

DRUPE

5b. Fruit dehiscent

7a. Fruit derived from one carpel FOLLICLE

7b. Fruit derived from a compound gynoecium CAPSULE

4b. Ovary wall (e.g., the outer layer of an apple 'core') of fruit papery, surrounded by a fleshy material that represents the coalescent parts of the stamens, petals, sepals, and (some believe) receptacle (examples:

apple, pear, quince)

POME

3b. Fruit typically dry and usually hardened when ripe

8a. Fruit indehiscent (does not open or dehisce when mature), generally with one seed

9a. Ovary wall of varying thickness, usually not bony

10a. Fruit not winged (examples: buttercup, 'seeds' of strawberry, sunflower family, sedges, grasses [ovary wall adherent to and surrounding seed, may be called caryopsis or grain])

ACHENE

10b. Fruit winged (examples: elm, tulip tree)

SAMARA

9b. Ovary wall hardened and bony

11a. Fruit usually > 5mm long (examples: oak, chestnut, hazelnut)

NUT

11b. Fruit small, usually < 5mm long (examples: borage and mint families [Boraginaceae and Lamiaceae]

NUTLET

8b. Fruit dehiscent (opens or dehisces when mature, usually along certain definite lines or sutures), with one or more seeds

12a. Fruit derived from a single carpel

13a. Fruit dehiscing along one side (examples: columbine, larkspur, magnolia, milkweed)

FOLLICLE

13b. Fruit dehiscing along two sides or breaking crosswise into one- seeded segments

14a. Fruit dehiscing along two sides (example: only the legume family [Fabaceae or Leguminosae])

LEGUMES

14b. Fruit breaking into one- seeded segments (example: only the legume family [Fabaceae or Leguminosae]

LOMENT

12b. Fruit derived from a compound gynoecium of two or more carpels (types of capsules)

15a. Fruit always 2-carpellate, two-celled, and with parietal placentation

16a. Fruit > 2-3 times longer than wide (example: only the mustard family [Brassicaceae or Cruciferae])

SILIQUE

16b. Fruit <2-3 times longer than wide (example: only the mustard family [Brassicaceae or Cruciferae])

SILICLE

15b. Fruit 2 or more carpellate, one or more celled, and with various types of placentation. CAPSULE

17a. Fruit dehiscing by pores (poricidal dehiscence; example: poppy)

PORICIDAL CAPSULE

17b. Fruit dehiscing along the septa or into the locules or by a lid.

18a. Fruit dehiscing by a lid (examples: Portulacaceae and some Caryophyllaceae)

CIRCUMSCISSILE CAPSULE

18b. Fruit dehiscing directly into the locules or along the septa.

19a. Fruit dehiscing directly into the locules (examples: iris, phlox, pyrola, violet, waterleaf)

LOCULICIDAL CAPSULE

19b. Fruit dehiscing along the septa

20a. Fruit dehiscing to form 1-seeded segments called mericarps (examples: carrot, maple, spurge)

SCHIZOCARPOUS CAPSULE or SCHIZOCARP

20b. Fruit dehiscing to form several-seeded segments (examples: peppers, figwort, rhododendron))

SEPTICIDAL CAPSULE