Proc. Nati. Acad. Sci. USA Vol. 76, No. 7, pp. 3358-3362, July 1979 Cell Biology

Pollen- interactions: Identification and characterization of surface components with recognition potential (cell surface receptors/ /adhesion/concanavalin A/tridacnin) ADRIENNE CLARKE*, PAUL GLEESON*t, SUSAN HARRISON*, AND R. BRUCE KNOX* *School of Botany, University of Melbourne, Parkville, Victoria 3052, Australia; and tDivision of Chemistry, Commonwealth Scientific and Industrial Organization, Parkville, Victoria 3052, Australia Communicated by Sir R. N. Robertson, April 5, 1979 ABSTIRACT Male-female recognition in flowering MATERIAL AND METHODS is initiated by mutual contact of and stigma surface components. Analysis of the surface macromolecules of both Cut flowers of Gladiolus gandavensis were maintained as de- stigma and pollen of Gladiolus gandavensis revealed a complex scribed (10). Pollen was collected and surface extracts were mixture of , glycoproteins, and glycolipids. The carbo- prepared in Tris/saline pH 7.4 buffer as described (10) (yield: hydrate-containing components amounted to 6% in pollen and 15 the pollen grains gave a 23% in stigma and contained the monosaccharides , mg/g of pollen). After extraction, , glucose, , and . All the mannose positive test with fluoroscein diacetate, indicating that the of both preparations was associated with a fraction that bound plasma membrane was intact. Stigma surface extracts were also to concanavalin A. The stigma surface contained an arabino- prepared in Tris/saline pH 7.4 buffer and 0.5% sodium deox- or protein as a major component. This ycholate as described (10) (yield: buffer extract, 0.2 mg/100 component has been isolated by affinity chromatography on stigmas; sodium deoxycholate extract, 10 mg/100 stigmas). tridacnin-Sepharose and shown to be similar in composition to Cytoplasmic streaming in the stigma was evident after removal a style canal component isolated in the same way. The capacity of the surface layer, so the plasma membrane was intact. of the stigma surface preparations to bind nonspecifically to of the and macromolecules from pollen and other sources has been dem- Analysis. Samples pollen stigma onstrated in vivo and in vitro. Specific binding of concanavalin extracts were hydrolyzed for 2 hr at 100'C in 2.5 M trifluo- A to the stigma surface decreases the adhesive capacity for roacetic acid. The released monosaccharides were reduced to pollen protein. The arabinogalactan of the stigma surface may the corresponding alditols, acetylated (14), and separated by act as an adhesive base. The pollen and stigma surfaces appar- gas/liquid chromatography on 100/200 Supelcoport coated ently complement one another to provide all the components with 3% SP 2340 (Supelco Inc, Bellefonte, PA). of an ideal adhesive. Electrophoresis. Sodium dodecyl sulfate/polyacrylamide gels were run according to Laemmli (15); they were stained is essential for the of life. This with Coomassie blue for protein, periodate-Schiff stain for Cell recognition continuity plant carbohydrate, and oil red 0 for lipid. Gels were also stained is well documented during sexual reproduction in flowering with Yariv artificial f3-glycosyl antigen by covering the gel with plants, when the receptive cells of the female stigma discrim- a solution of the antigen (1 mg/ml in dimethyl sulfoxide diluted inate between compatible and foreign (1-3). The rec- 1:10 with Tris/saline buffer), allowing it to stand at room ognition events are believed to be initiated by specific binding temperature for 24 hr, and then washing it with Tris/saline between pollen and stigma surface components (3). Information buffer. Electrophoresis on cellulose acetate strips was done in on the biochemical nature of the interacting surface compo- barbitone buffer (pH 8.8; I = 0.05), for 40 min at 4 mA/cm and nents is incomplete. Pollen sites have been shown cytochemi- the strips were stained by the same method. cally to contain various enzymes, antigens, and, in some species, Fractionation of Concanavalin A (Con A)-Binding Com- allergens (4). The materials diffusing from pollen also include ponents of Pollen and Stigma Surfaces. Con A-binding ma- low molecular weight , lipids, and pigments (5). terial from buffer extracts of pollen and stigmas was isolated For the stigma, characterization has been restricted to cyto- by affinity chromatography on Con A-Sepharose (Pharmacia) chemical detection of surface esterases (1, 6), antigens (7, 8), and 0.1 M a-methylmannoside as eluant. and lectin-binding sites (9, 10). A component binding to the Fractionation of Stigma Surface Preparation on Tridac- Yariv f3-glucosyl artificial carbohydrate antigen has also been nin-Sepharose. Specimens of Tridacna maxima, the small giant demonstrated cytochemically (11); this artificial antigen has clam, were collected from the Heron Island Research Station been shown to bind specifically to arabinogalactan proteins in (Queensland, Australia), and the galactose-binding lectin a wide range of plant tissues (12, 13). We now present an tridacnin was isolated from the hemolymph by affinity chro- analysis of the pollen and stigma surfaces as a basis for estab- matography on acid-treated Sepharose 6B (16) and coupled to lishing the components in which the specific binding properties Sepharose 4B. The stigma or style extract was applied directly may be vested. to the column in the presence of 10 mM CaCl2 and eluted with Gladiolus provides a model system in which the events of a calcium-free buffer (17). compatible pollination can be approached experimentally (10); Identity of in Pollen and Stigma Surface it has large accessible anthers and stigmas, abundant pollen, and Preparations. Double diffusion and immunoelectrophoresis the great advantage of year-round commercial availability. were carried out by the microslide method (Gelman Instrument Co., Ann Arbor, MI). The artificial antigen solutions were used at 1 mg/ml in dimethyl sulfoxide diluted 1:10 with Tris/saline The publication costs of this article were defrayed in part by page the and and charge payment. This article must therefore be hereby marked "ad- buffer; stigma pollen extracts, lectins, arabinoga- vertisement" in accordance with 18 U.S. C. §1734 solely to indicate this fact. Abbreviation: Con A, concanavalin A. 3358 Downloaded by guest on September 24, 2021 Cell Biology: Clarke et al. Proc. Natl. Acad. Sci. USA 76 (1979) 3359

lactan proteins were used at 10 mg/ml. The peanut lectin was Table 2. Monosaccharide composition of pollen and stigma from Industrie Biologique Franqaise (Clichy, France). The surface macromolecule preparations mouse myeloma J539 was grown by Rod Ceredig at the Walter Monosaccharide* Pollen wall Stigma surface and Eliza Hall Institute of Medical Research (Melbourne, Australia). Galactose 25 59 In Vivo Binding of Pollen Surface Components and Lec- Mannose 24 1 tins to Stigmas. Groups of 10 stigmas were briefly washed and Glucose 15t 11 incubated with the appropriate solution for 5 min, washed Arabinose 13 22 again, and assayed for radioactivity in a deep-well gamma Rhamnose 23 7 counter as described (10). The unlabeled protein solutions were Results are expressed as percentage (by weight) of each monosac- used at 5 mg/ml; the 125I-labeled protein solutions were diluted charide. to 6 X 106 cpm/ml before use. Binding of fluorescein isothio- * The same monosaccharides were detected in the dialysates of both cyanate-tridacnin (1.5 mg/ml) was carried out for 30 min and preparations. t was followed by buffer washing and examination by reflected Glucose was variable in different preparations. light fluorescent microscopy. and were in the Mr range 10,000-120,000. The Mr 10,000 In Vitro Binding of Stigma Surface and Pollen Wall component, like that in the pollen wall preparation, also stained Components. The possibility of specific binding between pollen for lipid. In addition, a diffuse band at Mr 120,000 stained for wall and stigma surface components was investigated by sep- carbohydrate but not protein, and this band also stained with arating and immobilizing the pollen wall components by im- the /3-glucosyl but not the a-galactosyl artificial carbohydrate munoelectrophoresis. After diffusion, the gels were washed in antigen, indicating the presence of an arabinogalactan or ara- buffered saline (14 hr at room temperature), and strips of binogalactan protein. Whatman no. 1 filter paper were laid over the precipitin bands Fractionation of Pollen Wall and Stigma Surface Prepa- on each side of the trough. 125I-Labeled stigma extract (5 ,ug) rations. The pollen and stigma surface preparations were was layered onto each of the filter paper strips and incubated fractionated on Con A-Sepharose. The Con A-bound fraction contained mannose as the in a moist chamber overnight at room temperature. The filter of the pollen preparation major with smaller amounts of galactose and glucose paper strips were then removed and the gels were washed for monosaccharide also present. The fraction not bound to Con A contained ga- 72 hr in buffered saline. The gels were then washed with water, lactose, glucose, arabinose, and rhamnose (Table 3). For the stained with Coomassie blue, and overlaid with strips of x-ray stigma surface preparation, mannose was also the major mo- film. The radioautographs were developed after 14-28 days. nosaccharide in the Con A-bound fraction; smaller proportions RESULTS of galactose, glucose, and arabinose were also present. In each case, all of the mannose was retained on Con A-Sepharose. Analysis of Pollen Wall and Stigma Surface Macromole- cules. Both preparations contained protein, carbohydrate, and Pollen a small amount of lipid (Table 1). The monosaccharide com- 70,000 50,000 22,000 position of the preparations is shown in Table 2. Pollen prepa- rations contained galactose, mannose, and rhamnose in ap- a 1 11111III1111II 1 1111 proximately equal amounts; arabinose and glucose were present in lower concentrations. The monosaccharide distributions of different Gladiolus pollen preparations were similar, with the exception of glucose which varied from ,10% to 20% of the bL II I total carbohydrate. The stigma surface preparation contained the same monosaccharides, but galactose was present as the major monosaccharide. Arabinose, glucose, and rhamnose were also present, with mannose as a minor constituent. cl I The complexity of the pollen wall and stigma surface prep- Stigma arations was revealed by sodium dodecyl sulfate gel electro- phoresis. Both preparations showed a number of protein- 70,000 50,000 22,000 staining components in the Mr range 10,000-120,000 (Fig. 1). Of the pollen wall components, five stained for carbohydrate. d I I 1 1I These components were in the Mr range 10,000-75,000; the Mr 10,000 component also stained for lipid. No staining of any of the bands with f-glucosyl artificial antigen was detected. Of the stigma surface components, nine stained for carbohydrate e li I 11 I 11 Table 1. Estimated protein, carbohydrate, and lipid content of e pollen wall and stigma surface preparations f Content, mg/100 mg dry weight Pollen wall Stigma surface* I Protein 10 20 Carbohydrate 6 23 Lipid 0.2 0.1 FIG. 1. Sodium dodecyl sulfate/polyacrylamide gel electropho- Total protein was measured with bovine serum albumin as stan- resis of pollen (a-c) and stigma (d-g) surface diffusates of Gladiolus. dard; total carbohydrate was measured with galactose as standard; (a and d) Stained for proteins with Coomassie blue; (b and e) stained lipid is weight of material extracted into chloroform/methanol, 2:1 for carbohydrate with periodic acid-Schiff reaction; (c and f) stained (vol/vol). for lipid with oil redO; (g) stained for arabinogalactan proteins with * These analyses were performed on sodium deoxycholate extracts the f-glucosyl artificial carbohydrate antigen. The positions of three of receptive stigmas. M, markers are indicated by arrows. Downloaded by guest on September 24, 2021 3360 Cell Biology: Clarke et al. Proc. Natl. Acad. Sci. USA 76 (1979) Table 3. Monosaccharide composition of pollen and stigma brilliant surface fluorescence when treated with this labeled surface preparations after fractionation on Con A-Sepharose tridacnin, which was not observed in the presence of 0.1 M Pollen Stigma surface lactose (Fig. 2). Analysis of the stigma surface bound to tri- Fraction Fraction Fraction Fraction dacnin-Sepharose showed that galactose was the major mono- Mono- Con A- not Con A- Con A- not Con A- saccharide and that small amounts of arabinose were present saccharide bound bound bound bound (Table 5). This arabinogalactan represented about 20% of the Galactose 15 38 13 67 total stigma surface preparation on a dry weight basis. Mannose 70 0 64 0 A style component was isolated in the same way, the mono- Glucose 15 21 8 28 saccharide composition of this preparation was similar to that Arabinose 0 29 15 5 of the stigma surface and was associated with protein (3%). Rhamnose 0 12* 0 0 Material precipitated from the style extract by the f3-glucosyl antigen gave a monosaccharide analysis similar to that of the Results are expressed as percentage (by weight) of each monosac- material bound to tridacnin-Sepharose (Table 5). The similarity charide. * Peak poorly resolved. Both the pollen and stigma surface preparations were ex- tracted with chloroform/methanol, 2:1 (vol/vol). The lipid fraction obtained accounted for 0.2 and 0.1% of the total dry weight of the pollen wall and stigma surface preparations, re- spectively (Table 1). The major carbohydrate associated with the fraction was glucose for both preparations (Table 4). On further analysis, the lipid fraction of the stigma surface prep- aration was found to have a low level of organically bound phosphate (3 ng/mg of stigma extract). Chromatography in chloroform/methanol/water, 65:25:4 (vol/vol), showed four well-separated components, each of which stained with a- naphthol, indicating the presence of carbohydrate, but none corresponded to either mono- or digalactosyl diglycerides. Isolation and Characterization of Stigma Surface Arabi- nogalactan Components. The stigma surface preparation gave a precipitin band in double diffusion tests with a number of galactose-binding macromolecules: tridacnin, peanut lectin, and the IgA protein of mouse myeloma J539. The material also bound to the f3-glucosyl artificial carbohydrate antigen which is known to interact with arabinogalactans and arabinogalactan proteins. A single red precipitin band was obtained with the f-glucosyl but not the a-galactosyl artificial carbohydrate an- tigen. Identical precipitin reactions were obtained with char- acterized arabinogalactan protein from pea, walnut, and carrot and the arbinogalactan protein from Gladiolus style canal, as well as with the fraction from the stigma surface not bound to Con A. The fraction that bound to Con A did not react with the 3-glucosyl artificial carbohydrate antigen. These results indicate that the stigma surface preparation contains a macromolecular component having accessible ter- minal galactosyl residues in the fraction that is not bound to Con A. The interaction of the preparation with the 3-glucosyl an- tigen indicated the presence of an arabinogalactan component, and its interaction with the lectin tridacnin suggested that it could be isolated by lectin affinity chromatography. Cyto- chemical evidence confirming the surface location of this ar- abinogalactan component was obtained using fluoroscein iso- thiocyanate-labeled tridacnin. Stigmatic papillae showed a Table 4. Monosaccharide composition of lipid fractions of pollen and stigma surface preparations Stigma surface preparation Mono- Pollen (sodium deoxycholate saccharide preparation extract) Galactose 8 0 Mannose 8 0 Glucose 84 85 FIG. 2. Binding of fluoroscein isothiocyanate-tridacnin to surface Arabinose 0 5 of stigmatic papillae of Gladiolus. (A) Appearance of untreated Rhamnose 0 10 papillae by scanning electron microscopy. (B) Fluorescence of surface after treatment with labeled tridacnin. (C) Absence of fluorescence Results are expressed as percentage (by weight) of each monosac- after treatment with labeled tridacnin in presence of 0.1 M lactose. charide in a chloroform/methanol, 2:1 extract (vol/vol). Bar= 100,m. Downloaded by guest on September 24, 2021 Cell Biology: Clarke et al. Proc. Natl. Acad. Sci. USA 76 (1979) 3361 Table 5. Stigma and style arabinogalactans The first function of the receptive female stigma is to trap Material bound to pollen. To perform this function it must present an adhesive tridacnin-Sepharose Style material surface so that pollen carried by a pollinating insect or wind- Stigma precipitated by borne pollen will be effectively captured. After capture, a series Monosaccharide surface Style fl-glucosyl antigen of interactive events may be initiated, the first of which is hy- dration of the pollen grain and release of wall proteins that bind Galactose 76.1 85.8 82.4 to receptors on the stigma surface (1, 2). Arabinose 20.0 14.2 17.6 The surface preparations we have examined represent buf- Glucose 4.0 Trace * fer-soluble extracellular material. The plasma membrane of Galactose/arabinose both the pollen protoplast and the stigmatic papillae were ratio 3.8 6.0 4.7 shown to be intact after the extraction procedure so that cyto- Results are expressed as percentage (by weight) of each monosac- plasmic components were excluded from the preparations. We charide, except for ratio. have demonstrated the presence of a complex array of proteins, * Not available due to presence of glucose in precipitating antigen. glycoproteins, and glycolipids at the stigma surface and from the pollen wall by sodium dodecyl sulfate/polyacrylamide gel of the stigma and style arabinogalactan components was also electrophoresis. The components that stained for both protein shown by cellulose acetate electrophoresis: both samples gave and carbohydrate (5 for the pollen preparation, 7 for the stigma single bands that stained with f3-glucosyl but not a-galactosyl preparation) are likely to be glycoproteins, and the low mo- artificial antigens. lecular weight component (Mr 10,000) of both preparations Adhesive Properties of Stigma Surface. Whole excised which stained for carbohydrate and lipid is likely to be gly- Gladiolus pistils bound 125I-labeled surface preparations from colipid in nature. This array of stigma surface Gladiolus pollen (mean i SD, 915 i 120 cpm per stigma) as glycoproteins well as pollen extracts from a related genus Crocosmia (1070 corresponds in number and molecular weight to those shown i 90 cpm per stigma) and from an unrelated genus Dianthus to bind to Con A (10). (1416 ± 172 cpm per stigma). Pretreatment of stigmas with The localization of these components at the unlabeled Gladiolus pollen surface preparation effectively stigma surface and within the pollen wall sites has been dem- diminished the binding as did pretreatment with the lectin Con onstrated by their binding to fluoroscein isothiocyanate-con- A. This effect was not observed when Con A was applied in the jugated Con A (unpublished data). These glycoproteins have presence of its complementary sugar ligand a-methylman- some of the same characteristics as those from animal cell sur- noside (Table 6). faces; all of the mannose of both the pollen and stigma prepa- In vitro binding of stigma and pollen surface components rations is bound to Con A, indicating its presence either as in- was shown by immobilizing pollen wall antigens in precipitin ternal 2-O-mannosyl residues or as terminal nonreducing arcs by immunoelectrophoresis and then applying 125I-labeled groups. In both preparations, the Con A-bound material con- stigma extract. Radioautography revealed that the labeled tained glucose and galactose as well as mannose. Galactose is material was bound to all pollen-antipollen precipitin arcs. The a common constituent of both animal and plant glycoproteins material also bound to a human IgG-rabbit anti-human IgG (20); glucose is not commonly found associated with glyco- precipitin arc, demonstrating the nonspecific adhesive prop- proteins, and the variable quantity of glucose found might be erties of the stigma surface preparation. Surface stigma extracts explained in terms of extracellular oligosaccharide in the pollen from Hibiscus, Zea, and Helianthus also showed the same ca- grains. pacity to bind to a range of pollen-antipollen precipitin arcs The chloroform/methanol-soluble lipid fraction of both when tested in the same manner. preparations contained glucose as the major monosaccharide. This observation, taken in conjunction with the very low content DISCUSSION of organically bound phosphate and the absence of both mono- Recognition reactions between animal cells and between cells and digalactosyl diglycerides, suggests that the glycolipid of lower plants are believed to be mediated by cell surface component might be a glucoside such as a sterol glucoside. proteins and glycoproteins (18, 19). In flowering plants, both The coincident staining of a high molecular weight stigma somatic and sexual recognition systems are likely to be based surface component with the 3-glucosyl artificial antigen and on similar types of molecular interaction (2). The interacting with periodate-Schiff reagent in sodium dodecyl sulfate/ surfaces during sexual recognition are those of the stigma and polyacrylamide gels indicates the presence of an arabinoga- the pollen grain. The stigma becomes receptive to compatible lactan or arabinogalactan protein. This was confirmed by iso- pollen after the flower opens, when material is secreted over lation of an arabinogalactan component by affinity chroma- the outer surface of the stigma cuticle. This is not a classic bi- tography on tridacnin-Sepharose. This component is apparently layered membrane in its ultrastructural appearance (2) but it related structurally to the arabinogalactan protein isolated from can be compared with a membrane in its involvement in an the style canal of Gladiolus (17). The monosaccharide analyses interactive reaction. (Table 5) and the electrophoretic behavior of both preparations were similar: they reacted in an apparently identical way with Table 6. Binding of Gladiolus pollen surface preparations to a number of galactose binding macromolecules, and they both stigma surfaces stained with the f3-glucosyl artificial antigens. However, they differed in the ratio of the monosaccharide components. 1251-Labeled Gladiolus Isolation of sufficient arabinogalactan from the stigma sur- pollen preparation, face for methylation analysis (1 ,ug per stigma) is difficult, but Stigma treatment cpm bound/stigma* the style canal material can be obtained more readily (150 ,ug None 915 i 120 per style) and has been shown to have a structure consistent with Unlabeled Gladiolus a 1,3-f-galactan backbone with side branches linked through pollen extract 362 + 45 C (0) 6 to the backbone galactose residues. The side branches Con A 549 + 51 are 1,6-linked galactose residues, some of which carry terminal Con A + a-methylmannoside 987 i 141 a-L-arabinofuranose residues (21). Protein is associated with * Mean + SD. the style arabinogalactan, but the nature of the linkage has not Downloaded by guest on September 24, 2021 336200 Cell Biology: Clarke et al. Proc. Nati. Acad. Sci. USA 76 (1979) been demonstrated. However, it has been established for a The parallels between the pollen-stigma and plant-host number of other arabinogalactan proteins (13). The polysac- pathogen interactive systems have been drawn by Pegg (26). charide component of the Lilium longiflorum stigmatic ex- Both depend on initial adhesion and recognition that initiates udate has been shown to contain an arabino-3,6-galactan growth of the invasive or fungal hypha. The possible component but differs from the Gladiolus stigma material in molecular basis for the adhesive stage of the host-pathogen having a higher arabinose content as well as rhamnose and interactions have recently been considered, and it appears that glucuronic acid as major components (22). both specific or lectin-mediated adhesion (27, 28) and non- The arabinogalactans isolated are major components of both specific glycoprotein-glycoprotein interactions (29) may be stigma and style-they represent about 20% of the total stigma involved. The same situation may be true in the pollen-stigma surface preparation and 40% of the total soluble material of the interaction, with some surface components participating in style. The secretions of both the style canal and the stigma pollen grain capture and adhesion and others being involved surface appear as the flower matures and becomes receptive. in the information exchange which initiates the cascade of Both the papillae and the secretory cells of the style canal are events. derived from epidermal cells (23), so that the structural simi- pollination larity of the secretory product is consistent with their common We gratefully acknowledge the generous gift of purified pea, walnut, developmental origin. However, in the two situations the ara- and carrot arabinogalactan proteins and artificial carbohydrate anti- binogalactan may fulfill different roles: the style canal material gensfrom Dr. M. A. Jermyn. We are indebted to Dr. Andrew Sinclair of L. longiflorum has been shown to provide nutrient for the for the lipid analyses and to Professor B. A. Stone and Dr. M. A. Jermyn growing pollen tube (24), and in addition it seems likely that for helpful discussions. This work was supported by a grant from the at the stigma surface it contributes to the general adhesive Australian Research Grants Committee. properties of the surface. General Adhesive Properties of the Stigma Surface. The 1. Heslop-Harrison, J. (1978) Proc. R. Soc. Ser. B 202,73-92. "sticky" nature of the receptive stigma surface has been shown 2. Clarke, A. E. & Knox, R. B. (1978) Q. Rev. Biol. 53,3-28. nonself 3. Nettancourt, D. de (1977) Incompatibility in Angiosperms by general binding of self and pollen preparations. (Springer, Berlin). Proteins and glycoproteins of animal origin also bind to variable 4. Knox, R. B., Heslop-Harrison, J. & Heslop-Harrison, Y. (1975) extents (10). The surface is apparently generally sticky, but in The Biology of the Male Gamete, eds. Duckett, J. G. & Racey, there is a defined capacity for adhesion demonstrated by de- P. A. (Academic, London), pp. 177-187. creased binding of the labeled pollen preparation after pre- 5. Stanley, R. G. & Linskens, H. F. (1974) Pollen: Biology Bio- treatment of the stigma with unlabeled pollen preparation. Con chemistry, Management (Springer, Berlin). 6. Mattsson, O., Knox, R. B., Heslop-Harrison, J. & Heslop-Harrison, A has previously been shown to bind specifically to stigma Y. (1974) Nature (London) 247,298-300. surface receptors (10), and this binding is now shown to decrease 7. Nasrallah, M. E. & Wallace, D. H. (1967) Heredity 22, 519- the adhesive capacity for pollen protein; the capacity is restored 527. when Con A is applied in the presence of its complementary 8. Nasrallah, M. E., Barber, J. T. & Wallace, D. H. (1970) Heredity ligand, suggesting that specific binding of Con A alters the to- 25,23-27. 9. Heslop-Harrison, Y. (1976) Micron 7,33-36. pography of the receptive surface in some way which results 10. Knox, R. B., Clarke, A. E., Harrison, S., Smith, P. & Marchalonis, in a less-ideal contact surface for general adhesion. This implies J. J. (1976) Proc. NatI. Acad. Scd. USA 73,2788-2792. that the stigma surface must be an efficient adhesive as well as 11. Clarke, A. E., Gleeson, P. A., Jermyn, M. A. & Knox, R. B. (1978) a carrier of receptors for pollen recognition. Aust. J. Plant Physiol. 5,573-588. we have identified on the 12. Jermyn, M. A. & Yeow, Y. M. (1975) Aust. J. Plant Physiol. 2, The groups of components that 501-531. stigma surface can be considered in terms of the classes of 13. Clarke, A. E., Anderson, R. & Stone, B. A. (1979) Phytochemlstry compounds that make up an ideal synthetic adhesive (25). 18,521-540. These are an adhesive base (which is usually a high molecular 14. Albersheim, P., Nevins, D. J., English, P. D. & Karr, A. (1967) weight branched polymer), a plasticizer such as glycerol or Carbohyd. Res. 5,340-345. monosaccharides the adhesive from 15. Laemmli, U. K. (1970) Nature (London) 227,680-682. (to prevent becoming 16. Baldo, B. A. & Uhlenbruck G. (1975) in Advances in Experi- brittle), a thickener (to increase the viscosity of the final com- mental Medicine and Biology, eds. Hildemann, W. M. & Ben- pound), a tackifier (such as a resin), and a wetting agent or edict, H. A. (Plenum, New York), pp. 3-11. detergent. On the stigma surface, the branched arabinogalactan 17. Gleeson, P. A., Jermyn, M. A. & Clarke, A. E. (1979) Anal. Bio- identified as a major component would fulfill the requirement chem. 92, 41-45. various free monosaccharides 18. Albersheim, P. & Anderson-Prouty, A. J. (1975) Annu. Rev. Plant for an adhesive base. The present Physiol. 26,31-52. on the stigma surface could serve as plasticizers, and the plas- 19. Talmadge, K. W. & Burger, M. M. (1975) M.T.P. Int. Rev. Sci.: ticizing effect might be reinforced by a contribution of mo- Biochem. Ser. One (Med. Tech. Publ. Co.) 5,43-93. nosaccharides from the pollen diffusate after initial pollen 20. Sharon, N. (1974) in Plant Carbohydrate Biochemistry, ed. capture (5). The glycoproteins of the stigma surface might be Pridham, J. B. (Academic, London), pp. 235-252. effective and the demonstrated on both 21. Gleeson, P. A. & Clarke, A. E. (1979) Biochem J. 181, in press. thickeners, glycolipids 22. Aspinall, G. 0. & Rosell, K. G. (1978) Phytochemistry 17, the pollen and the stigma surfaces would be ideal wetting 919-921. agents. Some component, analagous to resin to provide the 23. Clarke, A. E., Considine, J. A., Ward, R. & Knox, R. B. (1977) tackifier, is required to complete the requirements of an ideal Ann. Bot. 41,15-20. adhesive mixture, and the carotenoid pigments of the pollen 24. Labarca, C. & Loewus, F. (1973) Plant Physiol. 52,87-92. surface could fulfill this role. on initial contact of 25. Reynolds, G. E. T. (1971) in Aspects of Adhesion, ed. Alner D. Perhaps, pollen J. (Univ. of London Press, London), pp. 96-111. with stigma, the pollen contributes a pigment tackifier to the 26. Pegg, G. F. (1977) in Biochemistry Related to Speci- mixture on the stigma surface, thus enhancing its adhesive ficity in Host-Plant Pathogen Interactions, eds. Solheim, B. & properties. Efficient gluing is known to require roughened Raa, J. (Universitetsforlaget, Tromso, Norway). surfaces, and smooth surfaces do not accept adhesives efficiently 27. Ofek, I., Beachey, E. H. & Sharon, N. (1978) Trends. Biochem. The surface is for adhe- Sci. 3, 159-160. (25). pollen grain apparently adapted 28. Raa, J. (1977) in Cell Wall Biochemistry Related to Specificity sion because even relatively smooth wind-borne pollens have in Host-Plant Pathogen Interactions,-eds. Solheim, B. & Raa, surface patterning and provide a roughened surface for adhe- J. (Universitetsforlaget, Tromso, Norway), pp. 11-28. sion. 29. Edwards, P. A. W. (1978) Nature (London) 271, 248-249. Downloaded by guest on September 24, 2021