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The Plant Cell 19:2320-2321 (2007) © 2007 American Society of Plant Biologists
Elucidating the Function of Synergid Cells: A Regulatory Role for MYB98neckardt{at}aspb.org
Synergid cells are two specialized cells that lie adjacent to the egg cell in the female gametophyte of angiosperms and play an essential role in pollen tube guidance and function. The term synergid comes from the Greek "synergos," which means "working together" and was reportedly coined by Eduard Strasburger, a famous 19th century botanist who was one of the first to note that these cells somehow assist fertilization of the egg (Vesque, 1878
After a pollen grain germinates on the surface of the stigma, it forms a pollen tube that grows through the style and is guided through the micropyle (a minute opening at one end of the ovule). The pollen tube grows toward and into one of the synergid cells, which begins to degenerate as the pollen tube enters it (or shortly before). The pollen tube then ceases growth, ruptures, and releases its two sperm cells, which migrate to the egg and central cell, fertilization of which gives rise to the embryo and endosperm, respectively. The synergid cells are necessary for pollen tube guidance in the ovule, as shown by Higashiyama et al. (2001)
The synergid cell wall forms a highly thickened structure called the filiform apparatus at the micropylar end, consisting of numerous finger-like projections into the synergid cytoplasm. This structure greatly increases the surface area of the plasma membrane in this region, which is also associated with an elaborated endoplasmic reticulum. It is thought that the filiform apparatus mediates the transport of molecules into and out of the synergid cells (Willemse and van Went, 1984
In this issue of The Plant Cell, Punwani et al. (pages 2557–2568) show that the R2R3 Myb protein MYB98 functions as a transcriptional regulator of genes that are expressed in synergid cells and are required for formation of the filiform apparatus and pollen tube guidance. Kasahara et al. (2005)
Punwani et al. found that a fusion construct of MYB98:green fluorescent protein (MYB98:GFP) driven by the MYB98 promoter localized to the nuclei of synergid cells (see figure), and in vitro DNA mobility shift assays showed that MYB98 binds a consensus binding sequence for mammalian c-Myb, which has an R2R3 Myb domain highly similar to that of MYB98. The authors then used real-time RT-PCR to investigate the expression of 18 genes shown by Steffen et al. (2007)
These experiments strongly suggest that MYB98 functions as a positive regulator of transcription in synergid cells. Additional experiments are necessary to elucidate this function more precisely, for example, to identify the specific binding site and discover which genes are direct rather than indirect targets of MYB98. Punwani et al. show that MYB98 binds specifically to a core TAAC sequence present in the c-Myb consensus sequence. However, TAAC forms the core of many of the binding sites known for R2R3 binding proteins and, as noted by the authors, is found one or more times in virtually every promoter in the Arabidopsis genome. The use of a random binding site selection assay (e.g., Huang et al., 1993  ; Tang and Perry, 2003) would be an important step to establish the specific binding site of MYB98. In addition, although Punwani et al. show that MYB98 is required for the expression of 16 genes expressed in synergid cells, it is not clear if all of these genes are direct targets. Activation assays (e.g., Pasquali et al., 1994) testing the potential of MYB98 to activate transcription from the various target promoters would be one way to distinguish direct from indirect targets. In another set of experiments, the authors chose six of the synergid-expressed genes downregulated in myb98 mutant gametophyte tissue and investigated their subcellular localization patterns by transforming wild-type plants with GFP fusion constructs containing complete coding sequences and upstream regions. One of these genes, which encodes a Cys-rich protein lacking a putative signal peptide, was found to be expressed throughout the cytoplasm of synergid cells. Expression of the remaining five, all of which contained N-terminal signal peptides, was strongly associated with the filiform apparatus and not detected in the cytoplasm. Expression of a GFP construct containing only the putative signal peptide from one of these genes (driven by the MYB98 promoter) was also localized to the filiform apparatus. This suggests that an N-terminal signal peptide is sufficient to localize a synergid-expressed protein to the filiform apparatus.
The authors hypothesize that the synergid-expressed genes regulated by MYB98 (whether directly or indirectly) play roles in the formation and function of the filiform apparatus. Several of the Cys-rich proteins share similarity to other proteins known to play specialized roles in signaling. These include defensins, which are small Cys-rich proteins that have antimicrobial properties and function in signaling (Thomma et al., 2002
www.plantcell.org/cgi/doi/10.1105/tpc.107.055640
Higashiyama, T. (2002). The synergid cell: Attractor and acceptor of the pollen tube for double fertilization. J. Plant Res. 115: 149–160.[CrossRef][Web of Science][Medline] Higashiyama, T., Yabe, S., Sasaki, N., Nishimura, Y., Miyagishima, S.-y., Kuroiwa, H., and Kuroiwa, T. (2001). Pollen tube attraction by the synergid cell. Science 293: 1480–1483. Huang, B.-Q., and Russell, S.D. (1992). Female germ unit: Organization, isolation, and function. Int. Rev. Cytol. 140: 233–292.[CrossRef][Web of Science] Huang, H., Mizukami, Y., Yi, Y., and Ma, H. (1993). Isolation and characterization of the binding sequences for the product of the Arabidopsis floral homeotic gene AGAMOUS. Nucleic Acids Res. 21: 4769–4776. Kasahara, R.D., Portereiko, M.F., Sandaklie-Nikolova, L., Rabiger, D.S., and Drews, G.N. (2005). MYB98 is required for pollen tube guidance and synergid cell differentiation in Arabidopsis. Plant Cell 17: 2981–2992. Pasquali, G., Ouwerkerk, P.B.F., and Memlink, J. (1994). Versatile transformation vectors to assay the promoter activity of DNA elements in plants. Gene 149: 373–374.[CrossRef][Web of Science][Medline] Punwani, J.A., Rabiger, D.S., and Drews, G.N. (2007). MYB98 positively regulates a battery of synergid-expressed genes encoding filiform apparatus–localized proteins. Plant Cell 19: 2557–2568. Steffen, J.G., Kang, I.-H., Macfarlane, J., and Drews, G.N. (2007). Identification of genes expressed in the Arabidopsis female gametophyte. Plant J. 51: 281–292.[CrossRef][Web of Science][Medline] Tang, W., and Perry, S.E. (2003). Binding site selection for the plant MADS domain protein AGL15. An in vitro and in vivo study. J. Biol. Chem. 278: 28154–28159. Thomas, S.G., and Franklin-Tong, V.E. (2004). Self-incompatibility triggers programmed cell death in Papaver pollen. Nature 429: 305–309.[CrossRef][Medline] Thomma, B.P., Cammue, B.P., and Thevissen, K. (2002). Plant defensins. Planta 216: 193–202.[CrossRef][Web of Science][Medline] Vesque, J. (1878). Development du sac embryonnaire des phanérogamesangiosperms. Ann. Sci. Nat. Bot. 6: 237–285. Willemse, M.T.M., and van Went, J.L. (1984). The female gametophyte. In Embryology of Angiosperms, B.M. Johri, ed (Berlin: Springer-Verlag), pp. 159–196. Related articles in Plant Cell:
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