Laboratory of Molecular Information and Cellular Regulation
Group of Plant Reproductive Systems

ProfessorTetsuya Higashiyama
Cell-cell Signaling in Plant Reproduction
Associate ProfessorNarie Sasaki
Molecular Organization of Organelle Nucleoids
LecturerMasahiro Kanaoka
Action Mechanism and Molecular Evolution of Pollen Tube Attractants
Cell fate control in plant early development
▶Laboratory HP
Tetsuya Higashiyama Professor
Lab members

Among biological phenomena, reproduction is one of the most intriguing events. Flowering plants have evolved a unique reproduction mechanism called double fertilization. Double fertilization is not only a fascinating and sophisticated biological phenomenon, but is also directly important to human beings, e.g., in crop production. Its molecular mechanism, however, is not well understood. Our laboratory seeks to clarify the key molecular events required to achieve reproduction in plants and other eukaryotes, and thus to elucidate universal biological mechanisms pertaining to intercellular communication, heredity and development.

Intercellular Signaling in Pollen Tube Guidance and Double Fertilization

Studies of sexual reproduction in flowering plants are hampered by the thick and opaque layers of female tissues that enclose reproductive cells. We are using a unique plant, Torenia fournieri, whose female reproductive cells protrude from the tissue of the unfertilized seed (ovule). We succeeded in developing an in vitro Torenia system, wherein both pollen tube attraction to the egg apparatus and double fertilization occur in vitro.

Based on our in vitro Torenia system, we revealed mechanisms of pollen tube guidance. For example, by laser ablation of specific cells, we found that two synergid cells on the side of the egg cell emit diffusible pollen tube attractant(s). The attractant derived from the ovule tissue had been sought for more than 140 years. The attractant of Torenia was likely to be species-specific, even in closely related species. Expecting that the molecule might be a protein with rapid molecular evolution, we performed EST analysis of isolated synergid cells of Torenia. We found two defensin-like polypeptides expressed abundantly and specifically in the synergid cell. We finally identified these proteins as attractant molecules and named them as LUREs. This work was selected as a cover story in Nature. Based on identification of LUREs, various projects on pollen tube guidance are now in progress. There projects involves identification of “AMOR”, which is an factor required for capacitation of pollen tubes.

We have also developed live-cell imaging technique for studying double fertilization both in Torenia and Arabidopsis, which will be utilized for identification of novel factors required for double fertilization.

Organelle Nucleoids

Mitochondria also play important roles in sexual reproduction. They have their own DNA, which is packed into the nucleoid with histone-like proteins. For these studies, we are using Physarum polycephalum, which contains relatively large mitochondrial nucleoids. We developed a technique for isolating pure mitochondrial nucleoids from this species. The isolated nucleoids can undergo both RNA transcription and DNA duplication. Based on this technique, we will determine how molecular structures of the mitochondrial nucleoid contribute to the function of mitochondrial DNA and biological phenomena. In addition, we will utilize our techniques and information for studies of organelle nucleoids of malaria parasites, which will be also important for medical purposes.

Single Cell Analysis for Opening a New Frontier of Biology

Our studies are based on development of new techniques for analyses of targeted cells. For example, we have developed a laser-assisted thermal-expansion microinjector to deliver materials into the protruding female reproductive cells of Torenia. We are also trying to develop a deep-sequencing method using a targeted single cell. We are expecting that such single-cell based studies in unique biological models will establish a new field of biology. Please refer our lab homepage for the most recent information.


Mechanism of pollen tube guidance (left) and our in vitro system for studying this process (right)


  1. Higashiyama T. et al. (2001) Pollen tube attraction by the synergid cell. Science 293, 1480-1483.
  2. Mori T. et al. (2006) GENERATIVE CELL SPECIFIC 1 is essential for angiosperm fertilization. Nature Cell Biol. 8, 64-71.
  3. Nishimura Y. et al. (2006) Active digestion of sperm mitochondrial DNA in single living sperm revealed by optical tweezers. Proc. Natl. Acad. Sci. USA 103, 1382-1387.
  4. Higashiyama T. et al. (2006) Species preferentiality of the pollen tube attractant derived from the synergid cell of Torenia fournieri. Plant Physiol. 142, 481-491.
  5. Ingouff M. et al. (2007) Distinct dynamics of HISTONE3 variants between the two fertilization products in plants. Curr. Biol. 17, 1032-1037.
  6. Kanaoka M.M. et al. (2008) SCREAM/ICE1 and SCREAM2 specify three cell-state transitional steps leading to Arabidopsis stomatal differentiation. Plant Cell 20, 1775-1785.
  7. Sasaki N. et al. (2009) The Plasmodium HU homolog, which binds the plastid DNA sequence-independent manner, is essential for the parasite's survival. FEBS Lett. 583, 1446-1450.
  8. Okuda S., Tsutsui H., et al. (2009) Defensin-like polypeptide LUREs are pollen tube attractants secreted from synergid cells. Nature 458, 357-361.