For more than 140 years, pollen tube guidance in flowering plants has been thought to be mediated by chemoattractants derived from target ovules. However, there has been no convincing evidence of any particular molecule being the true attractant that actually controls the navigation of pollen tubes towards ovules. Emerging data indicate that two synergid cells on the side of the egg cell emit a diffusible, species-specific signal to attract the pollen tube at the last step of pollen tube guidance. Here we report that secreted, cysteine-rich polypeptides (CRPs) in a subgroup of defensin-like proteins are attractants derived from the synergid cells. We isolated synergid cells of Torenia fournieri, a unique plant with a protruding embryo sac, to identify transcripts encoding secreted proteins as candidate molecules for the chemoattractant(s). We found two CRPs, abundantly and predominantly expressed in the synergid cell, which are secreted to the surface of the egg apparatus. Moreover, they showed activity in vitro to attract competent pollen tubes of their own species and were named as LUREs. Injection of morpholino antisense oligomers against the LUREs impaired pollen tube attraction, supporting the finding that LUREs are the attractants derived from the synergid cells of T. fournieri.

The results indicated that this class of CRP proteins is a common pollen tube attractant in Torenia species. The sequence diversity of these proteins is important for species-specific pollen tube attraction.

Summary Gametophytic cells in the female embryo sac, the egg, the central and the synergid cells have unique roles in sexual reproduction in higher plants. To investigate their cell-specific features, an efficient method of distinguishing and isolating each gametophytic cell is required. By testing the activities of several cellulase enzymes, we modified our previous cell isolation method and succeeded in isolating each cell from the Torenia and Lindernia species. We also established genetic markers that show specific expression patterns in each gametophytic cell. These markers, together with the cell isolation method, will be useful for analyzing the function of gametophytic cells.