Continuous angiosperm pollen records from the Lower Cretaceous sequence of the Sorachi and Yezo Groups in the Teshionakagawa area of northern Hokkaido revise the first appearance times of four angiosperm pollen types in Japan: monosulcate, trichotomosulcate, tricolpate and tricolporate. The study shows that monosulcate, trichotomosulcate and tricolpate pollen types are present from at least the Barremian, while tricolporate type pollen is present from the late Albian. Compared with the first appearance times of Europe, North and South America, Africa and Asia, these times are about the same or even a little earlier. Notably, these results indicate the presence of the eudicotyledons in the Barremian of eastern Asia.

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Golgi vesicles (GVs) are closely involved in pollen tube growth. In the present study, we investigated the role of GV proteins, the main component of the isolated GVs, in germinating pollen. A GV-specific protein (GVP) was isolated by two-dimensional IEF-SDS-PAGE from a GV preparation extracted from Camellia japonica pollen. The isolated GVP had an apparent molecular mass of 70kDa and an isoelectric point of pI 5.2. The localization of GVP was investigated immunochemically using a mouse antiserum against GVP. The antibody reacted with the cytoplasm, and also with the new wall layer inside the intine, in the grain region of the germinating pollen, but not with the wall of the mature quiescent pollen grain. In germinating pollen, the antibody reacted with the inner callosic layer of the pollen tube wall, the GVs in the cytoplasm, and the callose plug. Similar results were also obtained from Camellia sasanquca, C. sinensis and Lilium longiflorum pollen. Therefore, it seems that GVP occurs in the callose-rich regions of the pollen wall, as well as in GVs; suggesting a relationship between GVP and callosic wall formation.

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Among proteins separated from an extract of Camellia japonica pollen by 2D-PAGE, three proteins, Cj 57, Cj 53 and Cj 50 with molecular weights of 57, 53 and 50kDa respectively, were detected as allergens by Western blotting using serum from a patient with camellia pollinosis. Moreover, the protein Cj 50 was identified as an enolase by mass spectrometry, and the amino acid sequence deduced by cloning of the Cj 50 gene confirmed that the protein is an enolase. Furthermore, the protein partially purified by column chromatography was found to have enolase activity. An antibody rised against Cj 50 recognized Cj 53 as well as Cj 50. Cj 53 may be an isoform of Cj 50 because it always was detected in the enolase fraction with Cj 50. These proteins were detected in the extracts of pollen grain, flower and leaf by Western blotting using the antibody, but Cj 53 was detected only in the pollen grain. Immuno-electron microscopy detected the labels of the antibody in the cytosol and generative cell nucleus of pollen, and in the cell nucleus of leaf cells. The immuno-labeling in the nucleus suggests that Cj 50 and/or Cj 53 have another physiological function besides enolase activity.

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Controversial arguments exist as to whether Nelumbo is native in Japan or introduced. We encountered considerable amounts of Nelumbo fossil pollen grains during the course of pollen analysis of the Karako Mire sediments, Japan. Pollen assemblages with a carbon-14 dating, and available stratigraphic data, indicate that the vegetation around the mire would have been an evergreen broad-leaved forest of Quercus, subgenera Cyclobalanopsis and Castanopsis, during the period 6000-2000 years B.P., and subsequently, for the last 2000 years, secondary Pinus forest. Nelumbo fossil pollen grains occur in more than half of samples studied, in notable amounts (7% or less). It is most likely that Nelumbo grew during a warm climate in a mire environment for more than 6000 years. These data are useful for the reconstruction of Nelumbo history in Japan.

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