Cytological and Proteomic Analysis of Ginkgo biloba Pollen Intine

The pollen intine plays important roles in pollen germination and tube growth,but related information in Ginkgo biloba remains unclear.We isolated and obtained de-exined pollen from G.biloba.Using fluorescent probes,we observed the strongest cellulose fluorescence in the pollen intine.De-esterified pectin immunolabeled with JIM5 was present throughout the entire cell wall,whereas esterified pectin recognized by the monoclonal antibody JIM7 was concentrated in some regions.Callose staining with aniline blue was observed across the entire surface of the pollen intine.These results were confirmed by Fourier Transform InfraRed(FTIR)analysis.We also used proteomic approaches to identify different proteins between mature and de-exined pollen(48h after hydration)in vitro.Based on mass spectrometry,de-exined pollen had more proteins than mature pollen,including calmodulin,serine hydroxymethyltransferase,β-galactosidase 6,and class IV chitinase.According to Gene Ontology(GO)analysis,the differentially expressed proteins were mainly associated with transportation,defense reaction,sugar metabolism,energy metabolism,signal transduction,and cell wall formation.These findings suggest that most proteins involved in pollen germination and pollen tube growth are synthesized during pollen hydration,indicating the important role of pollen hydration in the reproductive process of G.Biloba.

Sporophytic control of anther development and male fertility by glucose-6-phosphate/phosphate translocator 1(OsGPT1) in rice

Coordination between the sporophytic tissue and the gametic pollen within anthers is tightly controlled to achieve the optimal pollen fitness. Glucose-6-phosphate/phosphate translocator(GPT) transports glucose-6-phosphate, a key precursor of starch and/or fatty acid biosynthesis, into plastids. Here, we report the functional characterization of Os GPT1 in the rice anther development and pollen fertility. Pollen grains from homozygous osgpt1 mutant plants fail to accumulate starch granules, resulting in pollen sterility. Genetic analyses reveal a sporophytic effect for this mutation. Os GPT1 is highly expressed in the tapetal layer of rice anther. Degeneration of the tapetum, an important process to provide cellular contents to support pollen development, is impeded in osgpt1 plants. In addition, defective intine and exine are observed in the pollen from osgpt1 plants. Expression levels of multiple genes that are important to tapetum degeneration or pollen wall formation are significantly decreased in osgpt1 anthers. Previously, we reported that At GPT1 plays a gametic function in the accumulation of lipid bodies in Arabidopsis pollen. This report highlights a sporophytic role of Os GPT1 in the tapetum degeneration and pollen development. The divergent functions of Os GPT1 and At GPT1 in pollen development might be a result of their independent evolution after monocots and dicots diverged.

PECTATE LYASE-LIKE10 is associated with pollen wall development in Brassica campestris

PECTATE LYASE‐LIKE10（PLL10） was previously identified as one of the differentially expressed genes both in microspores during the late pollen developmental stages and in pistils during the fertilization process in Chinese cabbage（Brassica campestris ssp. chinensis）. Here, antisense‐RNA was used to study the functions of BcPLL10 in Chinese cabbage. Abnormal pollen was identified in the transgenic lines（bcpll10‐4, ‐5, and ‐6）. In fertilization experiments, fewer seeds were harvested when the antisense‐RNA lines were used as pollen donor. In vivo and in vitro pollen germination assays less germinated pollen tubes were observed in bcpll10 lines. Scanning electron microscopy observation verified that the tryphine materials were over accumulated around the pollen surface and sticked them together in bcpll10.Moreover, transmission electron microscopy observation revealed that the internal endintine was overdeveloped and predominantly occupied the intine, and disturbed thenormal proportional distribution of the two layers in the non‐germinal furrow region; and no obvious demarcation existed between them in the germinal furrow region in the bcpll10 pollen. Collectively, this study presented a novel PLL gene that played an important role during the pollen wall development in B. campestris, which may also possess potential importance for male sterility usage in agriculture.