Dataset Information

Electrophoretic profiling and immunocytochemical detection of pectins and arabinogalactan proteins in olive pollen during germination and pollen tube growth.

ABSTRACT:

Background and aims

Cell wall pectins and arabinogalactan proteins (AGPs) are important for pollen tube growth. The aim of this work was to study the temporal and spatial dynamics of these compounds in olive pollen during germination.

Methods

Immunoblot profiling analyses combined with confocal and transmission electron microscopy immunocytochemical detection techniques were carried out using four anti-pectin (JIM7, JIM5, LM5 and LM6) and two anti-AGP (JIM13 and JIM14) monoclonal antibodies.

Key results

Pectin and AGP levels increased during olive pollen in vitro germination. (1 ? 4)-?-d-Galactans localized in the cytoplasm of the vegetative cell, the pollen wall and the apertural intine. After the pollen tube emerged, galactans localized in the pollen tube wall, particularly at the tip, and formed a collar-like structure around the germinative aperture. (1 ? 5)-?-l-Arabinans were mainly present in the pollen tube cell wall, forming characteristic ring-shaped deposits at regular intervals in the sub-apical zone. As expected, the pollen tube wall was rich in highly esterified pectic compounds at the apex, while the cell wall mainly contained de-esterified pectins in the shank. The wall of the generative cell was specifically labelled with arabinans, highly methyl-esterified homogalacturonans and JIM13 epitopes. In addition, the extracellular material that coated the outer exine layer was rich in arabinans, de-esterified pectins and JIM13 epitopes.

Conclusions

Pectins and AGPs are newly synthesized in the pollen tube during pollen germination. The synthesis and secretion of these compounds are temporally and spatially regulated. Galactans might provide mechanical stability to the pollen tube, reinforcing those regions that are particularly sensitive to tension stress (the pollen tube-pollen grain joint site) and mechanical damage (the tip). Arabinans and AGPs might be important in recognition and adhesion phenomena of the pollen tube and the stylar transmitting cells, as well as the egg and sperm cells.

SUBMITTER: Castro AJ

PROVIDER: S-EPMC3718210 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Electrophoretic profiling and immunocytochemical detection of pectins and arabinogalactan proteins in olive pollen during germination and pollen tube growth.

Castro Antonio J AJ Suárez Cynthia C Zienkiewicz Krzysztof K Alché Juan de Dios Jde D Zienkiewicz Agnieszka A Rodríguez-García María Isabel MI

Annals of botany 20130526 3

<h4>Background and aims</h4>Cell wall pectins and arabinogalactan proteins (AGPs) are important for pollen tube growth. The aim of this work was to study the temporal and spatial dynamics of these compounds in olive pollen during germination.<h4>Methods</h4>Immunoblot profiling analyses combined with confocal and transmission electron microscopy immunocytochemical detection techniques were carried out using four anti-pectin (JIM7, JIM5, LM5 and LM6) and two anti-AGP (JIM13 and JIM14) monoclonal ...[more]

PMID: 23712452

Similar Datasets

Project description:Background and aimsA pollen grain contains a number of esterases, many of which are released upon contact with the stigma surface. However, the identity and function of most of these esterases remain unknown. In this work, esterases from olive pollen during its germination were identifided and functionally characterized.MethodsThe esterolytic capacity of olive (Olea europaea) pollen was examined using in vitro and in-gel enzymatic assays with different enzyme substrates. The functional analysis of pollen esterases was achieved by inhibition assays by using specific inhibitors. The cellular localization of esterase activities was performed using histochemical methods.Key resultsOlive pollen showed high levels of non-specific esterase activity, which remained steady after hydration and germination. Up to 20 esterolytic bands were identified on polyacrylamide gels. All the inhibitors decreased pollen germinability, but only diisopropyl fluorophosphate (DIFP) hampered pollen tube growth. Non-specific esterase activity is localized on the surface of oil bodies (OBs) and small vesicles, in the pollen intine and in the callose layer of the pollen tube wall. Acetylcholinesterase (AChE) activity was mostly observed in the apertures, exine and pollen coat, and attached to the pollen tube wall surface and to small cytoplasmic vesicles.ConclusionsIn this work, for the first time a systematic functional characterization of esterase enzymes in pollen from a plant species with wet stigma has been carried out. Olive pollen esterases belong to four different functional groups: carboxylesterases, acetylesterases, AChEs and lipases. The cellular localization of esterase activity indicates that the intine is a putative storage site for esterolytic enzymes in olive pollen. Based on inhibition assays and cellular localization of enzymatic activities, it can be concluded that these enzymes are likely to be involved in pollen germination, and pollen tube growth and penetration of the stigma.

Project description:Pollen germination, along with pollen tube growth, is an essential process for the reproduction of flowering plants. The germinating pollen with tip-growth characteristics provides an ideal model system for the study of cell growth and morphogenesis. As an essential step towards a detailed understanding of this important process, the objective of this study was to comprehensively analyze the transcriptome changes during pollen germination and pollen tube growth. Using Affymetrix Arabidopsis ATH1 Genome Arrays, this study is the first to show the changes in the transcriptome from desiccated mature pollen grains to hydrated pollen grains and then to pollen tubes of Arabidopsis thaliana. The number of expressed genes, either for total expressed genes or for specifically expressed genes, increased significantly from desiccated mature pollen to hydrated pollen and again to growing pollen tubes, which is consistent with the finding that pollen germination and tube growth was significantly inhibited in vitro by a transcriptional inhibitor. The results of GO analyses showed that expression of genes related to cell rescue, transcription, signal transduction and cellular transport were significantly changed, especially for up-regulation, during pollen germination and tube growth, respectively. In particular, genes of the CaM/CML, CHX and Hsp families showed the most significant changes during pollen germination and tube growth. These results demonstrate that the overall transcription of genes, both in the number of expressed genes and in the levels of transcription, was increased. Furthermore, the appearance of many novel transcripts during pollen germination as well as tube growth indicates that these newly expressed genes may function in this complex process. SUBMITTER_CITATION: Yi Wang, Wen-Zheng Zhang, Lian-Fen Song, Jun-Jie Zou, Zhen Su, and Wei-Hua Wu. Transcriptome analyses show changes in gene expression to accompany pollen germination and tube growth in Arabidopsis. Plant Physiol. September 5, 2008; 10.1104/pp.108.126375 Experiment Overall Design: Three samples are analyzed in this experiment. They are desiccated mature pollen grains (MP), hydrated pollen grains (HP) and growing pollen tubes (PT) of Arabidopsis thaliana, respectively. Each sample has two biological replicates, so that there are 6 data sets of ATH1 array in this experiment.

Project description:BACKGROUND: Arabinogalactan proteins (AGPs) are cell wall proteoglycans that have been shown to be important for pollen development. An Arabidopsis double null mutant for two pollen-specific AGPs (agp6 agp11) showed reduced pollen tube growth and compromised response to germination cues in vivo. A microarray experiment was performed on agp6 agp11 pollen tubes to search for genetic interactions in the context of pollen tube growth. A yeast two-hybrid experiment for AGP6 and AGP11 was also designed. RESULTS: The lack of two specific AGPs induced a meaningful shift in the gene expression profile. In fact, a high number of genes showed altered expression levels, strengthening the case that AGP6 and AGP11 are involved in complex phenomena. The expression levels of calcium- and signaling-related genes were found to be altered, supporting the known roles of the respective proteins in pollen tube growth. Although the precise nature of the proposed interactions needs further investigation, the putative involvement of AGPs in signaling cascades through calmodulin and protein degradation via ubiquitin was indicated. The expression of stress-, as well as signaling- related, genes was also changed; a correlation that may result from the recognized similarities between signaling pathways in both defense and pollen tube growth.The results of yeast two-hybrid experiments lent further support to these signaling pathways and revealed putative AGP6 and AGP11 interactors implicated in recycling of cell membrane components via endocytosis, through clathrin-mediated endosomes and multivesicular bodies. CONCLUSIONS: The data presented suggest the involvement of AGP6 and AGP11 in multiple signaling pathways, in particular those involved in developmental processes such as endocytosis-mediated plasma membrane remodeling during Arabidopsis pollen development. This highlights the importance of endosomal trafficking pathways which are rapidly emerging as fundamental regulators of the wall physiology.

Project description:BackgroundPollen tube growth and fertilization are key processes in angiosperm sexual reproduction. The transmitting tract (TT) of Nicotiana tabacum controls pollen tube growth in part by secreting pistil extensin-like protein III (PELPIII), transmitting-tract-specific (TTS) protein and 120 kDa glycoprotein (120 K) into the stylar extracellular matrix. The three arabinogalactan proteins (AGP) are referred to as stylar AGPs and are the focus of this research. The transmitting tract regulates pollen tube growth, promoting fertilization or rejecting pollen tubes.ResultsThe N-terminal domain (NTD) of the stylar AGPs is proline rich and polymorphic among Nicotiana spp. The NTD was predicted to be mainly an intrinsically disordered region (IDR), making it a candidate for protein-protein interactions. The NTD is also the location for the majority of the predicted O-glycosylation sites that were variable among Nicotiana spp. The C-terminal domain (CTD) contains an Ole e 1-like domain, that was predicted to form beta-sheets that are similar in position and length among Nicotiana spp. and among stylar AGPs. The TTS protein had the greatest amino acid and predicted O-glycosylation conservation among Nicotiana spp. relative to the PELPIII and 120 K. The PELPIII, TTS and 120 K genes undergo negative selection, with dn/ds ratios of 0.59, 0.29 and 0.38 respectively. The dn/ds ratio for individual species ranged from 0.4 to 0.9 and from 0.1 to 0.8, for PELPIII and TTS genes, respectively. These data indicate that PELPIII and TTS genes are under different selective pressures. A newly discovered AGP gene, Nicotiana tabacum Proline Rich Protein (NtPRP), was found with a similar intron-exon configuration and protein structure resembling other stylar AGPs, particularly TTS.ConclusionsFurther studies of the NtPRP gene are necessary to elucidate its biological role. Due to its high similarity to the TTS gene, NtPRP may be involved in pollen tube guidance and growth. In contrast to TTS, both PELPIII and 120 K genes are more diverse indicating a possible role in speciation or mating preference of Nicotiana spp. We hypothesize that the stylar AGPs and NtPRP share a common origin from a single gene that duplicated and diversified into four distinct genes involved in pollen-style interactions.

Dataset Information

Electrophoretic profiling and immunocytochemical detection of pectins and arabinogalactan proteins in olive pollen during germination and pollen tube growth.

Background and aims

Methods

Key results

Conclusions

Publications

Electrophoretic profiling and immunocytochemical detection of pectins and arabinogalactan proteins in olive pollen during germination and pollen tube growth.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets