Proteome Profiling of Maize Pollen Coats Reveals Novel Protein Components

Abstract

The pollen coat, which covers the exine wall of\ud pollen, is essential for initial sexual contact and thus successful fertilization in flowering plants. Pollen coat proteins (PCPs) not only mediate species specificity but are also needed for pollen–stigma recognition and pollen germination on the stigma. Maize (Zea mays L.) is one of the most common cereal crops in the world. To date, only a few PCPs from maize have been identified and characterized. In the present study, we extracted the pollen coat fraction from maize inbred line B73 with chloroform and purified the PCPs via a phenolbased\ud protocol prior to two-dimensional gel electrophoresis\ud (2-DE). By proteomic analysis, 26 protein spots were successfully identified and classified into 12 unique proteins. The protein composition of the maize pollen coat is distinctly different from those of the dicot plants Arabidopsis and rapeseed. Of the proteins identified in this study, eight (including p r o f i l i ns , caleos i n , Zea m 2 , β- expans i n - 1 0 , exopolygalacturonase, Rho GDP-dissociation inhibitor 1, Ras-related protein Rab-2-A, and putative subtilase) had not previously been observed in the maize pollen coat. Bioinformatic analysis showed that nine of the PCPs were secreted proteins and that most of them were extracellular. These PCPs are potentially involved in pollen germination and tube growth. The current study extracted and identified the pollen coat proteins of maize, one of the most important crops throughout the world. Proteome profiling of the maize pollen coat revealed many novel protein components potentially involved in pollen–stigma interactions and pollen germination. Our results provide basic knowledge and further the functional characterization of PCPs in wind-pollinated\ud species such as maize

The Pollen Coat Proteome: At the Cutting Edge of Plant Reproduction

“…The composition of the pollen coat proteome has been investigated in some plants including species of the Brassicaceae family [24,25], cereal crops, such as rice [26] and maize [27], and the olive tree [28]. The pollen coat can be effortlessly extracted by surface washing the intact pollen grains with different organic solvents such as cyclohexane [29], chloroform [27] and diethyl ether [30].…”

“…The pollen coat can be effortlessly extracted by surface washing the intact pollen grains with different organic solvents such as cyclohexane [29], chloroform [27] and diethyl ether [30]. Rinsing times as short as 10 se enable most of the pollen coating material to be removed.…”

“…Rinsing times as short as 10 se enable most of the pollen coating material to be removed. Proteins can be purified and solubilized from pollen coat preparations using sonication [29], precipitation with acetone [28], phenol-based extraction methods [31] and detergent-containing buffers [27]. Further analytical separation and identification of proteins can be achieved using either protein microsequencing- or one- or two-dimensional gel-based electrophoretic approaches in combination with mass spectrometry (MS) analysis.…”

“…EXPB1 (Zea m 1d) is the most abundant form in the maize pollen grain. Two of these β-expansins, namely EXPB1 (Zea m 1d) and EXPB10 (Zea m 1c), were consistently identified in the maize pollen coat fraction [27,40]. The predicted polypeptides of EXPB1 and EXPB10 genes are made up of 269 and 270 amino acids and show approximately 58 and 70% protein sequence identity with ryegrass Lol p 1 allergen, respectively.…”

“…In a recent study, Wu et al [27] scaled up maize pollen coat proteome profiling using a chloroform/phenol-based extraction method combined with an MS/MS-coupled two-dimensional electrophoresis (2-DE) strategy. This approach enabled up to 12 unique proteins to be identified (see Table 1).…”