The Predominant Protein on the Surface of Maize Pollen Is an Endoxylanase Synthesized by a Tapetum mRNA with a Long 5′ Leader

Abstract

In plants, the pollen coat covers the exine wall of the pollen and is the outermost layer that makes the initial contact with the stigma surface during sexual reproduction. Little is known about the constituents of the pollen coat, especially in wind-pollinated species. The pollen coat was extracted with diethyl ether from the pollen of maize (Zea mays L.), and a predominant protein of 35 kDa was identified. On the basis of the N-terminal sequence of this protein, a cDNA clone of the Xyl gene was obtained by reverse transcriptase-polymerase chain reaction. The deduced amino acid sequence of the 35-kDa protein shared similarities with the sequences of many microbial xylanases and a barley aleurone-layer xylanase. The 35-kDa protein in the pollen-coat extract was purified to homogeneity by fast protein liquid chromatography and determined to be an acidic endoxylanase that was most active on oat spelt xylan. Northern and in situ hybridization showed that Xyl was specifically expressed in the tapetum of the anther after the tetrad microspores had become individual microspores. Southern hybridization and gene-copy reconstruction studies showed only one copy of the Xyl gene per haploid genome. Analyses of the genomic DNA sequence of Xyl and RNase protection studies with the transcript revealed many regulatory motifs at the promoter region and an intron at the 5 leader region of the transcript. The Xyl transcript had a 562-nucleotide (nt) 5 leader, a 54-nt sequence encoding a putative signal peptide, a 933-nt coding sequence, and a 420-nt 3-untranslated sequence. The unusually long 5 leader had an open reading frame encoding a putative 175-residue protein whose sequence was most similar to that of a microbial arabinosidase. The maize xylanase is the first enzyme documented to be present in the pollen coat. Its possible role in the hydrolysis of the maize type II primary cell wall (having xylose, glucose, and arabinose as the major moieties) of the tapetum cells and the stigma surface is discussed.In plants, pollen is the male gametophyte that interacts with the female reproductive organ during sexual reproduction (1-4). It is produced in the anther of a flower, where diploid pollen mother cells undergo meiosis to produce haploid cells, termed microspores. Microspores mature in the anther locule, which is an enclosed, extracellular fluid space, to become pollen. During their maturation (microsporogenesis), the microspores interact with the tapetum, which consists of one layer of metabolically active cells enclosing the locule. The tapetum serves several important roles in the maturation of the microspores (5). Throughout microsporogenesis, the tapetum transfers nutrients to, and regulates the composition of, the locule fluid. In addition, at an early stage of microspore maturation, the tapetum secretes ␤-glucanase, which hydrolyzes the ␤-glucan (callose) cell wall enclosing the microspore tetrads (6). The released individual microspores continue to mature in the locule. Midway through microsporogenesis, the tapetum secr...

Tapetosomes in Brassica Tapetum Accumulate Endoplasmic Reticulum–Derived Flavonoids and Alkanes for Delivery to the Pollen Surface

“…Putative lipid-rich particles in tapetum cells of diverse species can be seen in electron microscopy images (Hesse et al, 1993), and flavonoids and neutral lipids are present universally on pollen of diverse species. Yet, tapetosomes are clearly absent in maize and other cereals (Bih et al, 1999;Wu et al, 2002). The maize pollen coat does not have oleosins, and its major constituent is an endoxylanase that is synthesized and stored in the tapetum cell cytosol for release upon PCD.…”

Pollen and Stigma Structure and Function: The Role of Diversity in Pollination

“…Further studies have identified medium-chain, long-chain, and very-long-chain fatty acids, proteins, glycoconjugates, carotenoids, and flavonoids (Piffanelli et al, 1998). In maize, the major protein component in the pollen coat is xylanase, which, upon its release at stigma binding, facilitates pollen tube invasion by hydrolyzing the xylan on the stigma surface (Bih et al, 1999). In B. oleracea, >100 pollen coat proteins, ranging in size from 5 to 50 kD, have been separated on denaturing gels.…”

“…A comparative study of tube emergence across taxa could yield insight into the evolution of the mechanisms necessary for different forms of tube emergence. Although apertures provide an important means of communication between pollen and the outside world, they also are sites of vulnerability to excess desiccation and invasion by fungal hyphae (Bih et al, 1999); consequently, some plants cover the apertures with specialized intines, lipid-rich pollen coatings, or opercula. Mutations that disrupt the regular pattern of apertures have been identified, but it is not currently feasible to use genetics to remove or add these features.…”

Proteomics Identification of Differentially Expressed Proteins Associated with Pollen Germination and Tube Growth Reveals Characteristics of Germinated Oryza sativa Pollen

“…The down-regulation of most of these proteins should result from their release from the germinating pollen into the medium (Table II), which is possibly similar to their situation in vivo. In the monocots maize and rice, ␤-1,4-xylanase is the major protein component in the pollen coat (14,53) and possibly facilitates pollen tube invasion by hydrolyzing the xylan on the stigma surface (53). Besides being responsible for degradation of demethylesterified pectin (7), polygalacturonase-mediated release of oligogalacturonides can act as a signal for different development processes (54).…”