Journal Article

Molecular Cloning of Two Exo-β-glucanases and Their in vivo Substrates in the Cell Walls of Lily Pollen Tubes

Hiroyuki Takeda, Takuo Yoshikawa, Xi-Zhen Liu, Naoki Nakagawa, Yi-Qin Li and Naoki Sakurai

in Plant and Cell Physiology

Published on behalf of Japanese Society of Plant Physiologists

Volume 45, issue 4, pages 436-444
Published in print April 2004 | ISSN: 0032-0781
Published online April 2004 | e-ISSN: 1471-9053 | DOI: http://dx.doi.org/10.1093/pcp/pch049
Molecular Cloning of Two Exo-β-glucanases and Their in vivo Substrates in the Cell Walls of Lily Pollen Tubes

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Full-length cDNA sequences of two exo-β-glucanases, LP-ExoI and LP-ExoII, secreted into cell walls of lily (Lilium longiflorum) pollen tube, were determined by RT-PCR. LP-ExoI exhibited over 80% similarity to LP-ExoII at both DNA and amino acid levels. RT-PCR showed that LP-ExoI transcripts were abundant in pollen grains and tubes, but could not be detected in leaf, stem, stigma, style, ovary, petal, filament, young root, young bud, and scale leaf of bulb. However, LP-ExoII transcripts ubiquitously existed in all the tissues tested. To determine the potential substrates of exo-β-glucanases, cell wall components of lily tissues were analyzed. Linkage analysis revealed that pollen tubes contained high levels of 3-Glc in hemicellulose (44.3%), while pollen grains had no detectable 3-Glc. The hemicellulose fraction of pollen tubes was treated with lichenase and the product was analyzed by HPLC-PAD to determine the origin of 3-Glc. Specific tetra-saccharide was liberated from hemicellulose of pollen tubes, suggesting the presence of 1,3 : 1,4-β-glucan in lily pollen tube hemicellulose. The structure of this 1,3 : 1,4-β-glucan may be different from cereal plant 1,3 : 1,4-β-glucan, since tri-saccharide was not detected in hemicellulose fraction after lichenase treatment. LP-ExoI and LP-ExoII, expressed in pollen grains and tubes, may be involved in the regulation of pollen tube elongation by hydrolyzing callose and 1,3 : 1,4-β-glucan within pollen tube walls.

Keywords: Keywords: Cell wall — Exo-β-glucanase — 1,3 : 1,4-β-glucan — Lichenase — Lily (Lilium longiflorum) — Pollen tube elongation.; Abbreviations: Ara, arabinose; DMSO, dimethyl sulfoxide; DW, deionized water; Fuc, fucose; Gal, galactose; Glc, glucose; HPLC-PAD, high performance liquid chromatography with pulsed amperometric detector; NMNO, N-morpholino N-oxide; Man, mannose; MOPS, 3-(N-morpholino) propanesulfonic acid; NS, neutral sugar; RACE, rapid amplification of cDNA ends; Rha, rhamnose; RT, reverse transcript; Xyl, xylose.

Journal Article.  6456 words.  Illustrated.

Subjects: Biochemistry ; Molecular and Cell Biology ; Plant Sciences and Forestry

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