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Plant and Cell Physiology Advance Access originally published online on July 4, 2009
Plant and Cell Physiology 2009 50(8):1558-1572; doi:10.1093/pcp/pcp100
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© The Author 2009. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oxfordjournals.org

Pollen-Specific SKP1-Like Proteins are Components of Functional SCF Complexes and Essential for Lily Pollen Tube Elongation

Liang-Chi Chang1,2, Cian-Ling Guo2, Yun-Shan Lin2, Hongyong Fu2, Co-Shing Wang3 and Guang-Yuh Jauh1,2,*

1Institute of Life Science, National Defense Medical Center, Taipei, Taiwan, ROC
2Institute of Plant and Microbial Biology, Academia Sinica, Taipei 115, Taiwan, ROC
3Graduate Institute of Biotechnology, National Chung-Hsing University, Taichung 402, Taiwan, ROC

*Corresponding author: E-mail, jauh{at}gate.sinica.edu.tw; Fax, +886-2-27827954.


  Abstract

The ubiquitin–proteasome pathway mediates protein degradation and is involved in diverse aspects of plant development and differentiation, including pollen tube elongation and self-incompatibility. We characterized three lily (Lilium longiflorum) SKP1-like genes, LSK1–LSK3, that are specifically expressed in late pollen developmental stages and the elongating pollen tube. The encoded peptide sequences reveal that LSK1–LSK3 share high identity with Arabidopsis ASK1 and contain a putative N-terminal CUL1- and a C-terminal F-box-interacting domain. Yeast two-hybrid and in vitro affinity binding assays revealed that the LSKs associate with lily CULLIN1. In addition, the LSK genes can functionally complement the yeast skp1 deletion mutant YDR328C. To investigate their biological functions in pollen tube elongation, an in vivo approach for green fluorescent protein (GFP)-tagged dominant-negative LSK1–LSK3 was developed. Microprojectile bombardment with N-terminally truncated LSK1–LSK3 (LSK1–LSK3{Delta}–GFP) significantly retarded pollen tube elongation in both in vitro germination and in vivo self- and cross-pollination after >12 h incubation. Interestingly, elongation of pollen tubes harboring overexpressed LSK2{Delta}–GFP and LSK3{Delta}–GFP was substantially inhibited within the self-pollinated styles. The elongation of most LSK2{Delta}–GFP-transformed pollen tubes could germinate only on the stigmatic surface of self style and showed statistically significant growth arrest as compared with control pollen tubes. Lily exhibits typical gametophytic self-incompatibility via an unknown mechanism, but LSK2 and LSK3 may be involved in this complex machinery. These results suggest critical roles for LSK1–LSK3 in regulating fundamental pollen tube elongation in vitro and in vivo and that the 26S proteasome-mediated protein pathway plays an important role in pollen tube elongation.

Keywords: Lily (Lilium longiflorum) - LLCUL1 - LSK - Pollen tube elongation - SCF complex - Self-incompatibility

Abbreviations: AChE, acetylcholinesterase; ChAT, choline acetyltransferase; CNBr, cyanogen bromide; FAP, FIM-associated protein; 5-FOA, 5-fluoro-orotic acid; GFP, green fluorescent protein; GSI, gametophytic self-incompatibility; GST, glutathione S-transferase; LLCUL1, lily CULLIN1; LSK, lily SKP1-like protein; RACE, rapid amplification of cDNA ends; RT–PCR, reverse transcrition–PCR; SI, self-incompatibilty; SKP, S-phase kinase-associated protein; SLF, S-locus F-box; SSI, sporophytic self-incompatibility; Ub, ubiquitin.

(Received June 2, 2009; Accepted July 1, 2009)
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