Comparison ofPetunia inflata S-Locus F-Box Protein (Pi SLF) with Pi SLF–Like Proteins Reveals Its Unique Function in S-RNase–Based Self-Incompatibility

Abstract

Petunia inflata possesses S-RNase-based self-incompatibility (SI), which prevents inbreeding and promotes outcrossing. Two polymorphic genes at the S-locus, S-RNase and P. inflata S-locus F-box (Pi SLF), determine the pistil and pollen specificity, respectively. To understand how the interactions between Pi SLF and S-RNase result in SI responses, we identified four Pi SLF-like (Pi SLFL) genes and used them, along with two previously identified Pi SLFLs, for comparative studies with Pi SLF 2 . We examined the in vivo functions of three of these Pi SLFLs and found that none functions in SI. These three Pi SLFLs and two other Pi SLFs either failed to interact with S 3 -RNase (a non-self S-RNase for all of them) or interacted much more weakly than did Pi SLF 2 in vitro. We divided Pi SLF 2 into FD1 (for Functional Domain1), FD2, and FD3, each containing one of the Pi SLF-specific regions, and used truncated Pi SLF 2 , chimeric proteins between Pi SLF 2 and one of the Pi SLFLs that did not interact with S 3 -RNase, and chimeric proteins between Pi SLF 1 and Pi SLF 2 to address the biochemical roles of these three domains. The results suggest that FD2, conserved among three allelic variants of Pi SLF, plays a major role in the strong interaction with S-RNase; additionally, FD1 and FD3 (each containing one of the two variable regions of Pi SLF) together negatively modulate this interaction, with a greater effect on interactions with self S-RNase than with non-self S-RNases. A model for how an allelic product of Pi SLF determines the fate of its self and non-self S-RNases in the pollen tube is presented.

NaStEP: A Proteinase Inhibitor Essential to Self-Incompatibility and a Positive Regulator of HT-B Stability inNicotiana alataPollen Tubes

“…(1) The S-RNase degradation model (Hua and Kao, 2006;Hua et al, 2007Hua et al, , 2008Kubo et al, 2010) focuses on S-RNase-SLF interactions that bring about preferential nonself S-RNase degradation. In this model, strong nonself S-RNase-SLF interactions lead to the degradation of nonself S-RNases by the ubiquitin-26S proteasome system, allowing pollen tubes to escape from its cytotoxic effect.…”

DNA Markers and the Molecular Mechanism of Self-(in)compatibility in Japanese Pear (<i>Pyrus pyrifolia</i> Nakai)

“…Similarly, 16-20 S-locus F-box (SLF) genes were identified in each S-haplotype in Petunia, a member of the Solanaceae family that exhibits the same S-RNasebased GSI as Japanese pear (Hua et al, 2007;Kubo et al, 2010Kubo et al, , 2015McCubbin et al, 2000;Wang et al, 2003Wang et al, , 2004Williams et al, 2014a). These SLFs were classified into 18 types (SLF1-SLF18) based on their phylogenetic relationships (Kubo et al, 2015).…”

Protein Interactions in S-RNase-Based Gametophytic Self-Incompatibility

“…Chromosome-walking, differential display or degenerate PCR-cloning approaches Wang et al, 2003Wang et al, ,2004Wheeler & Newbigin 2007;Zhou et al, 2003) in the Solanaceae and Plantaginaceae identified a number of SLF-like genes (SLFL) that were linked to the S-locus. These genes were thought not to be involved in self-incompatibility interactions specifically, since they did not show interaction with known S-RNases nor did they exhibit competitive interaction in transgenic assays (Hua et al, 2007;Meng et al, 2011). Hua et al, (2007) attempted to identify domains of SLF proteins that governed allelic specificity by iterative pairwise comparisons of SLF proteins with SLFL proteins.…”

“…These genes were thought not to be involved in self-incompatibility interactions specifically, since they did not show interaction with known S-RNases nor did they exhibit competitive interaction in transgenic assays (Hua et al, 2007;Meng et al, 2011). Hua et al, (2007) attempted to identify domains of SLF proteins that governed allelic specificity by iterative pairwise comparisons of SLF proteins with SLFL proteins. These comparisons identified three "SLFspecific" regions SR1, SR2 and SR3.…”

“…SBP1 (Hua & Kao 2006;Sims & Ordanic 2001;Sims et al, 2010) is a RING-HC protein. RING proteins are E3 ubiquitin ligases (Deshaies & Joazeiro 2009;Freemont 2000), and SBP1 has E3 ubiquitin ligase activity in vitro (Hua et al, 2007;Sims unpublished). AhSSK1 (Huang et al, 2006) and PhSSK1 are SKP1-like proteins (SKP1 is a scaffold component of SCF E3 ligases).…”