title: Functional characterization of two chimeric proteins between a Petunia inflata S-locus F-box protein, PiSLF2, and a PiSLF-like protein, PiSLFLb-S2.
abstract: Self-incompatible solanaceous species possess the S-RNase and SLF (S-locus F-box) genes at the highly polymorphic S-locus, and their products mediate S-haplotype-specific rejection of pollen tubes in the style. After a pollen tube grows into the style, the S-RNases produced in the style are taken up; however, only self S-RNase (product of the matching S-haplotype) can inhibit the subsequent growth of the pollen tube. Based on the finding that non-self interactions between PiSLF (Petunia inflata SLF) and S-RNase are stronger than self-interactions, and based on the biochemical properties of PiSLF, we previously proposed that a PiSLF preferentially interacts with its non-self S-RNases to mediate their ubiquitination and degradation, thereby only allowing self S-RNase to exert its cytotoxic function. We further divided PiSLF into three potential Functional Domains (FDs), FD1-FD3, based on sequence comparison of PiSLF and PiSLF-like proteins, and based on S-RNase-binding properties of these proteins and various truncated forms of PiSLF(2) (S(2) allelic variant of PiSLF). In this work, we examined the in vivo function of FD2, which we proposed to be responsible for strong, general interactions between PiSLF and S-RNase. We swapped FD2 of PiSLF(2) with the corresponding region of PiSLFLb-S(2) (S(2) allelic variant of a PiSLF-like protein), and expressed GFP-fused chimeric proteins, named b-2-b and 2-b-2, in S(2) S(3) transgenic plants. We showed that neither chimeric protein retained the SI function of PiSLF(2), suggesting that FD2 is necessary, but not sufficient, for the function of PiSLF. Moreover, since we previously found that b-2-b and 2-b-2 only interacted with S(3)-RNase ~50 and ~30%, respectively, as strongly as did PiSLF(2) in vitro, their inability to function as PiSLF(2) is also consistent with our model predicating on strong interaction between a PiSLF and its non-self S-RNases as part of the biochemical basis for S-haplotype-specific rejection of pollen tubes.
authors.affiliation: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, 403 Althouse Lab, University Park, PA 16802, USA.
authors.forename: Allison M
authors.initials: AM
authors.lastname: Fields
authors.forename: Ning
authors.initials: N
authors.lastname: Wang
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Hua
authors.forename: Xiaoying
authors.initials: X
authors.lastname: Meng
authors.forename: Teh-Hui
authors.initials: TH
authors.lastname: Kao
mesh_terms: D000595:Amino Acid Sequence; D018744:DNA, Plant; D018506:Gene Expression Regulation, Plant / Q000502:physiology; D018745:Genome, Plant; D049452:Green Fluorescent Proteins; D008969:Molecular Sequence Data; D032306:Petunia / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D053205:Pollen Tube / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D018744:DNA, Plant; D010940:Plant Proteins; D049452:Green Fluorescent Proteins