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CRISPR/Cas9-mediated knockout of PiSSK1 reveals essential role of S-locus F-box protein-containing SCF complexes in recognition of non-self S-RNases during cross-compatible pollination in self-incompatible Petunia inflata.
Plant Reprod. 2018 06; 31(2):129-143.PR

Abstract

KEY MESSAGE

Function of Petunia PiSSK1. Self-incompatibility (SI), an inbreeding-preventing mechanism, is regulated in Petunia inflata by the polymorphic S-locus, which houses multiple pollen-specific S-locus F-box (SLF) genes and a single pistil-specific S-RNase gene. S 2-haplotype and S 3-haplotype possess the same 17 polymorphic SLF genes (named SLF1 to SLF17), and each SLF protein produced in pollen is assembled into an SCF (Skp1-Cullin1-F-box) E3 ubiquitin ligase complex. A complete suite of SLF proteins is thought to collectively interact with all non-self S-RNases to mediate their ubiquitination and degradation by the 26S proteasome, allowing cross-compatible pollination. For each SCFSLF complex, the Cullin1 subunit (named PiCUL1-P) and Skp1 subunit (named PiSSK1), like the F-box protein subunits (SLFs), are pollen-specific, raising the possibility that they also evolved specifically to function in SI. Here we used CRISPR/Cas9-meditated genome editing to generate frame-shift indel mutations in PiSSK1 and examined the SI behavior of a T 0 plant (S 2 S 3) with biallelic mutations in the pollen genome and two progeny plants (S 2 S 2) each homozygous for one of the indel alleles and not carrying the Cas9-containing T-DNA. Their pollen was completely incompatible with pistils of seven otherwise-compatible S-genotypes, but fully compatible with pistils of an S 3 S 3 transgenic plant in which production of S3-RNase was completely suppressed by an antisense S 3-RNase gene, and with pistils of immature flower buds, which produce little S-RNase. These results suggest that PiSSK1 specifically functions in SI and support the hypothesis that SLF-containing SCF complexes are essential for compatible pollination.

Authors+Show Affiliations

Intercollege Graduate Degree Program in Plant Biology, The Pennsylvania State University, University Park, PA, 16802, USA.Intercollege Graduate Degree Program in Plant Biology, The Pennsylvania State University, University Park, PA, 16802, USA. txk3@psu.edu. Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA, 16802, USA. txk3@psu.edu.

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, Non-P.H.S.

Language

eng

PubMed ID

29192328

Citation

Sun, Linhan, and Teh-Hui Kao. "CRISPR/Cas9-mediated Knockout of PiSSK1 Reveals Essential Role of S-locus F-box Protein-containing SCF Complexes in Recognition of Non-self S-RNases During Cross-compatible Pollination in Self-incompatible Petunia Inflata." Plant Reproduction, vol. 31, no. 2, 2018, pp. 129-143.
Sun L, Kao TH. CRISPR/Cas9-mediated knockout of PiSSK1 reveals essential role of S-locus F-box protein-containing SCF complexes in recognition of non-self S-RNases during cross-compatible pollination in self-incompatible Petunia inflata. Plant Reprod. 2018;31(2):129-143.
Sun, L., & Kao, T. H. (2018). CRISPR/Cas9-mediated knockout of PiSSK1 reveals essential role of S-locus F-box protein-containing SCF complexes in recognition of non-self S-RNases during cross-compatible pollination in self-incompatible Petunia inflata. Plant Reproduction, 31(2), 129-143. https://doi.org/10.1007/s00497-017-0314-1
Sun L, Kao TH. CRISPR/Cas9-mediated Knockout of PiSSK1 Reveals Essential Role of S-locus F-box Protein-containing SCF Complexes in Recognition of Non-self S-RNases During Cross-compatible Pollination in Self-incompatible Petunia Inflata. Plant Reprod. 2018;31(2):129-143. PubMed PMID: 29192328.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - CRISPR/Cas9-mediated knockout of PiSSK1 reveals essential role of S-locus F-box protein-containing SCF complexes in recognition of non-self S-RNases during cross-compatible pollination in self-incompatible Petunia inflata. AU - Sun,Linhan, AU - Kao,Teh-Hui, Y1 - 2017/11/30/ PY - 2017/10/10/received PY - 2017/11/21/accepted PY - 2017/12/2/pubmed PY - 2018/9/28/medline PY - 2017/12/2/entrez KW - CRISPR/Cas9 KW - Petunia inflata KW - PiSSK1 KW - S-locus F-box protein KW - Self-incompatibility KW - Skp1-like protein SP - 129 EP - 143 JF - Plant reproduction JO - Plant Reprod VL - 31 IS - 2 N2 - KEY MESSAGE: Function of Petunia PiSSK1. Self-incompatibility (SI), an inbreeding-preventing mechanism, is regulated in Petunia inflata by the polymorphic S-locus, which houses multiple pollen-specific S-locus F-box (SLF) genes and a single pistil-specific S-RNase gene. S 2-haplotype and S 3-haplotype possess the same 17 polymorphic SLF genes (named SLF1 to SLF17), and each SLF protein produced in pollen is assembled into an SCF (Skp1-Cullin1-F-box) E3 ubiquitin ligase complex. A complete suite of SLF proteins is thought to collectively interact with all non-self S-RNases to mediate their ubiquitination and degradation by the 26S proteasome, allowing cross-compatible pollination. For each SCFSLF complex, the Cullin1 subunit (named PiCUL1-P) and Skp1 subunit (named PiSSK1), like the F-box protein subunits (SLFs), are pollen-specific, raising the possibility that they also evolved specifically to function in SI. Here we used CRISPR/Cas9-meditated genome editing to generate frame-shift indel mutations in PiSSK1 and examined the SI behavior of a T 0 plant (S 2 S 3) with biallelic mutations in the pollen genome and two progeny plants (S 2 S 2) each homozygous for one of the indel alleles and not carrying the Cas9-containing T-DNA. Their pollen was completely incompatible with pistils of seven otherwise-compatible S-genotypes, but fully compatible with pistils of an S 3 S 3 transgenic plant in which production of S3-RNase was completely suppressed by an antisense S 3-RNase gene, and with pistils of immature flower buds, which produce little S-RNase. These results suggest that PiSSK1 specifically functions in SI and support the hypothesis that SLF-containing SCF complexes are essential for compatible pollination. SN - 2194-7961 UR - https://www.unboundmedicine.com/medline/citation/29192328/CRISPR/Cas9_mediated_knockout_of_PiSSK1_reveals_essential_role_of_S_locus_F_box_protein_containing_SCF_complexes_in_recognition_of_non_self_S_RNases_during_cross_compatible_pollination_in_self_incompatible_Petunia_inflata_ L2 - https://dx.doi.org/10.1007/s00497-017-0314-1 DB - PRIME DP - Unbound Medicine ER -