U-box E3 ligases (PUBs) regulate plant self-incompatibility (SI), hormone signaling, and stress responses. By mining time-course transcriptome data from Brassica oleracea following self-pollination (0-60 min), we identified BoPUB3L, a PUB gene that is rapidly and specifically upregulated by self-pollination. The 2214 bp open reading frame encodes a 737-aa protein (81.27 kD; pI 6.04) containing a single U-box and five ARM repeats, lacking a signal peptide and transmembrane domain, and localizing to the nucleus. The BoPUB3L promoter contains multiple cis-acting elements related to light responsiveness, abscisic acid, auxin, and gibberellin responses, as well as meristem-associated expression. BoPUB3L is most highly expressed in the style, followed by sepals, petals, and buds, as confirmed by GUS staining. Transcript abundance increased continuously during the first 30 min after self-pollination and peaked at 15 min, reaching a level 15.65-fold higher than that after cross-pollination. Yeast two-hybrid, pull-down, and BiFC assays further showed that BoPUB3L interacts with the kinase domain of the S-locus receptor kinase (SRK). Together, these results identify BoPUB3L as a previously uncharacterized component of the SI response in B. oleracea and provide new insight into the molecular basis of self-incompatibility in crucifers.
Zuo et al. (Mon,) studied this question.