A 5.5-kb LTR-retrotransposon insertion inside phytochrome B gene (CsPHYB) results in long hypocotyl and early flowering in cucumber (Cucumis sativus L.)

Abstract

The novel spontaneous long hypocotyl and early flowering (lhef) mutation in cucumber is due to a 5551-bp LTR-retrotransposon insertion in CsPHYB gene encoding PHYTOCHROME B, which plays a major role in regulating photomorphogenic hypocotyl growth and flowering. Hypocotyl length and flowering time are important for establishing high-quality seedlings in modern cucumber production, but little is known for the underlying molecular mechanisms of these two traits. In this study, a spontaneous cucumber long hypocotyl and early flowering mutant was identified and characterized. Based on multiple lines of evidence, we show that cucumber phytochrome B (CsPHYB) is the candidate gene for this mutation, and a 5551-bp LTR-retrotransposon insertion in the first exon of CsPHYB was responsible for the mutant phenotypes. Uniqueness of the mutant allele at CsPHYB was verified in 114 natural cucumber lines. Ectopic expression of the CsPHYB in Arabidopsis phyB mutant rescued the long hypocotyl and early flowering phenotype of phyB-9 mutant. The wild-type CsPHYB protein was localized on the membrane and cytoplasm under white light condition, whereas in the nucleus under red light, it is consistent with its roles as a red-light photoreceptor in Arabidopsis. However, the mutant csphyb protein was localized on the membrane and cytoplasm under both white and red-light conditions. Expression dynamics of CsPHYB and several cell elongation-related genes were positively correlated with hypocotyl elongation; the transcription levels of key positive and negative regulators for flowering time were also consistent with the anthesis dates in the mutant and wild-type plants. Yeast two hybrid and bimolecular fluorescence complementation assays identified physical interactions between CsPHYB and phytochrome interacting factor 3/4 (CsPIF3/4). These findings will provide new insights into the roles of the CsPHYB in cucumber hypocotyl growth and flowering time.