Intron-retained alternatively spliced VIN3-LIKE 2 gene from Chimonanthus praecox promotes flowering in transgenic Arabidopsis

Abstract

Vernalization is a process of acquiring or accelerating the flowering ability by prolonged cold exposure. VERNALIZATION INSENSITIVE3 (VIN3) is induced by chilling and is extremely important for the vernalization response in Arabidopsis thaliana. However, the issue of the induction of the VIN3-LIKE genes in wintersweet (Chimonanthus praecox) has been largely neglected. In the present study, we explored the molecular regulation of the PHD type finger protein-encoding gene CpVIL2 in relation to the growth and development of wintersweet in Arabidopsis. In wintersweet, quantitative real-time PCR (qRT-PCR) analysis showed that the relative expression of CpVIL2-As2i (intron-retained alternatively spliced in the second intron) was extremely higher in the pistils than in the other tissues. And the relative CpVIL2-As2i expression in flower buds (FBs) treated at 8 °C was higher than that of FBs in December, 2016 under natural conditions, which was not detected in non-flowering FBs at 16 °C. In Arabidopsis, the expression patterns of the CpVIL2-As2i gene were detected at first in CpVIL2-As2i pro::GUS (β-glucuronidase) lines, with predominantly higher expression in flowers and inflorescence. Meanwhile, the hormone-induced expression profiles of the CpVIL2-As2i promoter were confirmed using exogenous induction by abscisic acid (ABA) and indole acetic acid (IAA) phytohormones, where the GUS enzyme activity obviously decreased compared with that of control. In comparison with Arabidopsis/Col-0, early flowering was detected in ectopic 35S::CpVIL2-As2i lines. Overall, these results demonstrated the function of the CpVIL2-As2i gene, at the same time, provided us with new insights into the molecular mechanisms of early flowering and complex regulatory networks of vernalization in wintersweet.