Pirin1 Is a Non-Circadian Regulated Transcript and Protein, but Highly Responsive to Light/Dark Periods in the Seed-to-Seedling Transition in Arabidopsis thaliana

Abstract

As non-motile organisms, plants must respond to constantly changing environmental conditions through dynamic means. The Arabidopsis thaliana Pirin1 (PRN1) is an enzyme that regulates the UV-screening compound quercetin and exhibits multiple light-dependent mutant phenotypes. However, regulation of the PRN1 gene or protein is not well studied. Herein, we investigated the light regulation of expression of the PRN1 transcript and protein early in development under multiple light conditions (constant darkness, constant white light, 12:12, 16:8, and circadian conditions). PRN1 transcript was more highly expressed in continuous darkness than in continuous light or 16:8 conditions in 7-day seedlings. However, in these identical samples, protein detected by a specific antibody revealed quite different protein band patterns, with larger bands than that observed for in vitro-translated PRN1 alone. In entrainment studies, exploration of 12:12 expression indicated that transcript generally was reduced in “evening” of the photoperiod. PRN1 protein expression revealed multiband patterns, varying over time and light conditions. PRN1 transcript did not display typical diurnal or circadian regulation; protein from the same samples indicated unique patterns in the free-running light versus free-running dark. Purified in vitro-translated PRN1 protein briefly treated with cell extract produced higher molecular mass bands, indicating posttranslational modification. prn1 mutant data indicated that full transcript was not produced, and any protein resulting was likely to be targeted to the proteasome; data indicated that for wild-type (WT) PRN1, the lower molecular weight bands (~36–40 kD) were likely active protein. RNA-seq analysis of 6-day-old dark-grown WT versus prn1 mutant seedlings indicated few significantly altered genes. High sequence conservation of PRN1 among plants indicates that Arabidopsis is a model system to understand its regulation.