Altered expression levels of long non-coding natural antisense transcripts overlapping the UGT73C6 gene affect rosette size in Arabidopsis thaliana

Abstract

Natural antisense long non-coding RNAs (lncNATs) are involved in the regulation of gene expression in plants, modulating different relevant developmental processes and responses to various stimuli. We have identified and characterized two lncNATs (NAT1(UGT73C6) and NAT2(UGT73C6), collectively NATs(UGT73C6)) from Arabidopsis thaliana that are transcribed from a gene fully overlapping UGT73C6, a member of the UGT73C subfamily of genes encoding UDP-glycosyltransferases (UGTs). Expression of both NATs(UGT73C6) is developmentally controlled and occurs independently of the transcription of UGT73C6 in cis. Downregulation of NATs(UGT73C6) levels through artificial microRNAs results in a reduction of the rosette area, while constitutive overexpression of NAT1(UGT73C6) or NAT2(UGT73C6) leads to the opposite phenotype, an increase in rosette size. This activity of NATs(UGT73C6) relies on its RNA sequence and, although modulation of UGT73C6 in cis cannot be excluded, the observed phenotypes are not a consequence of the regulation of UGT73C6 in trans. The NATs(UGT73C6) levels were shown to affect cell proliferation and thus individual leaf size. Consistent with this concept, our data suggest that the NATs(UGT73C6) influence the expression levels of key transcription factors involved in regulating leaf growth by modulating cell proliferation. These findings thus reveal an additional regulatory layer on the process of leaf growth. In this work, we characterized at the molecular level two long non-coding RNAs (NATs(UGT73C6)) that are transcribed in the opposite direction to UGT73C6, a gene encoding a glucosyltransferase involved in brassinosteroid homeostasis in A. thaliana. Our results indicate that NATs(UGT73C6) expression influences leaf growth by acting in trans and by modulating the levels of transcription factors that are involved in the regulation of cell proliferation.