XBAT35, a novel Arabidopsis RING E3 ligase exhibiting dual targeting of its splice isoforms, is involved in ethylene-mediated regulation of apical hook curvature.
The Arabidopsis XBAT35 is one of five structurally related ankyrin repeat-containing Really Interesting New Gene (RING) E3 ligases involved in ubiquitin-mediated protein degradation, which plays key roles in a wide range of cellular processes. Here, we show that the XBAT35 gene undergoes alternative splicing, generating two transcripts that are constitutively expressed in all plant tissues. The two splice variants derive from an exon skipping event that excludes an in-frame segment from the XBAT35 precursor mRNA, giving rise to two protein isoforms that differ solely in the presence of a nuclear localization signal (NLS). Transient expression assays indicate that the isoform lacking the NLS localizes in the cytoplasm of plant cells, whereas the other is targeted to the nucleus, accumulating in nuclear speckles. Both isoforms are functional E3 ligases, as assessed by in vitro ubiquitination assays. Two insertion mutant alleles and RNA-interference (RNAi) silencing lines for XBAT35 display no evident phenotypes under normal growth conditions, but exhibit hypersensitivity to the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC) during apical hook exaggeration in the dark, which is rescued by an inhibitor of ethylene perception. Independent expression of each XBAT35 splice variant in the mutant background indicates that the two isoforms may differentially contribute to apical hook formation but are both functional in this ethylene-mediated response. Thus, XBAT35 defines a novel player in ethylene signaling involved in negatively regulating apical hook curvature, with alternative splicing controlling dual targeting of this E3 ubiquitin ligase to the nuclear and cytoplasmic compartments.
Instituto Gulbenkian de Ciência, Rua da Quinta Grande, 6,  2780-156 Oeiras,  Portugal.
Sofia D
SD
Carvalho
Rita
R
Saraiva
Teresa M
TM
Maia
Isabel A
IA
Abreu
Paula
P
Duque
D017398:Alternative Splicing; D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D003593:Cytoplasm / Q000201:enzymology; D005030:Ethylenes / Q000378:metabolism*; D005091:Exons / Q000235:genetics; D018506:Gene Expression Regulation, Plant; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019913:Nuclear Localization Signals; D021381:Protein Transport; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D036226:Seedlings / Q000033:anatomy & histology* / Q000254:growth & development; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D005030:Ethylenes; D007527:Isoenzymes; D019913:Nuclear Localization Signals; D012333:RNA, Messenger; C036216:ethylene; D044767:Ubiquitin-Protein Ligases