Barley (Hordeum vulgare L.) β-Tubulin gene HvTUBULIN8 has pleiotropic effects on spike and leaf development

Abstract Barley lateral spikelet morphology and grain shape play a key role in grain physical quality and yield. Several genes and QTLs for these traits have been cloned or fine mapped previously. Here, we report the phenotypic and genotypic analysis of a barley mutant with Round Lateral Spikelet (rls) from c.v. Edamai 934. The mutant had round lateral spikelet, short but round grain, shortened awn, thick glume and dark green leaves. Histocytologic and ultrastructural analysis revealed that the difference of grain shape of rls was caused by change of cell arrangement in glume, and the dark leaf color resulted from enlarged chloroplast. HvTUBULIN8 (HvTUB8) was identified as the functional gene for rls by combination of RNA-seq, map-based cloning, virus-induced-gene-silencing (VIGS) and protein subcellular location. A single G-A substitution in exon of HvTUB8 resulted in change of Cysteine to Tyrosine. Furthermore, the mutant isoform of HvTUB8 could be detected in both nucleus and cytoplasm, whereas the wild was only in cytoplasm of wheat protoplasts. Being consistent with the rare phenotype, the “A” allele of HvTUB8 was only detected in rls, but not in a worldwide barley germplasm panel consisting about 400 accessions. VIGS confirmed that HvTUB8 was essential to maintain spike integrity. RNA-sequencing results suggest that HvTUB8 control spike morphogenesis via ethylene homeostasis and signaling, and control leaf color through chlorophyll metabolism. Collectively, our results demonstrated that HvTUB8 has pleiotropic effects on barley spike and leaf morphology and provided insight of a novel mechanism of HvTUB8 in spike and leaf development.