Auxin-related regulation of barley floral organ fate: genetic and phenotypic insights from tweaky spike mutants.

Mutant-based research is a cornerstone for elucidating gene function in barley (Hordeum vulgare) and other cereals. In this study, we exploited a unique collection of barley tweaky spike (tw) mutants to identify the locus controlling floret and spike development. We combined genetic mapping with structural variant and gene-expression analyses across a tw allelic series, supported by functional validation, and quantified endogenous auxin levels alongside auxin-application assays to test phytohormone involvement. Analysis of independent tw allelic mutants revealed that the phenotype results from loss of function of HvAP2L5, most often due to complete open reading frame (ORF) deletions, and, in one allele, due to transcriptional silencing. tw mutants showed a reproducible endogenous indole-3-acetic acid deficit, and exogenous auxin could partially phenocopy and/or rescue core tw-associated traits, indicating compromised auxin homeostasis. We conclude that HvAP2L5 is essential for normal spike/floret development and that its loss disrupts auxin balance. Auxin imbalance is a compelling working model that can account for most of the pleiotropic features of tw mutants.