Abstract
Plant-specific LOB-domain (LBD) transcription factors are crucial in post-embryonic root initiation. In cereals, the fibrous root system comprises primary and seminal roots that develop during embryogenesis and lateral- and crown roots that develop post-embryonically from root or stem, respectively. In rice, the CROWNROOTLESS1 (CRL1) LBD transcription factor is the core regulator of crown root initiation and a direct target of the auxin response factor (ARF)-mediated auxin signaling pathway. Orthologs of CRL1 have been identified and characterized in several species, where their role in crown root initiation has been validated. In barley, we identified two genes phylogenetically closely related to the rice CRL1 genes that we named HvCRL1 and HvCRL1L1. Using a crown root inducible system (CRIS), we identified that both HvCRL1 and HvCRL1L1 are expressed in response to auxin during the early steps of crown root differentiation in stem base, with HvCRL1 transcripts being accumulated quickly during the first hour of treatment. Transient activation assays in rice proto- plast showed that HvCRL1 could bind the LBD-box, a consensus DNA sequence recognized by LBD transcription factors, whereas HvCRL1L1 did not. Both genes can partially complement the crl1 rice mutant. Loss-of-function mutation in each gene drastically impairs crown root formation in barley. These data show that HvCRL1 and HvCRL1L1 are both involved in the regulation of crown root formation in barley but that these two transcription factors likely act through distinct and complementary pathways in this developmental process.