CS-20-1 - Rootless1 Is a Putative bHLH Transcription Factor Modulating Post-embryonic Crown Root Development in Maize

Root system architecture in cereals consists of crown roots (CRs), which are critical for water and nutrient uptake efficiencies. Yet the molecular and genetic control of CR development remains obscure. The classic maize (Zea mays) mutant rootless1 (rt1) identified by Jenkins (1930) had almost no crown roots. 90 years later, we have identified the gene controlling rt1 as a bHLH transcription factor using Ac/Ds mutant alleles and complementation experiments with rt1. Ds mutants exhibit a reduction of only aboveground CRs, compared to the more severe phenotype of the original mutant. Nanopore sequencing of rt1 revealed an Indel in the 5’ region of ZmRt1 leading to its reduced expression. Histology of the seedling coleoptilar node showed that rt1 mutants appear to initiate fewer CRs than wild-type. In situ hybridizations of wild-type nodes show diffuse expression in initiating CR primordia, but a highly specific expression in the endodermal cell file in emerging primordia, suggesting roles in both initiation and development of CRs. This pattern overlaps with SCARECROW1 (ZmScr1), but is distinct from WUSCHEL-RELATED HOMEOBOX 5. rt1 mutants also displayed stunted growth of primary roots, indicating that ZmRt1 is involved in both CR and primary root development. Strikingly, rt1 mutants downregulated ZmScr1 expression, suggesting a potential role for ZmScr1 in ZmRt1-mediated CR development. Ongoing experiments seek to identify regulatory targets of Rt1, as well as co-expression patterns with other key organ initiation and growth regulators. Since QTL for CR number (CRN) overlapped with Rt1 in two separate studies with distinct germplasm, we next queried whether Rt1 could be involved in CRN quantitative variation and identified teosinte alleles of Rt1 that modulated CRN. The mounting body of evidence thus suggests that Rt1 is a major genetic regulator of CRs in maize.