Identification and Expression Analysis of Medicago truncatula Isopentenyl Transferase Genes (IPTs) Involved in Local and Systemic Control of Nodulation

Abstract

Cytokinins are essential for legume plants to establish a nitrogen-fixing symbiosis with rhizobia. Recently, the expression level of cytokinin biosynthesis IPTs (ISOPENTENYLTRANSFERASES) genes was shown to be increased in response to rhizobial inoculation in Lotus japonicus, Medicago truncatula and Pisum sativum. In addition to its well-established positive role in nodule primordium initiation in root cortex, cytokinin negatively regulates infection processes in the epidermis. Moreover, it was reported that shoot-derived cytokinin inhibits the subsequent nodule formation through AON (autoregulation of nodulation) pathway. In L. japonicus, LjIPT3 gene was shown to be activated in the shoot phloem via the components of AON system, negatively affecting nodulation. However, in M. truncatula, the detailed analysis of MtIPTs expression, both in roots and shoots, in response to nodulation has not been performed yet, and the link between IPTs and AON has not been studied so far. In this study, we performed an extensive analysis of MtIPTs expression levels in different organs, focusing on the possible role of MtIPTs in nodule development. MtIPTs expression dynamics in inoculated roots suggest that besides its early established role in the nodule primordia development, cytokinin may be also important for later stages of nodulation. According to expression analysis, MtIPT3, MtIPT4, and MtIPT5 are activated in the shoots in response to inoculation. Among these genes, MtIPT3 is the only one the induction of which was not observed in leaves of the sunn-3 mutant defective in CLV1-like kinase, the key component of AON, suggesting that MtIPT3 is activated in the shoots in an AON-dependent manner. Taken together, our findings suggest that MtIPTs are involved in the nodule development at different stages, both locally in inoculated roots and systemically in shoots, where their expression can be activated in an AON-dependent manner.

Local and Systemic Effect of Cytokinins on Soybean Nodulation and Regulation of Their Isopentenyl Transferase (IPT) Biosynthesis Genes Following Rhizobia Inoculation

“…The BLAST searches were broadened to include the databases for common bean, L. japonicus , M. truncatula and the non-legume Arabidopsis thaliana . This confirmed the presence of 23 putative IPT genes in M. truncatula , making it the largest gene family compared with the eight, six and nine members identified in common bean, L. japonicus and A. thaliana , respectively ( Takei et al, 2001 ; Azarakhsh et al, 2018 ) ( Supplementary Tables S3 – S6 ).…”

Responses of mature symbiotic nodules to the whole-plant systemic nitrogen signaling

“…Ethylene has been implicated in the negative control of nodule formation and nodule infection ( Guinel, 2015 ; Reid et al , 2018 ). Modulating shoot methyl jasmonate or cytokinin (CK) accumulation has suggested roles for these molecules in the systemic control of nodulation in Lotus japonicus ( Nakagawa and Kawaguchi, 2006 ; Kinkema and Gresshoff, 2008 ; Sasaki et al , 2014 ; Azarakhsh et al , 2018 ). CLE and CEP hormone peptides have been implicated in the long-distance control of nodule formation and AON as well as in root formation and nitrate acquisition ( Mortier et al ., 2012 ; Okamoto et al , 2016 ; Taleski et al , 2018 ; Laffont et al , 2019 ).…”

Lotus SHAGGY‐like kinase 1 is required to suppress nodulation in Lotus japonicus

“…In our study, the relative transcript levels of the cytokinin biosynthetic gene IPT3 were unaffected in lsk1‐1 roots. Medicago truncatula, MtIPT3 , is induced in the shoots in an AON‐dependent manner (Azarakhsh et al ., ), whereas in Glycine max (soybean) this induction is independent of AON (Mens et al ., ). Similarly, expression of the cytokinin receptor kinase, LHK1, and which is known to mediate cytokinin signals during nodulation (Held et al ., ), and which was recently reported to link miR2111 to AON (Tsikou et al ., ), as well as CKX3, a primarily tZ‐type cytokinin‐catabolizing enzyme, were also unaltered in LSK1 knock‐down plants.…”