Medicago truncatula Genome Encoded Secreted Peptides Affect Host-Rhizobia Interactions

Plant hormones are endogenous signaling compounds that regulate plant growth and development. A recent surge in interest in small secreted peptide hormones has uncovered their roles in symbiotic nodule development (CEP family) and autoregulation of nodule number (CLE family). However, a complete inventory of all peptides encoded in the newly annotated Medicago genome is missing; primarily because genome annotation pipelines often overlook short ORFs (Open Reading Frames). Therefore we re-annotated the latest version of the Medicago genome (V4.0 and V3.5) to specifically include genes encoding an N-terminal secretion signal with total length less than 200 amino acids. Further, we analyzed expression data from 118 RNA-seq libraries (See Lundquist P. poster for more info) to validate these predicted ORFs. To explore function, we selected peptide coding genes induced at least 1.5 fold under low nitrogen conditions and rhizobial infection. Only the candidate genes whose expression we confirmed by quantitative RT-PCR were then synthesized. These peptides were added to solid (agarose) plant growth media and their effects on nodulation tested after seven days. We have screened 28 novel peptides from 10 different families and found that some affect nodulation. We further found that these effects could be seen with as little as 10 nM of added peptide. These peptides did not affect the growth of rhizobia in culture or bacterial attachment to root hairs. Thus they appear to be novel regulators of nodule development.