The Sinorhizobium (Ensifer) fredii HH103 Nodulation Outer Protein NopI Is a Determinant for Efficient Nodulation of Soybean and Cowpea Plants

Abstract

The type III secretion system (T3SS) is a specialized secretion apparatus that is commonly used by many plant and animal pathogenic bacteria to deliver proteins, termed effectors, to the interior of the host cells. These effectors suppress host defenses and interfere with signal transduction pathways to promote infection. Some rhizobial strains possess a functional T3SS, which is involved in the suppression of host defense responses, host range determination, and symbiotic efficiency. The analysis of the genome of the broad-host-range rhizobial strain Sinorhizobium fredii HH103 identified eight genes that code for putative T3SS effectors. Three of these effectors, NopL, NopP, and NopI, are Rhizobium specific. In this work, we demonstrate that NopI, whose amino acid sequence shows a certain similarity with NopP, is secreted through the S. fredii HH103 T3SS in response to flavonoids. We also determined that NopL can be considered an effector since it is directly secreted to the interior of the host cell as demonstrated by adenylate cyclase assays. Finally, the symbiotic phenotype of single, double, and triple nopI, nopL, and nopP mutants in soybean and cowpea was assayed, showing that NopI plays an important role in determining the number of nodules formed in both legumes and that the absence of both NopL and NopP is highly detrimental for symbiosis.IMPORTANCE The paper is focused on three Rhizobium-specific T3SS effectors of Sinorhizobium fredii HH103, NopL, NopP, and NopI. We demonstrate that S. fredii HH103 is able to secrete through the T3SS in response to flavonoids the nodulation outer protein NopI. Additionally, we determined that NopL can be considered an effector since it is secreted to the interior of the host cell as demonstrated by adenylate cyclase assays. Finally, nodulation assays of soybean and cowpea indicated that NopI is important for the determination of the number of nodules formed and that the absence of both NopL and NopP negatively affected nodulation. KEYWORDS:Sinorhizobium fredii, cowpea, effector, nodulation, soybean, symbiosis, type III secretion S oil bacteria, collectively known as rhizobia, induce the formation of specialized structures, called nodules, on the roots or stems of host legumes. Within these nodules, rhizobia differentiate into bacteroids that are able to reduce atmospheric nitrogen to ammonia, which is assimilated by the plant. In exchange, rhizobia are provided with a carbon source and an appropriate environment to promote bacterial growth (1).

Diversity of phytobeneficial traits revealed by whole‐genome analysis of worldwide‐isolated phenazine‐producing Pseudomonas spp.

“…or Erwinia spp., to mediate the translocation of effectors into plant cells to manipulate host defence responses (Alfano and Collmer, ). T3SSs are also used by several rhizobia to deliver nodulation outer proteins (Nops) into plant cells (Deakin and Broughton, ), which can suppress host immune response (Jiménez‐Guerrero et al, ) and contribute to host range determination (Yasuda et al, ) and nodulation efficiency (Jiménez‐Guerrero et al, ). T3SSs are present in numerous plant‐beneficial Pseudomonas spp.…”

Formulation of a Highly Effective Inoculant for Common Bean Based on an Autochthonous Elite Strain of Rhizobium leguminosarum bv. phaseoli, and Genomic-Based Insights Into Its Agronomic Performance

“…Rhizobia with a functional T3SS (R. etli CFN42 T , S. fredii HH103, B. diazoefficiens USDA110, or Rhizobium sp. NGR234) secrete nodulation outer proteins (Nops) in the presence of flavonoids, inducing the transcription of nodulation genes (Jiménez-Guerrero et al, 2017).…”

Transcriptomic Studies of the Effect of nod Gene-Inducing Molecules in Rhizobia: Different Weapons, One Purpose

“…The rest of genes controlled by the remaining TB code for effector proteins, namely nopC , nopD , nopI , nopM , nopM2 , nopL , nopP , and nopT ( Table S4 ) ( Figure 5 ). In fact, many of the proteins encoded by these genes were previously detected in supernatants of S. fredii HH103 obtained from cultures induced with genistein [ 117 , 118 ]. These findings indicate that, in the presence of inducing flavonoids, this bacterium produce a fully functional T3SS.…”