A Putative Type III Secretion System Effector Encoded by the MA20_12780 Gene in Bradyrhizobium japonicum Is-34 Causes Incompatibility with Rj ₄ Genotype Soybeans

Abstract

eThe nodulation of Bradyrhizobium japonicum Is-34 is restricted by Rj 4 genotype soybeans (Glycine max). To identify the genes responsible for this incompatibility, Tn5 mutants of B. japonicum Is-34 that were able to overcome this nodulation restriction were obtained. Analysis of the Tn5 mutants revealed that Tn5 was inserted into a region containing the MA20_12780 gene. In addition, direct disruption of this gene using marker exchange overcame the nodulation restriction by Rj 4 genotype soybeans. The MA20_12780 gene has a tts box motif in its upstream region, indicating a possibility that this gene encodes a type III secretion system (T3SS) effector protein. Bioinformatic characterization revealed that the MA20_12780 protein contains the small ubiquitin-like modifier (SUMO) protease domain of the C48 peptidase (ubiquitin-like protease 1 [Ulp1]) family. The results of the present study indicate that a putative T3SS effector encoded by the MA20_12780 gene causes the incompatibility with Rj 4 genotype soybeans, and they suggest the possibility that the nodulation restriction of B. japonicum Is-34 may be due to Rj 4 genotype soybeans recognizing the putative T3SS effector (MA20_12780 protein) as a virulence factor. S oybeans (Glycine max) are an important agricultural crop and are well known for establishing a symbiosis with nitrogenfixing bacteria (rhizobia), such as Bradyrhizobium and Ensifer species (1). The symbionts form nodules in the roots of the soybean plant and provide fixed nitrogen to the soybean. In return, the soybean provides carbohydrates to the symbionts (2). The nodulation of the symbionts is controlled by soybean genes that are referred to as Rj or rj genes. To date, eight Rj or rj genes (rj 1 , Rj 2 , Rfg1, Rj 3 , Rj 4 , rj 5 , rj 6 , and rj 7 ) have been identified (3). For example, the dominant Rj 4 gene in soybeans restricts the nodulation of specific strains of Bradyrhizobium bacteria (e.g., Bradyrhizobium japonicum Is-34 and Bradyrhizobium elkanii USDA 61) almost completely (3-5). The Rj 4 genotype may be an important trait of soybeans in the agricultural fields of Japan, because Glycine max cv. Fukuyutaka, harboring the Rj 4 gene, is the most cultivated cultivar in Japan, according to the 2012 statistics of the Ministry of Agriculture, Forestry, and Fisheries (MAFF) of Japan (http://www .maff.go.jp/j/seisan/ryutu/daizu/d_data/pdf/010.pdf [in Japanese; accessed January 2015]), and 24% of soybeans introduced into the United States from Japan exhibited the Rj 4 genotype (6). In addition, Ͼ60% of soybean cultivars introduced from Southeast Asia had the Rj 4 gene (6, 7). One possible explanation for the high abundance of the Rj 4 genotype in soybeans is its ability to restrict the nodulation of B. elkanii strains. B. elkanii strains are considered to be poor symbiotic partners of soybeans (7), because rhizobitoxine, produced by B. elkanii, induces chlorosis in soybeans (6). Recently, a product of the Rj 4 gene was identified as a thaumatin-like protein (TLP) (4, 7). Products of all the Rj (or r...

High-quality permanent draft genome sequence of the Bradyrhizobium elkanii type strain USDA 76T, isolated from Glycine max (L.) Merr

“…The Type III secretion system (T3SS) [55] can either promote or impair the establishment of symbiosis, depending on the legume host [56], and has been characterized as a host determinant for rj1, Rfg1, Rj2 and Rj4 soybean cultivars [33, 57, 58]. The dominant soybean genes Rj2 and Rj4 restrict nodulation with specific strains of Bradyrhizobium [33].…”

NopD of Bradyrhizobium sp. XS1150 Possesses SUMO Protease Activity

“…The C-terminal regions of the BEL2_5 (USDA61) and MA20_12780 (Is-34) proteins are related to NopD (Figure 1B) and thus are predicted to possess SUMO protease activity. Mutant strains lacking these proteins gained the ability to induce nodules on Rj4/Rj4 soybeans, suggesting that NopD family proteins possess asymbiotic effector activity on soybean genotypes expressing Rj4 (Faruque et al, 2015;Tsurumaru et al, 2015;Yasuda et al, 2016). Rj4 encodes a specific thaumatin-like protein that only differs in few amino acids from homologs (Tang et al, 2016).…”

Identification of a gene encoding a sugar transporter as a potential target of a novel LysR‐type transcriptional regulator rluD from rhizobium required for symbiotic nodulation in soybean (Glycine max Merr.)

“…Although several studies have reported mapping nodulation‐related traits or biological nitrogen fixation traits in legumes, the candidate genes that are most likely involved in symbiotic nodulation have not been studied (Diaz et al., ; Heilig et al., ; Kamfwa et al., ). The candidate genes for nodule‐related QTLs that were confirmed and cloned were Rfg1 , Rj2 and Rj4 (Tang et al., ; Tsurumaru et al., ), which were a variety of dominant genes shown to restrict nodulation in soybean, especially by Brandyrhizobium (Sugawara et al., ; Tang et al., ). However, mining candidate genes by QTL mapping involved in symbiosis signal pathway transmission for revealing symbiotic mechanisms was also considerable and efficient.…”