Prediction of the in planta Phakopsora pachyrhizi secretome and potential effector families.

Asian soybean rust (ASR), caused by the obligate biotrophic fungus Phakopsora pachyrhizi, can cause losses greater than 80%. Despite its economic importance, there is no soybean cultivar with durable ASR resistance. In addition, the P. pachyrhizi genome is not yet available. However, the availability of other rust genomes, as well as the development of sample enrichment strategies and bioinformatics tools, has improved our knowledge of the ASR secretome and its potential effectors. In this context, we used a combination of laser capture microdissection (LCM), RNAseq and a bioinformatics pipeline to identify a total of 36 350 P. pachyrhizi contigs expressed in planta and a predicted secretome of 851 proteins. Some of the predicted secreted proteins had characteristics of candidate effectors: small size, cysteine rich, do not contain PFAM domains (except those associated with pathogenicity) and strongly expressed in planta. A comparative analysis of the predicted secreted proteins present in Pucciniales species identified new members of soybean rust and new Pucciniales- or P. pachyrhizi-specific families (tribes). Members of some families were strongly up-regulated during early infection, starting with initial infection through haustorium formation. Effector candidates selected from two of these families were able to suppress immunity in transient assays, and were localized in the plant cytoplasm and nuclei. These experiments support our bioinformatics predictions and show that these families contain members that have functions consistent with P. pachyrhizi effectors.

Genomic and transcriptomic characterization of the transcription factor family R2R3-MYB in soybean and its involvement in the resistance responses to Phakopsora pachyrhizi.

Myb genes constitute one of the largest transcription factor families in the plant kingdom. Soybean MYB transcription factors have been related to the plant response to biotic stresses. Their involvement in response to Phakopsora pachyrhizi infection has been reported by several transcriptional studies. Due to their apparently highly diverse functions, these genes are promising targets for developing crop varieties resistant to diseases. In the present study, the identification and phylogenetic analysis of the soybean R2R3-MYB (GmMYB) transcription factor family was performed and the expression profiles of these genes under biotic stress were determined. GmMYBs were identified from the soybean genome using bioinformatic tools, and their putative functions were determined based on the phylogenetic tree and classified into subfamilies using guides AtMYBs describing known functions. The transcriptional profiles of GmMYBs upon infection with different pathogen were revealed by in vivo and in silico analyses. Selected target genes potentially involved in disease responses were assessed by RT-qPCR after different times of inoculation with P. pachyrhizi using different genetic backgrounds related to resistance genes (Rpp2 and Rpp5). R2R3-MYB transcription factors related to lignin synthesis and genes responsive to chitin were significantly induced in the resistant genotypes.

Genome-wide analysis of the Hsp20 gene family in soybean: comprehensive sequence, genomic organization and expression profile analysis under abiotic and biotic stresses.

BACKGROUND: The Hsp20 genes are associated with stress caused by HS and other abiotic factors, but have recently been found to be associated with the response to biotic stresses. These genes represent the most abundant class among the HSPs in plants, but little is known about this gene family in soybean. Because of their apparent multifunctionality, these proteins are promising targets for developing crop varieties that are better adapted to biotic and abiotic stresses. Thus, in the present study an in silico identification of GmHsp20 gene family members was performed, and the genes were characterized and subjected to in vivo expression analysis under biotic and abiotic stresses. RESULTS: A search of the available soybean genome databases revealed 51 gene models as potential GmHsp20 candidates. The 51 GmHsp20 genes were distributed across a total of 15 subfamilies where a specific predicted secondary structure was identified. Based on in vivo analysis, only 47 soybean Hsp20 genes were responsive to heat shock stress. Among the GmHsp20 genes that were potentials HSR, five were also cold-induced, and another five, in addition to one GmAcd gene, were responsive to Meloidogyne javanica infection. Furthermore, one predicted GmHsp20 was shown to be responsive only to nematode infection; no expression change was detected under other stress conditions. Some of the biotic stress-responsive GmHsp20 genes exhibited a divergent expression pattern between resistant and susceptible soybean genotypes under M. javanica infection. The putative regulatory elements presenting some conservation level in the GmHsp20 promoters included HSE, W-box, CAAT box, and TA-rich elements. Some of these putative elements showed a unique occurrence pattern among genes responsive to nematode infection. CONCLUSIONS: The evolution of Hsp20 family in soybean genome has most likely involved a total of 23 gene duplications. The obtained expression profiles revealed that the majority of the 51 GmHsp20 candidates are induced under HT, but other members of this family could also be involved in normal cellular functions, unrelated to HT. Some of the GmHsp20 genes might be specialized to respond to nematode stress, and the predicted promoter structure of these genes seems to have a particular conserved pattern related to their biological function.