Transcriptome-wide association mapping provides insights into the genetic basis and candidate genes governing flowering, maturity and seed weight in rice bean (Vigna umbellata).

BACKGROUND: Rice bean (Vigna umbellata), an underrated legume, adapts to diverse climatic conditions with the potential to support food and nutritional security worldwide. It is used as a vegetable, minor food crop and a fodder crop, being a rich source of proteins, minerals, and essential fatty acids. However, little effort has been made to decipher the genetic and molecular basis of various useful traits in this crop. Therefore, we considered three economically important traits i.e., flowering, maturity and seed weight of rice bean and identified the associated candidate genes employing an associative transcriptomics approach on 100 diverse genotypes out of 1800 evaluated rice bean accessions from the Indian National Genebank. RESULTS: The transcriptomics-based genotyping of one-hundred diverse rice bean cultivars followed by pre-processing of genotypic data resulted in 49,271 filtered markers. The STRUCTURE, PCA and Neighbor-Joining clustering of 100 genotypes revealed three putative sub-populations. The marker-trait association analysis involving various genome-wide association study (GWAS) models revealed significant association of 82 markers on 48 transcripts for flowering, 26 markers on 22 transcripts for maturity and 22 markers on 21 transcripts for seed weight. The transcript annotation provided information on the putative candidate genes for the considered traits. The candidate genes identified for flowering include HSC80, P-II PsbX, phospholipid-transporting-ATPase-9, pectin-acetylesterase-8 and E3-ubiquitin-protein-ligase-RHG1A. Further, the WRKY1 and DEAD-box-RH27 were found to be associated with seed weight. Furthermore, the associations of PIF3 and pentatricopeptide-repeat-containing-gene with maturity and seed weight, and aldo-keto-reductase with flowering and maturity were revealed. CONCLUSION: This study offers insights into the genetic basis of key agronomic traits in rice bean, including flowering, maturity, and seed weight. The identified markers and associated candidate genes provide valuable resources for future exploration and targeted breeding, aiming to enhance the agronomic performance of rice bean cultivars. Notably, this research represents the first transcriptome-wide association study in pulse crop, uncovering the candidate genes for agronomically useful traits.

HvMPK4 phosphorylates HvWRKY1 to enhance its suppression of barley immunity to powdery mildew fungus.

Mitogen-activated protein kinase (MAPK) cascades play important roles in disease resistance in model plant species. However, the functions of MAPK signaling pathways in crop disease resistance are largely unknown. Here we report the function of HvMKK1-HvMPK4-HvWRKY1 module in barley immune system. HvMPK4 is identified to play a negative role in barley immune response against Bgh, as virus-induced gene silencing of HvMPK4 results in enhanced disease resistance whilst stably overexpressing HvMPK4 leads to super-susceptibility to Bgh infection. Furthermore, the barley MAPK kinase HvMKK1 is found to specifically interact with HvMPK4, and the activated HvMKK1DD variant specifically phosphorylates HvMPK4 in vitro. Moreover, the transcription factor HvWRKY1 is identified to be a downstream target of HvMPK4 and phosphorylated by HvMPK4 in vitro in the presence of HvMKK1DD. Phosphorylation assay coupled with mutagenesis analyses identifies S122, T284, and S347 in HvWRKY1 as the major residues phosphorylated by HvMPK4. HvWRKY1 is phosphorylated in barley at the early stages of Bgh infection, which enhances its suppression on barley immunity likely due to enhanced DNA-binding and transcriptional repression activity. Our data suggest that the HvMKK1-HvMPK4 kinase pair acts upstream of HvWRKY1 to negatively regulate barley immunity against powdery mildew.

WRKY1 acts as a key component improving resistance against Alternaria solani in wild tomato, Solanum arcanum Peralta.

Early blight (EB), caused by Alternaria solani, is a major threat to global tomato production. In comparison with cultivated tomato (Solanum lycopersicum), a wild relative, S. arcanum exhibits strong resistance against EB. However, molecular cascades operating during EB resistance in wild or cultivated tomato plants are largely obscure. Here, we provide novel insight into spatio-temporal molecular events in S. arcanum against A. solani. Transcriptome and co-expression analysis presented 33-WRKYs as promising candidates of which 12 SaWRKYs displayed differential expression patterns in resistant and susceptible accessions during EB disease progression. Among these, SaWRKY1 exhibited induced expression with significant modulation in xyloglucan endotrans hydrolase 5 (XTH5) and MYB2 expressions that correlated with the disease phenotypes. Electro-mobility shift assay confirmed physical interaction of recombinant SaWRKY1 to SaXTH5 and SaMYB2 promoters. Comparative WRKY1 promoter analysis between resistant and susceptible plants revealed the presence of crucial motifs for defence mechanism exclusively in resistant accession. Additionally, many defence-related genes displayed significant expression variations in both the accessions. Further, WRKY1 overexpressing transgenic plants exhibited higher levels of EB resistance while RNAi silencing lines had increased susceptibility to A. solani with altered expression of XTH5 and MYB2. Overall, these findings demonstrate the positive influence of WRKY1 in improving EB resistance in wild tomato and this could be further utilized as a potential target through genetic engineering to augment protection against A. solani in crop plants.

Klebsiella sp. confers enhanced tolerance to salinity and plant growth promotion in oat seedlings (Avena sativa).

Plant growth and yield is adversely affected by soil salinity. Salt tolerant plant growth-promoting rhizobacteria (PGPR) strain IG 3 was isolated from rhizosphere of wheat plants. The isolate IG 3 was able to grow in presence of NaCl ranging from 0 to 20% in Luria Bertani medium. The present study was planned to evaluate the role of inoculation of PGPR strain IG 3 and its efficacy in augmenting salt tolerance in oat (Avena sativa) under NaCl stress (100mM). The physiological parameter such as shoot length, root length, shoot dry weight, root dry weight and relative water content (RWC) were remarkably higher in IG 3 inoculated plants in comparison to un-inoculated plants under NaCl stress. Similarly, the biochemical parameters such as proline content, electrolyte leakage and malondialdehyde (MDA) content and activities of antioxidant enzymes were analyzed and found to be notably lesser in IG 3 inoculated oat plants in contrast to un-inoculated plants under salt stress. Inoculation of IG 3 strain to oat seedlings under salt stress positively modulated the expression profile of rbcL and WRKY1 genes. Root colonization of root surface and interior was demonstrated using scanning electron microscopy and tetrazolium staining, respectively. Due these outcomes, it could be implicated that inoculation of PGPR strain IG 3 enhanced plant growth under salt stress condition. This study demonstrates that PGPR play an imperative function in stimulating salt tolerance in plants and can be used as biofertilizer to enhance growth of crops in saline areas.

Cloning and sequence analysis of 10-deacetylbaccatin III-10-O-acetyl transferase gene and WRKY1 transcription factor from taxol-producing endophytic fungus Lasiodiplodia theobromea.

In this study, we have isolated an endophytic fungal strain Lasiodiplodia theobromae from non-Taxus host plant Piper nigrum. The strain L. theobromae identity was confirmed by morphological characteristics and internal transcribed spacer sequence analysis. Taxol produced by L. theobromae was observed to be identical to the authentic taxol as analyzed by chromatography and spectroscopy methods. The quantity of taxol produced by the fungus was estimated to be 247 µg L-1, and fungal taxol showed potent cytotoxic activity towards cancer cell line. Evidence to support the independent production of taxol by L. theobromea, the gene encoding 10-deacetylbacccation-III-O-acetyltransferase (DBAT), as well as, for the first time, open reading frame (ORF) of WRKY1 transcription factor (TF) were cloned and sequenced. The predicted amino sequence of L. theobromae dbat gene shared high homology with the taxol-producing plant and fungal dbat gene. Not only dbat gene, ORF of WRKY1 TF too shared high homology with Taxus chinensis WRKY1 TF ORF. To the best of our knowledge, this is the first report on cloning of dbat gene and its transcription factor from endophytes of non-Taxus host plant.