Isolation and characterization of a newly discovered plant growth-promoting endophytic fungal strain from the genus Talaromyces.

In the Kandi zone of Punjab, India, root and rhizospheric soil samples were collected from the local vegetation near the Shivalik mountain foothills. Fifteen fungal colonies exhibiting distinct cultural morphology on Potato Dextrose Agar (PDA) plates were selected for plant-microbe interaction studies. Among these, the isolate HNB9 was identified as a nonpathogenic root colonizer. Morphological and molecular analyses confirmed HNB9 as Talaromyces albobiverticillius, characterized by the secretion of a red pigment as a secondary metabolite. Plants colonized with T. albobiverticillius HNB9 exhibited enhanced growth, manifesting in increased shoot and root length compared to untreated controls. This study unveiled the first evidence that a species from the Talaromyces genus, specifically T. albobiverticillius, possesses dual capabilities of root colonization and plant growth promotion. Moreover, HNB9 demonstrated the production of plant growth-regulating compounds like Indole Acetic Acid (IAA) and proficient solubilization of crucial nutrients (Phosphorous, Zinc, and Silica) through plate culture methods. This finding represents a significant contribution to the understanding of root-colonizing fungi with plant growth-promoting attributes, challenging the existing knowledge gap within the Talaromyces genus.

Interaction studies of Serendipita indica and Zhihengliuella sp. ISTPL4 and their synergistic role in growth promotion in rice.

Rapid urbanization and globalization demand increasing agricultural productivity. Soil nutrient supply capacity is continuously decreasing due to soil erosion, degradation, salt deposition, undesired element, metal deposition, water scarcity, and an uneven nutrient delivery system. Rice cultivation requires a large amount of water which is becoming detrimental due to these activities. There is a need to increase its productivity. Microbial inoculants are becoming increasingly important in achieving sustainable agricultural production systems. The current study was conducted to investigate the interaction between the root endophytic fungus Serendipita indica (S. indica) and the actinobacterium Zhihengliuella sp. ISTPL4 (Z. sp. ISTPL4) and their synergistic effects on the growth of rice (Oryza sativa L). Both S. indica and Z. sp. ISTPL4 showed positive interactions. Growth of S. indica was observed at different days after Z. sp. ISTPL4 inoculation, and stimulated growth of S. indica was observed when Z. sp. ISTPL4 was inoculated at 5 dafi (days after fungal inoculation). Z. sp. ISTPL4 promoted the growth of S. indica as it increased spore germination. Furthermore, confocal and scanning electron microscopy (SEM) analyses showed a 27% increase in the spore size of S. indica in the presence of Z. sp. ISTPL4. In a liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis increased production of alanine and glutamic acid was observed in their sequential co-culture as compared with individual cultures. Sequential inoculation of S. indica and Z. sp. ISTPL4 significantly increased the biochemical and physical characteristics of rice as compared with their individual inoculum. Biochemical parameters such as chlorophyll content, total soluble sugar, and flavonoid content in the rice increased by up to 57%, 47%, and 39%, respectively, in the presence of the combined inoculum of S. indica and Z. sp. ISTPL4. This will be the first study, to the best of our knowledge, which shows the fungus and actinobacterium interaction and their synergistic roles in the growth promotion of rice. Furthermore, this novel combination can also be used to boost the growth of other crops to increase the agricultural yield.

Biopriming with Piriformospora indica ameliorates cadmium stress in rice by lowering oxidative stress and cell death in root cells.

Piriformospora indica is known for plant growth promotion and abiotic stress alleviation potential in several agricultural crops. However, a systemic analysis is warranted to explore potential application of this important fungus to augment heavy metal tolerance in rice. The present study explores potential of P. indica in ameliorating the effect of cadmium (Cd) stress in rice cultivars N22 and IR64. Seedlings inoculated with P. indica recorded significantly higher root-shoot length and biomass as compared to non-inoculated plants under control and Cd stress, respectively. Moreover, P. indica inoculated stressed roots accumulated more Cd as compared to non-inoculated stressed roots in both the varieties. Interestingly, cell death and reactive oxygen species (ROS) accumulation were significantly lower in the inoculated plant roots as compare with non-inoculated roots under Cd stress. The results emphasized significantly higher accumulation of Cd in fungal spores could reduce ROS accumulation in root cells resulting in lower cell death.

Role of exchange protein directly activated by cAMP (EPAC1) in breast cancer cell migration and apoptosis.

Despite the current progress in cancer research and therapy, breast cancer remains the leading cause of mortality among half a million women worldwide. Migration and invasion of cancer cells are associated with prevalent tumor metastasis as well as high mortality. Extensive studies have powerfully established the role of prototypic second messenger cAMP and its two ubiquitously expressed intracellular cAMP receptors namely the classic protein kinaseA/cAMP-dependent protein kinase (PKA) and the more recently discovered exchange protein directly activated by cAMP/cAMP-regulated guanine nucleotide exchange factor (EPAC/cAMP-GEF) in cell migration, cell cycle regulation, and cell death. Herein, we performed the analysis of the Cancer Genome Atlas (TCGA) dataset to evaluate the essential role of cAMP molecular network in breast cancer. We report that EPAC1, PKA, and AKAP9 along with other molecular partners are amplified in breast cancer patients, indicating the importance of this signaling network. To evaluate the functional role of few of these proteins, we used pharmacological modulators and analyzed their effect on cell migration and cell death in breast cancer cells. Hence, we report that inhibition of EPAC1 activity using pharmacological modulators leads to inhibition of cell migration and induces cell death. Additionally, we also observed that the inhibition of EPAC1 resulted in disruption of its association with the microtubule cytoskeleton and delocalization of AKAP9 from the centrosome as analyzed by in vitro imaging. Finally, this study suggests for the first time the mechanistic insights of mode of action of a primary cAMP-dependent sensor, Exchange protein activated by cAMP 1 (EPAC1), via its interaction with A-kinase anchoring protein 9 (AKAP9). This study provides a new cell signaling cAMP-EPAC1-AKAP9 direction to the development of additional biotherapeutics for breast cancer.

A novel Pfs38 protein complex on the surface of Plasmodium falciparum blood-stage merozoites.

BACKGROUND: The Plasmodium genome encodes for a number of 6-Cys proteins that contain a module of six cysteine residues forming three intramolecular disulphide bonds. These proteins have been well characterized at transmission as well as hepatic stages of the parasite life cycle. In the present study, a large complex of 6-Cys proteins: Pfs41, Pfs38 and Pfs12 and three other merozoite surface proteins: Glutamate-rich protein (GLURP), SERA5 and MSP-1 were identified on the Plasmodium falciparum merozoite surface. METHODS: Recombinant 6-cys proteins i.e. Pfs38, Pfs12, Pfs41 as well as PfMSP-165 were expressed and purified using Escherichia coli expression system and antibodies were raised against each of these proteins. These antibodies were used to immunoprecipitate the native proteins and their associated partners from parasite lysate. ELISA, Far western, surface plasmon resonance and glycerol density gradient fractionation were carried out to confirm the respective interactions. Furthermore, erythrocyte binding assay with 6-cys proteins were undertaken to find out their possible role in host-parasite infection and seropositivity was assessed using Indian and Liberian sera. RESULTS: Immunoprecipitation of parasite-derived polypeptides, followed by LC-MS/MS analysis, identified a large Pfs38 complex comprising of 6-cys proteins: Pfs41, Pfs38, Pfs12 and other merozoite surface proteins: GLURP, SERA5 and MSP-1. The existence of such a complex was further corroborated by several protein-protein interaction tools, co-localization and co-sedimentation analysis. Pfs38 protein of Pfs38 complex binds to host red blood cells (RBCs) directly via glycophorin A as a receptor. Seroprevalence analysis showed that of the six antigens, prevalence varied from 40 to 99%, being generally highest for MSP-165 and GLURP proteins. CONCLUSIONS: Together the data show the presence of a large Pfs38 protein-associated complex on the parasite surface which is involved in RBC binding. These results highlight the complex molecular interactions among the P. falciparum merozoite surface proteins and advocate the development of a multi-sub-unit malaria vaccine based on some of these protein complexes on merozoite surface.

Proteomic Identification and Analysis of Arginine-Methylated Proteins of Plasmodium falciparum at Asexual Blood Stages.

Plasmodium falciparum undergoes a tightly regulated developmental process in human erythrocytes, and recent studies suggest an important regulatory role of post-translational modifications (PTMs). As compared with Plasmodium phosphoproteome, little is known about other PTMs in the parasite. In the present study, we performed a global analysis of asexual blood stages of Plasmodium falciparum to identify arginine-methylated proteins. Using two different methyl arginine-specific antibodies, we immunoprecipitated the arginine-methylated proteins from the stage-specific parasite lysates and identified 843 putative arginine-methylated proteins by LC-MS/MS. Motif analysis of the protein sequences unveiled that the methylation sites are associated with the previously known methylation motifs such as GRx/RGx, RxG, GxxR, or WxxxR. We identified Plasmodium homologues of known arginine-methylated proteins in trypanosomes, yeast, and human. Hydrophilic interaction liquid chromatography (HILIC) was performed on the immunoprecipitates from the trophozoite stage to enrich arginine-methylated peptides. Mass spectrometry analysis of immunoprecipitated and HILIC fractions identified 55 arginine-methylated peptides having 62 methylated arginine sites. Functional classification revealed that the arginine-methylated proteins are involved in RNA metabolism, protein synthesis, intracellular protein trafficking, proteolysis, protein folding, chromatin organization, hemoglobin metabolic process, and several other functions. Summarily, the findings suggest that protein methylation of arginine residues is a widespread phenomenon in Plasmodium, and the PTM may play an important regulatory role in a diverse set of biological pathways, including host-pathogen interactions.

Visualization and quantification of Plasmodium falciparum intraerythrocytic merozoites.

Malaria, a leading parasitic killer, is caused by Plasmodium spp. The pathology of the disease starts when Plasmodium merozoites infect erythrocytes to form rings, that matures through a large trophozoite form and develop into schizonts containing multiple merozoites. The number of intra-erythrocytic merozoites is a key-determining factor for multiplication rate of the parasite. Counting of intraerythrocytic merozoites by classical 2-D microscopy method is error prone due to insufficient representation of merozoite in one optical plane of a schizont. Here, we report an alternative 3-D microscopy based automated method for counting of intraerythrocytic merozoites in entire volume of schizont. This method offers a considerable amount of advantages in terms of both, ease and accuracy.