Assessment of Biological Activities of Fungal Endophytes Derived Bioactive Compounds Isolated from Amoora rohituka.

Fungal endophytes have remarkable potential to produce bioactive compounds with numerous pharmacological significance that are used in various disease management and human welfare. In the current study, a total of eight fungal endophytes were isolated from the leaf tissue of Amoora rohituka, and out of which ethyl acetate (EA) extract of Penicillium oxalicum was found to exhibit potential antioxidant activity against DPPH, nitric oxide, superoxide anion and hydroxyl free radicals with EC50 values of 178.30 ± 1.446, 75.79 ± 0.692, 169.28 ± 0.402 and 126.12 ± 0.636 µg/mL, respectively. The significant antioxidant activity of EA extract of P. oxalicum is validated through highest phenolic and flavonoid content, and the presence of unique bioactive components observed through high-performance thin layer chromatography (HPTLC) fingerprinting. Moreover, EA extract of P. oxalicum also displayed substantial anti-proliferative activity with IC50 values of 56.81 ± 0.617, 37.24 ± 1.26 and 260.627 ± 5.415 µg/mL against three cancer cells HuT-78, MDA-MB-231 and MCF-7, respectively. Furthermore, comparative HPTLC fingerprint analysis and antioxidant activity of P. oxalicum revealed that fungal endophyte P. oxalicum produces bioactive compounds in a host-dependent manner. Therefore, the present study signifies that fungal endophyte P. oxalicum associated with the leaf of A. rohituka could be a potential source of bioactive compounds with antioxidant and anticancer activity.

A rapid, efficient, and low-cost BiFC protocol and its application in studying in vivo interaction of seed-specific transcription factors, RISBZ and RPBF.

Proteins regulate cellular and biological processes in all living organisms. More than 80% of the proteins interact with one another to perform their respective functions; therefore, studying the protein-protein-interaction has gained attention in functional characterization studies. Bimolecular fluorescence complement (BiFC) assay is widely adopted to determine the physical interaction of two proteins in vivo. Here, we developed a simple, yet effective BiFC assay for protein-protein-interaction using transient Agrobacterium-mediated-transformation of onion epidermal cells by taking case study of Rice-P-box-Binding-Factor (RPBF) and rice-seed-specific-bZIP (RISBZ) in vivo interaction. Our result revealed that both the proteins, i.e., RISBZ and RPBF, interacted in the nucleus and cytosol. These two transcription factors are known for their coordinate/synergistic regulation of seed-protein content via concurrent binding to the promoter region of the seed storage protein (SSP) encoding genes. We further validated our results with BiFC assay in Nicotiana by agroinfiltration method, which exhibited similar results as Agrobacterium-mediated-transformation of onion epidermal cells. We also examined the subcellular localization of RISBZ and RPBF to assess the efficacy of the protocol. The subcellular localization and BiFC assay presented here is quite easy-to-follow, reliable, and reproducible, which can be completed within 2-3 days without using costly instruments and technologies that demand a high skill set.