Biotic elicitors enhance diosgenin production in Helicteres isora L. suspension cultures via up-regulation of CAS and HMGR genes

In an attempt to find an alternative and potent source of diosgenin, a steroidal saponin in great demand for its pharmaceutical importance, Helicteres isora suspension cultures were explored for diosgenin extraction. The effect of biotic elicitors on the biosynthesis of diosgenin, in suspension cultures of H. isora was studied. Bacterial as well as fungal elicitors such as Escherichia coli, Bacillus subtilis, Saccharomyces cerevisiae and Aspergillus niger were applied at varying concentrations to investigate their effects on diosgenin content. The HPLC based quantification of the treated samples proved that amongst the biotic elicitors, E. coli (1.5%) proved best with a 9.1-fold increase in diosgenin content over respective control cultures. Further, the scaling-up of the suspension culture to shake-flask and ultimately to bioreactor level were carried out for production of diosgenin. During all the scaling-up stages, diosgenin yield obtained was in the range between 7.91 and 8.64 mg l−1, where diosgenin content was increased with volume of the medium. The quantitative real-time PCR (qRT-PCR) analysis showed biotic elicitors induced the expression levels of regulatory genes in diosgenin biosynthetic pathway, the 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) and cycloartenol synthase (CAS), which can be positively correlated with elicited diosgenin contents in those cultures. The study holds significance as H. isora represents a cleaner and easy source of diosgenin where unlike other traditional sources, it is not admixed with other steroidal saponins, and the scaled-up levels of diosgenin achieved herein have the potential to be explored commercially.     

Characterizing Mutational Signatures in Human Cancer Cell Lines Reveals Episodic APOBEC Mutagenesis

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A growth factor-dependent nuclear kinase phosphorylates p27(Kip1) and regulates cell cycle progression

The cyclin-dependent kinase inhibitor, p27(Kip1), which regulates cell cycle progression, is controlled by its subcellular localization and subsequent degradation. p27(Kip1) is phosphorylated on serine 10 (S10) and threonine 187 (T187). Although the role of T187 and its phosphorylation by Cdks is well-known, the kinase that phosphorylates S10 and its effect on cell proliferation has not been defined. Here, we identify the kinase responsible for S10 phosphorylation as human kinase interacting stathmin (hKIS) and show that it regulates cell cycle progression. hKIS is a nuclear protein that binds the C-terminal domain of p27(Kip1) and phosphorylates it on S10 in vitro and in vivo, promoting its nuclear export to the cytoplasm. hKIS is activated by mitogens during G(0)/G(1), and expression of hKIS overcomes growth arrest induced by p27(Kip1). Depletion of KIS using small interfering RNA (siRNA) inhibits S10 phosphorylation and enhances growth arrest. p27(-/-) cells treated with KIS siRNA grow and progress to S/G(2 )similar to control treated cells, implicating p27(Kip1) as the critical target for KIS. Through phosphorylation of p27(Kip1) on S10, hKIS regulates cell cycle progression in response to mitogens.     

An immunochemical method for detection and analysis of changes in methylome

DNA methylation is an important epigenetic modification involved in the ability of an organism to respond to stress and adaptation. It has been implicated in development, differentiation, oncogenesis, chromatin remodelling, nutrigenomics, and appears to play a pivotal role in many regulatory and adaptive functions. It is therefore important to analyze the status of DNA methylation and its changes under various developmental, carcinogenic, pharmacological, and environmental conditions. In this report we describe an immunochemical method for the detection of genome wide DNA methylation and its alterations under various conditions along with the analysis of DNA methyltransferase activity. The ability of this approach to detect and provide a map of methylomic changes in a genome facilitates assessment of various agents and conditions which can alter this important epigenetic signal. This experimental system permits rapid evaluation of potential target genes which would be modulated by DNA methylation changes and thus the gene networks that govern the processes.     

Indirect organogenesis and plant regeneration in Helicteres isora L., an important medicinal plant

Abstract    Helicteres isora is a medicinal plant effective against asthma, diabetes, hypolipidemia, HIV, polio besides a good source of diosgenin. Seed dormancy and low natural fruit production rate make this plant a perfect candidate for developing an in vitro regeneration method. However, to date, no such work has been procured in this plant. An efficient method for plant regeneration via shoot organogenesis from callus cultures has been developed using nodal explants in H. isora. Murashige and Skoog (MS) media counting 2,4-Dichlorophenoxyacetic acid (2,4-D, 2.26 to 13.57 μM), Indole-3-acetic acid (IAA, 2.85 to 17.13 μM), Indole-3-butyric acid (IBA, 2.46 to 14.70 μM), 6-Benzylaminopurine (BA, 2.22 to 13.32 μM) and Kinetin (Kin, 2.32 to 13.92 μM) either singly or in the following combinations (IAA + BA; IAA + Kin, and BA + Kin) produced granular callus except BA + Kin which resulted in compact, hard, greenish-white (CHGW) callus. The optimum CHGW callus (2.62 g fresh weight/ explant) was produced on MS media with 13.32 μM BA + 2.32 μM Kin with over 93% callus induction frequency. Optimum shoot organogenesis (67% frequency) was achieved in CHGW callus with lower level of BA (2.22 μM) and Kin (2.32 μM) and produced 3.2 shoots/0.5 g callus within 35 d of culture. Microshoots were rooted successfully (62% frequency) after 35 d of culture on 1/2MS containing 4.90 μM IBA and hardened off. Antioxidant enzymes such as catalase, peroxidase, polyphenol oxidase, and biochemical parameters viz. hydrogen peroxide, reducing and nonreducing sugars, starch, proteins, phenols, and proline contents were studied in regenerating and nonregenerating CHGW calluses to establish a correlation between these parameters and shoot morphogenesis. All the enzyme activities and biochemical parameters were found more in regenerating callus than in nonregenerating except phenols.     

Oculocutaneous Albinism Type 1: The Last 100 Years

Research on human albinism has been central to many of the major discoveries in human genetics. These include the first evidence that Mendel's rules of genetic segregation apply to humans, first published in 1903. Contrary to initial thought that albinism is caused by mutations in a single gene, we now know that the genetics of albinism are complex. The complexity of albinism was hinted at, in early publications, but has only recently been fully appreciated with the advent of molecular techniques. Currently, 12 different genes have been identified, that when mutated, result in a different type of albinism. Oculocutaneous albinism type 1 (OCA1), resulting from mutations of the tyrosinase gene, is genetically and biochemically the best understood type of albinism. Though much of the research in albinism has involved OCA1, there are many unanswered questions about OCA1 and albinism, in general. The next 100 yr should still provide many surprises as did the first 100 yr.     

An exploration of microbial and associated functional diversity in the OMZ and non-OMZ areas in the Bay of Bengal

Depletion of oxygen in certain marine areas creates oxygen minimum zones (OMZs), which can alter the species composition and abundance. We have carried out high-throughput 16S rRNA gene amplicon profiling from the Bay of Bengal (BOB) OMZ and non-OMZ areas. Typically, a total of 35 families of micro-organisms were identified as biomarkers for OMZ and non-OMZ regions in the BOB. Our analysis has identified families Pseudoalteromonadaceae, OM60 and Synechococcaceae to be abundant in oxygenated water, whereas organisms belonging to families Pelagibacteraceae and Caulobacteraceae, which are involved in sulphur and nitrogen metabolism, were prominent in the OMZ areas. Predictive functional analysis for these identified bacteria clearly that suggested an abundance of microbes with assimilatory sulphur-reducing genes (cysl and csH) in the non-OMZ, while bacteria involved in dissimilatory sulphate reduction (known to carry aprA and aprB genes) were enriched in the OMZ areas. Comparative analysis with OMZ areas from Peru and Chile revealed that OMZ areas in the BOB are characterized by specific and distinctive bacterial diversity. Overall, the current analysis provides valuable documentation about the bacterial populations and their characteristics, which can generate pointers for their functional significance in the BOB.