In vitro organogenesis and plant formation in Acacia sinuata

In vitro morphogenesis via organogenesis was achieved from callus cultures derived from hypocotyl explants of Acacia sinuata on MS (Murashige and Skoog, 1962) medium. Calli were induced from hypocotyl explants excised from 7-day-old seedlings on MS medium containing 3% sucrose, 0.8% agar, 6.78 μM 2,4-dichlorophenoxyacetic acid and 2.22 μM 6-benzylaminopurine. Regeneration of adventitious buds from callus was achieved when they were cultured on MS medium supplemented with 10% coconut water, 13.2 μM 6-benzylaminopurine and 3.42 μM indoleacetic acid. Addition of gibberellic acid (1.73 μM) favored shoot elongation. Regenerated shoots produced prominent roots when transferred to half strength MS medium supplemented with 7.36 μM indolebutyric acid. Rooted plantlets, thus developed were hardened and successfully established in the soil. This protocol yielded an average of 20 plants per hypocotyl explant over a period of 4 months. This revised version was published online in June 2006 with corrections to the Cover Date.     

Indole-3-piperazinyl derivatives: novel chemical class of 5-HT(6) receptor antagonists

N₁-Arylsulfonyl-3-piperazinyl indole derivatives were designed and identified as a novel class of 5-HT₆ receptors ligands. All the compounds have high affinity and antagonist activity towards 5-HT₆ receptor. The compound 7a (K_i = 3.4 nM, functional assay IC₅₀ = 310 nM) shows enhanced cognitive effect when tested in NORT and Morris water maze models. Synthesis, SAR and PK profile of these novel compounds constitute the subject matter of this Letter.     

Deciphering the ovarian cancer ascites fluid peptidome

Background

Conventional proteomic approaches have thus far been unable to identify novel serum biomarkers for ovarian cancer that are more sensitive and specific than the current clinically used marker, CA-125. Because endogenous peptides are smaller and may enter the circulation more easily than proteins, a focus on the low-molecular-weight region may reveal novel biomarkers with enhanced sensitivity and specificity. In this study, we deciphered the peptidome of ascites fluid from 3 ovarian cancer patients and 3 benign individuals (ascites fluid from patients with liver cirrhosis).

Results

Following ultrafiltration of the ascites fluids to remove larger proteins, each filtrate was subjected to solid phase extraction and fractionated using strong cation exchange chromatography. The resultant fractions were analyzed using an Orbitrap mass spectrometer. We identified over 2000 unique endogenous peptides derived from 259 proteins. We then catalogued over 777 peptides that were found only in ovarian cancer ascites. Our list of peptides found in ovarian cancer specimens includes fragments derived from the proteins vitronectin, transketolase and haptoglobin.

Conclusions

Peptidomics may uncover previously undiscovered disease-specific endogenous peptides that warrant further investigation as biomarkers for ovarian cancer.     

Clinical potential of human-induced pluripotent stem cells

The recent establishment of induced pluripotent stem (iPS) cells promises the development of autologous cell therapies for degenerative diseases, without the ethical concerns associated with human embryonic stem (ES) cells. Initially, iPS cells were generated by retroviral transduction of somatic cells with core reprogramming genes. To avoid potential genotoxic effects associated with retroviral transfection, more recently, alternative non-viral gene transfer approaches were developed. Before a potential clinical application of iPS cell-derived therapies can be planned, it must be ensured that the reprogramming to pluripotency is not associated with genome mutagenesis or epigenetic aberrations. This may include direct effects of the reprogramming method or “off-target” effects associated with the reprogramming or the culture conditions. Thus, a rigorous safety testing of iPS or iPS-derived cells is imperative, including long-term studies in model animals. This will include not only rodents but also larger mammalian model species to allow for assessing long-term stability of the transplanted cells, functional integration into the host tissue, and freedom from undifferentiated iPS cells. Determination of the necessary cell dose is also critical; it is assumed that a minimum of 1 billion transplantable cells is required to achieve a therapeutic effect. This will request medium to long-term in vitro cultivation and dozens of cell divisions, bearing the risk of accumulating replication errors. Here, we review the clinical potential of human iPS cells and evaluate which are the most suitable approaches to overcome or minimize risks associated with the application of iPS cell-derived cell therapies.     

Comparative Genomics and Synteny Analysis of <Emphasis Type="BoldItalic">KCS17</Emphasis>-<Emphasis Type="BoldItalic">KCS18</Emphasis> Cluster Across Different Genomes and Sub-genomes of Brassicaceae for Analysis of Its Evolutionary History

Comparative genomics-based synteny analysis has proved to be an effective strategy to understand evolution of genomic regions spanning a single gene (micro-unit) to large segments encompassing hundreds of kilobases to megabases. Brassicaceae is in a unique position to contribute to understanding genome and trait evolution through comparative genomics because whole genome sequences from as many as nine species have been completed and are available for analysis. In the present work, we compared genomic loci surrounding the KCS17-KCS18 cluster across these nine genomes. KCS18 or FAE 1 gene encodes beta-ketoacyl synthase, (β-KCS) a membrane-bound enzyme that catalyses the key rate-limiting step during synthesis of VLCFAs such as erucic acid (C22) present in seed oil in Brassicaceae by elongating carbon chain from C18 to C22; knowledge on role of KCS17 in plant development is however lacking. Synteny across the genomic segments harbouring FAE1 showed variable levels of gene retention ranging between 26% (Arabidopsis thaliana and Brassica napus C03) and 89% (between A. thaliana and Brassica rapa A01), and gene density ranged between 1 gene/2.86 kb and 1 gene/4.88 kb. Interestingly, in diploid Brassica species, FAE1 was retained in only one of the sub-genomes in spite of the presence of three sub-genomes created as a result of genome triplication; in contrast, FAE1 was present at three loci, with four copies in Camellina sativa which is also known to have experienced a recent genome triplication revealing contrasting fates upon duplication. The organization of KCS17 and KCS18 as head-to-tail cluster was conserved across most of the species, except the C genome containing Brassicas, namely B. oleracea and B. napus, where disruptions because of other genes were observed. Even in the conserved blocks, the distance between KCS17 and KCS18 varied; the functional implication of the organization of KCS17-KCS18 as a cluster vis-à-vis fatty acid biosynthesis needs to be dissected, as the cis-regulatory region is expected to be present in the intergenic space. Phylogenetic analysis of KCS gene family along with KCS17-KCS18 from members of Brassicaceae reveals their ancestral relationship with KCS8-KCS9 block. Further comparative functional analysis between KCS8, KCS9, KCS16, KCS17 and KCS18 across evolutionary time-scale will be required to understand the conservation or diversification of roles of these members of KCS family in fatty acid biosynthesis during course of evolution.     

Functional analysis of calcium-binding EF-hand motifs of visinin-like protein-1

Visinin-like protein-1 (VILIP-1), a myristoylated calcium sensor protein with three EF-hand motifs, modulates adenylyl cyclase activity. It translocates to membranes when a postulated "calcium-myristoyl switch" is triggered by calcium-binding to expose its sequestered myristoyl moiety. We investigated the contributions of the EF-hand motifs to the translocation of VILIP-1 to membranes and to the modulation of adenylyl cyclase activity. Mutation of residues crucial for binding calcium within each one of the EF-hand motifs indicated that they all contributed to binding calcium. Simultaneous mutations of all of the three EF-hand motifs completely abolished VILIP-1's ability to bind calcium, attenuated but did not eliminate its modulation of adenylyl cyclase activity, and abolished its calcium-dependence for association with cellular membranes. These results show that the calcium-binding EF-hand motifs of VILIP-1 do not have an essential role in modulating adenylyl cyclase activity but instead have a structural role in activating the "calcium-myristoyl switch" of VILIP-1.     

Intra- and interspecific interactions between Lysiphlebus testaceipes and Lipolexis scutellaris (Hymenoptera: Aphidiidae) reared on Toxoptera citricida (Homoptera: Aphididae)

Classical biological control of the brown citrus aphid Toxoptera citricida Kirkaldy in Florida has involved the release of Lipolexis scutellaris Mackauer. Lysiphlebus testaceipes Cresson is already present in Florida and also parasitizes T. citricida. Because parasitoid-parasitoid interactions may affect the establishment of a newly introduced parasitoid species, intra- and interspecific larval interactions of both parasitoids were studied in the laboratory using T. citricida reared on citrus. Five time intervals were allowed between successive oviposition opportunities. Early developmental times were determined for both parasitoids: eggs of L. testaceipes and L. scutellaris hatched after 54.3 and 61.7 h, while molt to second instar occurred after 73.3 and 87.1 h, respectively, after oviposition at 22 degrees C. At intervals <12 h, both parasitoids had a greater tendency to multiparasitize than to superparasitize, and the tendency to superparasitize or multiparasitize decreased with an increase in time between successive oviposition opportunities. Of the 10 interspecific interactions studied, 5 produced a winning wasp species, 3 of which could be explained by the hypothesis of physical conflict. A combination of development time, age of competing larvae, and oviposition sequence were responsible for the outcomes observed. Neither parasitoid proved to be intrinsically superior when interspecific competition occurs in second- and third-instar T. citricida, indicating there is no reason to suggest that they cannot coexist in Florida citrus groves.     

Genetic association analysis of KCNQ3 and juvenile myoclonic epilepsy in a South Indian population

Juvenile myoclonic epilepsy (JME) is a common subtype of idiopathic generalized epilepsy that shows a complex pattern of inheritance. We have tested the association between JME phenotype and an intragenic marker in KCNQ3 by using the transmission disequilibrium test in 119 probands and their parents. Mutations in KCNQ3 are known to cause benign familial neonatal convulsions and are involved in the physiologically important M current in neurons. Our results provide suggestive evidence of allelic association between JME and KCNQ3 (P-value=0.008) and raise an interesting possibility of a genetic contribution to JME, viz., of a gene that causes a monogenic form of human epilepsy.