Esterases as a marker for growth of BY-2 tobacco cells and early somatic embryos of the Norway spruce

Growth is one of the basic properties of biological systems. The methods which are commonly used for the determination of growth are usually difficult and not very accurate. In the present work we decided to use esterase activity as a growth marker in tobacco suspension culture (BY-2 line) and in early somatic embryos of Norway spruce (clone 2/32) grown on a semi-solid medium. Esterase activity correlates well with the classical growth characteristics of BY-2 and spruce early somatic embryos. Determination of esterase activity is based on spectrophotometric and spectrofluorimetric detection of reaction products, which arise from the enzymatic hydrolysis of two substrates (p -nitrophenyl acetate and fluorescein diacetate) by esterase. The spectrophotometric method enabled us to detect approximately 10⁴ BY-2 cells and 25 spruce embryos whereas the more sensitive spectrofluorimetric method allowed us to detect approximately 800 BY-2 cells and 5 early somatic embryos of Norway spruce.     

Small molecule biaryl FSH receptor agonists. Part 2: Lead optimization via parallel synthesis

Potent small molecule biaryl diketopiperazine FSH receptor agonists such as 10c (EC(50)=13 nM) and 11f (EC(50)=1.2 nM) were discovered through the design, synthesis and evaluation of three biaryl diketopiperazine optimization libraries with over 300 compounds. These libraries were prepared via solid-phase parallel synthesis using a cyclization-release method.     

Glycosphingolipids in endocytic membrane transport

Endocytosis leads to the internalisation of both lipids and proteins and their delivery to specific subcellular locations. This involves sorting processes that are not completely understood, but may involve interactions between lipids and proteins as well as pH and calcium gradients. This article discusses the importance of endocytosis in glycosphingolipid (GSL) synthesis as well as the potential roles of GSLs in endocytic membrane transport. Although the accumulation of GSLs in storage diseases clearly disrupts endocytic transport, increasing evidence also supports a role for GSLs in endocytosis in normal cells.     

Identification of promoters bound by c-Jun/ATF2 during rapid large-scale gene activation following genotoxic stress

The NH2-terminal Jun kinases (JNKs) function in diverse roles through phosphorylation and activation of AP-1 components including ATF2 and c-Jun. However, the genes that mediate these processes are poorly understood. A model phenotype characterized by rapid activation of Jun kinase and enhanced DNA repair following cisplatin treatment was examined using chromatin immunoprecipitation with antibodies against ATF2 and c-Jun or their phosphorylated forms and hybridization to promoter arrays. Following genotoxic stress, we identified 269 genes whose promoters are bound upon phosphorylation of ATF2 and c-Jun. Binding did not occur following treatment with transplatin or the JNK inhibitor SP600125 or JNK-specific siRNA. Of 89 known DNA repair genes represented on the array, 23 are specifically activated by cisplatin treatment within 3-6 hr. Thus, the genotoxic stress response occurs at least partly via activation of ATF2 and c-Jun, leading to large-scale coordinate gene expression dominated by genes of DNA repair.     

Cadmium-induced changes in antioxidative systems,hydrogen peroxide content,and lipid peroxidation in<Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

To study the relationship between cadmium (Cd)-induced phytotoxicity and oxidative stress, we grew Cd-sensitive wild-type (WT) and Cd-resistant type (RT) seedlings ofArabidopsis thaliana on MS media containing up to 500 μM CdCl₂. The resistant seedlings showed higher biomasses and lower hydrogen peroxide and lipid peroxidation levels, the latter expressed in terms of malondialdehyde (MDA) production. These results indicate that RT plants experience lower oxidative stress when exposed to Cd. Furthermore, compared with the WT, RT seedlings have significantly higher activities of superoxide dismutase (SOD) and enzymes related to hydrogen peroxide removal, e.g., guaiacol peroxidase (GPX), ascorbate peroxidase (APX), and glutathione reductase (GR). These differential responses suggest that such phytotoxicity could be induced by oxidative stress, and that lower accumulations of hydrogen peroxide confer Cd tolerance in seedlings.     

Cloning, characterization and expression of CDK5RAP1_v3 and CDK5RAP1_v4, two novel splice variants of human CDK5RAP1

Two novel splice variants of CDK5RAP1, named CDK5RAP1_v3 and CDK5RAP1_v4, were isolated through the large-scale sequencing analysis of a human fetal brain cDNA library. The CDK5RAP1_v3 and CDK5RAP1_v4 cDNAs are 1923bp and 1792bp in length, respectively. Sequence analysis revealed that CDK5RAP1_v4 lacked 1 exon, which was present in CDK5RAP1_v3, with the result that these cDNAs encoded different putative proteins. The deduced proteins were 574 amino acids (designated as CDK5RAP1_v3) and 426 amino acids (CDK5RAP1_v4) in length, and shared the 420 N-terminal amino acids. RT-PCR analysis showed that human CDK5RAP1_v3 was widely expressed in human tissues. The expression level of CDK5RAP1_v3 was relatively high in placenta and lung, whereas low levels of expression were detected in heart, brain, liver, skeletal muscle, pancreas, spleen, thymus, small intestine and peripheral blood leukocytes. In contrast, human CDK5RAP1_v4 was mainly expressed in brain, placenta and testis.     

A graph theoretical approach for predicting common RNA secondary structure motifs including pseudoknots in unaligned sequences

MOTIVATION: RNA structure motifs contained in mRNAs have been found to play important roles in regulating gene expression. However, identification of novel RNA regulatory motifs using computational methods has not been widely explored. Effective tools for predicting novel RNA regulatory motifs based on genomic sequences are needed. RESULTS: We present a new method for predicting common RNA secondary structure motifs in a set of functionally or evolutionarily related RNA sequences. This method is based on comparison of stems (palindromic helices) between sequences and is implemented by applying graph-theoretical approaches. It first finds all possible stable stems in each sequence and compares stems pairwise between sequences by some defined features to find stems conserved across any two sequences. Then by applying a maximum clique finding algorithm, it finds all significant stems conserved across at least k sequences. Finally, it assembles in topological order all possible compatible conserved stems shared by at least k sequences and reports a number of the best assembled stem sets as the best candidate common structure motifs. This method does not require prior structural alignment of the sequences and is able to detect pseudoknot structures. We have tested this approach on some RNA sequences with known secondary structures, in which it is capable of detecting the real structures completely or partially correctly and outperforms other existing programs for similar purposes. AVAILABILITY: The algorithm has been implemented in C++ in a program called comRNA, which is available at http://ural.wustl.edu/softwares.html