Nucleotide analogues to investigate RNA structure and function

RNA plays an essential cellular role in nearly every aspect of the transmission and expression of genetic information, including regulatory roles that have significance for cellular development. Access to RNA bearing synthetic modifications has allowed biological chemists to probe deep into the inner workings of cellular processes. Here, we describe recent advances in harnessing the power of nucleotide analogues to obtain mechanistic and biological insights into RNA structure, function and dynamics.     

Myotrophin-kappaB DNA interaction in the initiation process of cardiac hypertrophy

To investigate how cardiac hypertrophy and heart failure develop, we isolated and characterized a candidate initiator, the soluble 12-kDa protein myotrophin, from rat and human hearts. Myotrophin stimulates protein synthesis and myocardial cell growth associated with increased levels of hypertrophy marker genes. Recombinant myotrophin from the cloned gene showed structural/functional motifs, including ankyrin repeats and putative phosphorylation sites for protein kinase C (PKC) and casein kinase II. One repeat, homologous with I kappaB, interacts with rel/NF-kappaB in vitro. We analyzed the interaction of recombinant myotrophin and nuclear extracts prepared from neonatal and adult cardiomyocytes; gel mobility shift assay showed that myotrophin bound to kappaB DNA. To define PKC's role in myotrophin-induced myocyte growth, we incubated neonatal rat myocytes (normal and stretch) with specific inhibitors and found that myotrophin inhibits [3H]leucine incorporation into myocytes and different hypertrophic gene expression in neonatal myocytes. Using confocal microscopy, we observed that a basal level of myotrophin was present in both cytoplasm and nucleus under normal conditions, but under cyclic stretch, myotrophin levels became elevated in the nucleus. Myotrophin gene levels were upregulated when myocytes underwent cyclic stretch or were treated with tumor necrosis factor-alpha (TNF-alpha) or interleukin-1beta and also when excised beating hearts were exposed to high pressure. Our data showed that the myotrophin-kappaB interaction was increased with age in spontaneously hypertensive rats (SHRs) only. Our data provide evidence that myotrophin-kappaB DNA interaction may be an important step in initiating cardiac hypertrophy.     

Evolution of the chordate muscle actin gene

Summary The ascidians Styela plicata, S. clava, and Mogula citrina are urochordates. The larvae of urochordates are considered to morphologically resemble the ancestral vertebrate. We asked whether larval and adult ascidian muscle actin sequences are nonmusclelike as in lower invertebrates, musclelike as in vertebrates, or possess characteristics of both. Nonmuscle and muscle actin cDNA clones from S. plicata were sequenced. Based on 27 diagnostic amino acids, which distinguish vertebrate muscle actin from other actins, we found that the deduced protein sequences of ascidian muscle actins exhibit similarities to both invertebrate and vertebrate muscle actins. A comparison to muscle actins from different vertebrate and invertebrate phylogenetic groups suggested that the urochordate muscle actins represent a transition from a nonmusclelike sequence to a vertebrate musclelike sequence. The ascidian adult muscle actin is more similar to skeletal actin and the larval muscle actin is more similar to cardiac actin, which indicates that the divergence of the skeletal and cardiac isoforms occurred before the emergence of urochordates. The muscle actin gene may be a powerful probe for investigating the chordate lineage. Offprint requests to: C.R. Tomlinson     

Capturing the Mechanism Underlying TOP mRNA Binding to LARP1

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Column Test in Field Application Study Using Biostimulant Ball in Contaminated Coastal Sediment

This study consisted of a field column test to evaluate the effectiveness of the slow release and long-term outcomes of a biostimulant ball (BSB) in contaminated coastal sediment. The BSB was prepared using biostimulating agents consisting of sulphate, nitrate and acetate with uncontaminated dredged sediment, and cellulose acetate (CA) and polysulfone (PS). SEM images were captured to investigate the physical properties of the PS and CA-coated biostimulant ball, and doing so was observed to have a great influence in the release rate of the nutrients. The CA-coated BSB had large pores with an irregular shape inside and a honeycomb structure outside. The PS-coated BSB had no pores inside and had small pores outside the coating, and the release rate of the PS-coated BSB was slower with more long-term effects than the CA-coated BSB. The CA- and PS-coated BSB sample showed that the chemical oxygen demand (COD), total phosphorous (TP) and total nitrogen (TN) had been effectively reduced, and conversely, pH is not significantly affected when using the BSB. Moreover, the fractions of heavy metals (Cd, Zn, Cu, Pb, Cr and Fe) were highly influenced when using the PS-coated BSB added sample than when using the CA-coated BSB. Therefore, application of CA- and PS-coated BSB is a promising method for cleaning contaminated coastal sediment.     

Plant Growth-Promoting Chitinolytic Paenibacillus elgii Responds Positively to Tobacco Root Exudates

Bacterial strains from chitin/chitosan-rich soils, from two industries, were screened for their chitinolytic, antifungal, and mineral phosphate solubilization abilities. The isolate SMA-1-SDCH02, positive for all three properties, was selected and identified as Paenibacillus elgii based on morphological and biochemical characters and supported by 16S rRNA gene sequence analysis. P. elgii enhanced the growth of groundnut in terms of shoot height, root length, total chlorophyll, and fresh and dry weight when applied alone or in combination with chitosan. The plant growth-promoting activity of P. elgii was seen in tobacco in a specially designed gnotobiotic setup indicating its capability to promote growth of at least groundnut and tobacco. Metabolite changes in the bacteria, studied using attenuated total reflectance-infrared (ATR-IR) spectroscopy, revealed split bands of amide I at the 1659- and 1636-cm⁻¹ regions when grown in minimal media amended with tobacco root exudates. The difference in ATR-IR bands in the presence of tobacco root exudates indicated production of compounds with differences in functional groups.     

The 5′-Nor Aristeromycin Analogues of 5′-Deoxy-5′-Methylthioadenosine and 5′-Deoxy-5′-Thiophenyladenosine

To extend the potential of 5′-noraristeromycin (and its enantiomer) as potential antiviral candidates, the enantiomers of the carbocyclic 5′-nor derivatives of 5′-methylthio-5′-deoxyadenosine and 5′-phenylthio-5′-deoxyadenosine have been synthesized and evaluated. None of the compounds showed meaningful antiviral activity.