Influence of Exon Duplication on Intron and Exon Phase Distribution

Nonrandomness in the intron and exon phase distributions in a sample of 305 human genes has been found and analyzed. It was shown that exon duplications had a significant effect on the exon phase nonrandomness. All of the nonrandomness is probably due to both the processes of exon duplication and shuffling. A quantitative estimation of exon duplications in the human genome and their influence on the intron and exon phase distributions has been analyzed. According to our estimation, the proportion of duplicated exons in the human genome constitutes at least 6% of the total. Generalizing the particular case of exon duplication to the more common event of exon shuffling, we modeled and analyzed the influence of exon shuffling on intron phase distribution. Received: 28 March 1997 / Accepted: 9 July 1997     

The Ll.LtrB intron from Lactococcus lactis excises as circles in vivo: insights into the group II intron circularization pathway

Group II introns are large ribozymes that require the assistance of intron-encoded or free-standing maturases to splice from their pre-mRNAs in vivo. They mainly splice through the classical branching pathway, being released as RNA lariats. However, group II introns can also splice through secondary pathways like hydrolysis and circularization leading to the release of linear and circular introns, respectively. Here, we assessed in vivo splicing of various constructs of the Ll.LtrB group II intron from the Gram-positive bacterium Lactococcus lactis. The study of excised intron junctions revealed, in addition to branched intron lariats, the presence of perfect end-to-end intron circles and alternatively circularized introns. Removal of the branch point A residue prevented Ll.LtrB excision through the branching pathway but did not hinder intron circle formation. Complete intron RNA circles were found associated with the intron-encoded protein LtrA forming nevertheless inactive RNPs. Traces of double-stranded head-to-tail intron DNA junctions were also detected in L. lactis RNA and nucleic acid extracts. Some intron circles and alternatively circularized introns harbored variable number of non-encoded nucleotides at their splice junction. The presence of mRNA fragments at the splice junction of some intron RNA circles provides insights into the group II intron circularization pathway in bacteria.     

MHC gene configuration variation in geographically disparate populations of California sea lions (Zalophus californianus)

Major histocompatibility complex (MHC) class II DRB genotypes were examined in two geographically isolated populations of California sea lions (Zalophus californianus) (Gulf of California and California coastal Pacific Ocean). Genomic DNA from 227 California sea lions was examined using eight sequence-specific primer (SSP) pairs flanking the putative peptide-binding site. A total of 40 different Zaca-DRB genotype configurations were identified among the 227 individuals. Using SSP-PCR, significant differences were found between coastal California and Gulf of California Zalophus populations in numbers of DRB sequences per individual and configuration of sequences within individuals. Additionally, unique local patterns of MHC diversity were identified among the Midriff Island animals. These population differences are consistent with either ecologically distinct patterns of selection pressures and/or geographical isolation. The consequences of these partitioned MHC configurations at the population level are as yet unknown; however, the worldwide increase in emerging marine diseases lends urgency to their examination.     

High expression of the mRNA of cytochrome P450 and phase II enzymes in the lung and kidney tissues of cattle

The objective of this study was to investigate the tissue-specific mRNA expression of different cytochrome P450 (CYP) isoforms, UDP glucuronsyl transferase 1A1 (UGT1A1) and glutathione-S-transferase (GSTA1) in the different tissues (liver, mammary gland, lungs, spleen, kidney cortex, heart, masseter muscle and tongue) of cattle, using quantitative real-time polymerase chain reaction (qPCR). CYP1A1-like mRNA was expressed in all of the tissues examined, including the liver, with the highest expression level in the kidney. CYP1A2-, 2E1- and 3A4-like mRNAs were only expressed hepatically. Interestingly, significant expression of CYP2B6-like mRNA was recorded in the lung tissue, while CYP2C9-like mRNA was expressed in the liver and kidney tissues of the cattle examined. UGT1A1- and GSTA1-like mRNAs were expressed in all of the examined tissues, except the mammary glands, and the highest expression levels were recorded in the kidney. The high expression of UGT1A1 in the lung tissue and GSTA1 in the liver tissue was unique to cattle; this has not been reported for rats or mice. The findings of this study strongly suggest that the liver, kidneys and lungs of cattle are the major organs contributing to xenobiotics metabolism.     

Enhancement of YD spin relaxation by the CaMn4 cluster in photosystem II detected at room temperature: A new probe for the S-cycle

The long-lived, light-induced radical Y_D^• of the Tyr161 residue in the D2 protein of Photosystem II (PSII) is known to magnetically interact with the CaMn₄ cluster, situated ∼ 30 Å away. In this study we report a transient step-change increase in Y_D^• EPR intensity upon the application of a single laser flash to S₁ state-synchronised PSII-enriched membranes from spinach. This transient effect was observed at room temperature and high applied microwave power (100 mW) in samples containing PpBQ, as well as those containing DCMU. The subsequent decay lifetimes were found to differ depending on the additive used. We propose that this flash-induced signal increase was caused by enhanced spin relaxation of Y_D^• by the OEC in the S₂ state, as a consequence of the single laser flash turnover. The post-flash decay reflected S₂ → S₁ back-turnover, as confirmed by their correlations with independent measurements of S₂ multiline EPR signal and flash-induced variable fluorescence decay kinetics under corresponding experimental conditions. This flash-induced effect opens up the possibility to study the kinetic behaviour of S-state transitions at room temperature using Y_D^• as a probe.     

Synthesis of novel acridine and bis acridine sulfonamides with effective inhibitory activity against the cytosolic carbonic anhydrase isoforms II and VII

4-Amino-N-(4-sulfamoylphenyl)benzamide was synthesized by reduction of 4-nitro-N-(4-sulfamoylphenyl)benzamide and used to synthesize novel acridine sulfonamide compounds, by a coupling reaction with cyclic-1,3-diketones and aromatic aldehydes. The new compounds were investigated as inhibitors of the metalloenzyme carbonic anhydrase (CA, EC 4.2.1.1), and more precisely the cytosolic isoforms hCA I, II and VII. hCA I was inhibited in the micromolar range by the new compounds (K_Is of 0.16–9.64 μM) whereas hCA II and VII showed higher affinity for these compounds, with K_Is in the range of 15–96 nM for hCA II, and of 4–498 nM for hCA VII. The structure–activity relationships for the inhibition of these isoforms with the acridine–sulfonamides reported here were also elucidated.     

Drosophila p24 homologues eclair and baiser are necessary for the activity of the maternally expressed Tkv receptor during early embryogenesis

p24 proteins are assumed to play an important role in the transport of secreted and transmembrane proteins into membranes. However, only few cargo proteins are known that partially, but in no case completely require p24 proteins for membrane transport. Here, we show that two p24 proteins are essential for dorsoventral patterning of Drosophila melanogaster embryo. Mutations in the genes, eclair (eca) and baiser (bai), encoding two p24 proteins reduce signalling by the TGF-beta homologue, Dpp, in early embryos. This effect is strictly maternal and specific to early embryogenesis, as Dpp signalling in other contexts is not notably affected. We provide genetic evidence that in the absence of eca or bai function in the oocyte, the maternally expressed type I TGF-beta receptor Tkv is not active. We propose that during early embryogenesis eca and bai are specifically required for the activity of the maternal Tkv, while the zygotic Tkv is not affected in the mutant embryos. Mutations in either eca or bai are sufficient for the depletion of Tkv activity and no enhancement of the phenotypes was observed in embryos derived from oocytes mutant for both genes. The dependence of maternal Tkv protein on the products of p24 genes may serve as an in vivo model for studying p24 proteins.     

Oil/water and pre-emulsified oil/water (PIT) dispersions in a stirred vessel: Implications for fermentations

This study examines dispersions of rapeseed oil (RSO) in water by mechanical agitation under conditions mimicking those found in certain antibiotic fermentations; for example, in the presence of air, antifoam, and finely divided CaCO(3) particles. A problem with residual oil has been reported for such fermentations, and it has been suggested that the use of pre-emulsified oil can reduce this problem. Hence, the dispersion of a pre-emulsified oil produced by the "phase inversion temperature (PIT) method" has been evaluated. In both cases, the volume fraction of oil was 2%. For the RSO systems, a relatively high agitation speed was required to disperse the oil, especially in the presence of the particles and, when the agitation was stopped, separation occurred rapidly. The Sauter mean drop diameters depended on the system, being at an average energy dissipation rate of approximately 0.9 W kg(-1), 180 microm for RSO/water, 130 microm for RSO/water(antifoam)/air, 580 microm for RSO/water/CaCO(3), and 850 microm for RSO/water(antifoam)/air/CaCO(3). For the same four systems, the PIT emulsion, once dispersed, was very stable and the drop size was essentially independent of the operating conditions, with a Sauter mean diameter of approximately 0.3 microm. The implications of these findings for fermentations in which oil is used as a carbon source are assessed.