Isolation and characterization of some novel genes of the apolipoprotein A-I family in Japanese eel, <Emphasis Type="Italic">Anguilla japonica</Emphasis>

Apolipoproteins such as apolipoprotein (apo) A-I, apoA-IV, and apoE are lipid binding proteins synthesized mainly in the liver and the intestine and play an important role in the transfer of exogenous or endogenous lipids through the circulatory system. To investigate the mechanism of lipid transport in fish, we have isolated some novel genes of the apoA-I family, apoIA-I (apoA-I isoform) 1–11, from Japanese eel by PCR amplification. Some of the isolated genes of apoIA-I corresponded to 28kDa-1 cDNAs which had already been deposited into the database and encoded an apolipoprotein with molecular weight of 28 kDa in the LDL, whereas others seemed to be novel genes. The structural organization of all apoIA-Is consisted of four exons separated by three introns. ApoIA-I10 had a total length of 3232 bp, whereas other genes except for apoIA-I9 ranged from 1280 to 1441 bp. The sequences of apoIA-Is at the exon-intron junctions were mostly consistent with the consensus sequence (GT/AG) at exon-intron boundaries, whereas the sequences of 3′ splice acceptor in intron 1 of apoIA-I1-7 were (AC) but not (AG). The deduced amino acid sequences of all apoIA-Is contained a putative signal peptide and a propeptide of 17 and 5 amino acid residues, respectively. The mature proteins of apoIA-I1-3, 7, and 8 consisted of 237 amino acids, whereas those of apoIA-I4-6 consisted of 239 amino acids. The mature apoIA-I10 sequence showed 65% identity to amino acid sequence of apoIA-I11 which was associated with an apolipoprotein with molecular weight of 23 kDa in the VLDL. All these mature apoIA-I sequences satisfied the common structural features depicted for the exchangeable apolipoproteins such as apoA-I, apoA-IV, and apoE but apoIA-I11 lacked internal repeats 7, 8, and 9 when compared with other members of apoA-I family. Phylogenetic analysis showed that these novel apoIA-Is isolated from Japanese eel were much closer to apoA-I than apoA-IV and apoE, suggesting new members of the apoA-I family.     

Successive Isolation of Proteinase Hyperproductive Mutants of Aspergillus sojae

Hydrochloric acid treatment of methyl 3-(4-isobutylphenyl)-3-methylglycidate and methyl 2-hydroxy-3-(4-isobutylphenyl)-3-butenoate, a rearrangement product of the former, in acetic acid gave 3-(4-isobutylphenyl)-3-methylpyruvic acid and 2-(4-isobutylphenyl)-pro-panal. The same treatment of 2-hydroxy-3-(4-isobutylphenyl)-3-butenoic acid gave 2-(4-isobutylphenyl)-propanal. Both 3-(4-isobutylphenyl)-3-methylpyruvic acid and 2-(4-iso-butylphenyl)-propanal were oxidized to 2-(4-isobutylphenyl)-propionic acid.     

The RNA ligands for mouse proline-rich RNA-binding protein (mouse Prrp) contain two consensus sequences in separate loop structure

Mouse proline-rich RNA-binding protein (mPrrp) is a mouse ortholog of Xenopus Prrp, which binds to a vegetal localization element (VLE) in the 3′-untranslated region (3′-UTR) of Vg1 mRNA and is expected to be involved in the transport and/or localization of Vg1 mRNA to the vegetal cortex of oocytes. In mouse testis, mPrrp protein is abundantly expressed in the nuclei of pachytene spermatocytes and round spermatids, and shifts to the cytoplasm in elongating spermatids. To gain an insight into the function of mPrrp in male germ cells, we performed in vitro RNA selection (SELEX) to determine the RNA ligand sequence of mPrrp. This analysis revealed that many of the selected clones contained both of two conserved elements, AAAUAG and GU_1–3AG. RNA-binding study on deletion mutants and secondary structure analyses of the selected RNA revealed that a two-loop structure containing the conserved elements is required for high-affinity binding to mPrrp. Furthermore, we found that the target mRNAs of Xenopus Prrp contain intact AAAUAG and GU_1–3AG sequences in the 3′-UTR, suggesting that these binding sequences are shared by Prrps of Xenopus and mouse.     

Brap2 functions as a cytoplasmic retention protein for p21 during monocyte differentiation

The cell cycle inhibitor p21 plays an important role in monocytic cell differentiation, during which it translocates from the nucleus to cytoplasm. This process involves the negative regulation of the p21 nuclear localization signal (NLS). Here, we sought to determine the relationship between the cytoplasmic translocation of p21 and another molecule, Brap2, a cytoplasmic protein which binds the NLS of BRCA1 and was recently reported to inactivate KSR in the Ras-activating signal pathway under the name of IMP. We report that p21 and Brap2 directly interact, both in vitro and in vivo, in a manner requiring the NLS of p21 and the C-terminal portion of Brap2. When it is cotransfected with Brap2, p21 is expressed in the cytoplasm. Monocytic differentiation of the promyelomonocytic cell lines U937 and HL60 is associated with the upregulation of Brap2 expression concomitantly with the upregulation and cytoplasmic relocalization of p21. Our results underscore the role played by Brap2 in the process of cytoplasmic translocation of p21 during monocyte differentiation.     

Flavone-catalyzed apoptosis in Scutellaria baicalensis

In response to mechanical damage, roots of Scutellaria baicalensis undergo cell death within 24 h. The flavone baicalein was identified as the factor regulating apoptosis in the damaged roots of S. baicalensis. Plant apoptosis is known to be triggered by oxidative damage of DNA through oxidative bursts, whereas baicalein causes apoptosis in Scutellaria cells by a copper-dependent oxidation of nuclear DNA without inducing an oxidative burst. S. baicalensis possesses an interesting system for quickly producing this apoptosis-inducing flavone in its cells. Intact Scutellaria cells contain little baicalein but store a large amount of baicalin (baicalein 7-O-β-D-glucuronide). Stress treatment of Scutellaria cells immediately initiates hydrolysis of baicalin by endogenous β-glucuronidase, and the resulting baicalein is immediately translocated to the nucleus, leading to apoptosis. Thus, S. baicalensis possesses a unique apoptosis-inducing system that is linked with metabolism of baicalin.     

Non-systemic expression of a stress-responsive maize polyubiquitin gene (Ubi-1) in transgenic rice plants

We have used the promoter, 1st exon and 1st intron of the maize polyubiquitin gene (Ubi-1) for rice transformation experiments and revealed the characteristic expression of Ubi-1 gene: (1) Ubi-1 gene is not regulated systemically but rather individual cells respond independently to the heat or physical stress; (2) Ubi-1 gene changes its tissue-specific expression in response to stress treatment; (3) the expression of Ubi-1 gene is dependent on cell cycle.     

Synthesis of a polymide containing a glucose unit in the main chain

A new type polyamide containing a glucose unit in the main chain has been synthesized by the polymerization of C₁, C₃, C₄ blocked C₆-carboxymethylglucosamine, prepared from chitin. The deblocking procedure gave the water-soluble polyamide, of MW 1.5 × 10⁴, which can be regarded as a model for the recognition site of lectin.     

Alicyclic acid metabolism in plants 8. Association, of two enzymes of the shikimate pathway in shoots of Phaseolus mungo seedlings

The association of two enzymes involved in the shikimate pathway,3-dehydroquinate hydro-lyase (EC 4.2.1.10 [EC] ) and shikimate: NADPoxidoreductase (EC 1.1.1.25 [EC] ), was studied with shoots of etiolated4-day-old Phaseolus mungo seedlings. The enzymes were not separableby ammonium sulfate fractionation, sucrose density gradientcentrifugation, polyacrylamide gel electrophoresis and chromatographyon Sephadex G-100 and DEAE-Sephadex A-50. The results are discussedin relation to the channelling function of metabolites in thealicyclic acid metabolism in higher plants. (Received October 28, 1975; )