Production of Dwarf Lettuce by Overexpressing a Pumpkin Gibberellin 20-Oxidase Gene

We investigated the effect of overexpressing a pumpkin gibberellin (GA) 20-oxidase gene encoding an enzyme that forms predominantly biologically inactive products on GA biosynthesis and plant morphology in transgenic lettuce (Lactuca sativa cv Vanguard) plants. Lettuce was transformed with the pumpkin GA 20-oxidase gene downstream of a strong constitutive promoter cassette (El2-35S-Omega). The transgenic plants in which the pumpkin gene was detected by polymerase chain reaction were dwarfed in the T(2) generation, whereas transformants with a normal growth phenotype did not contain the transgene. The result of Southern-blot analysis showed that the transgene was integrated as a single copy; the plants segregated three dwarfs to one normal in the T(2) generation, indicating that the transgene was stable and dominant. The endogenous levels of GA(1) and GA(4) were reduced in the dwarfs, whereas large amounts of GA(17) and GA(25), which are inactive products of the pumpkin GA 20-oxidase, accumulated in these lines. These results indicate that a functional pumpkin GA 20-oxidase is expressed in the transgenic lettuce, resulting in a diversion of the normal pathway of GA biosynthesis to inactive products. Furthermore, this technique may be useful for controlling plant stature in other agricultural and horticultural species.     

Maturation and spawning processes of anadromous and resident pond smelt in lake ogawara

Pond smelt,Hypomesus nipponensis McAllister, in Lake Ogawara demonstrate alternative life history strategies, as evidenced by the coexistence of anadromous and resident fish. However, it is unknown if anadromous and resident groups interbreed. In this study, maturation and spawning processes were examined and compared between anadromous and resident groups. Histological observations indicated negligible variation in the maturational stage composition of oocytes, the frequency of oocyte diameter being unimodal for all specimens at different maturational stages. Oocytes were absent in the ovaries of spent fish. Accordingly, the species can be considered a semelparous spawner with unimodal oocyte diameter distribution. Temporal changes in the proportion of spent fish were compared between anadromous and resident groups. Spawning of both groups began in late March and peaked over April 8–12. Although both groups did not differ significantly in the period of peak spawning, anadromous fish finished spawning earlier than resident ones. Anadromous fish were not able to spawn upon migration into Lake Ogawara, and quickly matured after immigration, contrasting with resident fish.     

Functional analysis of the p57 KIP2 gene mutation in Beckwith-Wiedemann syndrome

p57 ^KIP2 is a potent tight-binding inhibitor of several G₁ cyclin/cyclin-dependent kinase (Cdk) complexes, and is a negative regulator of cell proliferation. The gene encoding p57 ^KIP2 is located at 11p15.5, a region implicated in both sporadic cancers and Beckwith-Wiedemann syndrome (BWS). Previously we demonstrated that p57 ^KIP2 is imprinted and only the maternal allele is expressed in both mice and humans. We also showed mutations found in p57 ^KIP2 in patients with BWS that were transmitted from the patients’ carrier mothers, indicating that the expressed maternal allele was mutant and that the repressed paternal allele was normal. In the study reported here, we performed functional analysis of the two mutated p57 ^KIP2 genes. We showed that the nonsense mutation found in the Cdk inhibitory domain in a BWS patient rendered the protein inactive with consequent complete loss of its role as a cell cycle inhibitor and of its nuclear localization. We also showed that the mutation in the QT domain, although completely retaining its cell cycle regulatory activity, lacked nuclear localization and was thus prevented from performing its role as an active cell cycle inhibitor. Consequently, no active p57 ^KIP2 would have existed, which might have caused the disorders in BWS patients. Received: 7 November 1998 / Accepted: 19 December 1998     

Isolation and characterization of a thermophilic Bacillus sp. JF8 capable of degrading polychlorinated biphenyls and naphthalene

Bacillus sp. strain JF8, which was isolated from compost, utilizes naphthalene and biphenyl as carbon sources at 60 degrees C. Biphenyl grown cells of strain JF8 barely degraded naphthalene while naphthalene grown cells did not degrade p-chlorobiphenyl, suggesting the existince of two independent degradation pathways. Isolation of JF8N, a mutant strain which can not utilize biphenyl as a carbon source while retaining the ability to utilize naphthalene, supports this hypothesis. Biphenyl grown cells of strain JF8 can degrade several polychlorinated biphenyl congeners including tetra- and pentachlorobiphenyl. bph and nah probes from mesophilic organisms failed to hybridize to strain JF8 DNA.     

Purification and characterization of a form of cytochrome P-450 with high specificity for aflatoxin B1 from 3-methylcholanthrene-treated hamster liver

A form of cytochrome P-450 highly active in inducing mutagenicity of aflatoxin B1 was purified to a specific content of 15.1 nmol/mg of protein from 3-methylcholanthrene-treated hamster liver. This species of cytochrome P-450, having its absorption maximum at 448.5 nm in carbon monoxide-complex of reduced form and low spin ferric ion, is of molecular weight of 56,000 and distinctly different in physicochemical and catalytic properties from major forms of cytochrome P-450 purified from phenobarbital- or 3-methylcholanthrene-treated rat liver. In the induction of aflatoxin B1 mutagenicity, this hamster cytochrome P-450 is 50 times more potent than those from rat liver.     

Lysine 624 of the amyloid precursor protein (APP) is a critical determinant of amyloid β peptide length: support for a sequential model of γ-secretase intramembrane proteolysis and regulation by the amyloid β precursor protein (APP) juxtamembrane region

γ-Secretase is a multiprotein intramembrane cleaving aspartyl protease (I-CLiP) that catalyzes the final cleavage of the amyloid β precursor protein (APP) to release the amyloid β peptide (Aβ). Aβ is the primary component of senile plaques in Alzheimer's disease (AD), and its mechanism of production has been studied intensely. γ-Secretase executes multiple cleavages within the transmembrane domain of APP, with cleavages producing Aβ and the APP intracellular domain (AICD), referred to as γ and ε, respectively. The heterogeneous nature of the γ cleavage that produces various Aβ peptides is highly relevant to AD, as increased production of Aβ 1-42 is genetically and biochemically linked to the development of AD. We have identified an amino acid in the juxtamembrane region of APP, lysine 624, on the basis of APP695 numbering (position 28 relative to Aβ) that plays a critical role in determining the final length of Aβ peptides released by γ-secretase. Mutation of this lysine to alanine (K28A) shifts the primary site of γ-secretase cleavage from 1-40 to 1-33 without significant changes to ε cleavage. These results further support a model where ε cleavage occurs first, followed by sequential proteolysis of the remaining transmembrane fragment, but extend these observations by demonstrating that charged residues at the luminal boundary of the APP transmembrane domain limit processivity of γ-secretase.     

Smad3 deficiency in mast cells provides efficient host protection against acute septic peritonitis

Mast cells play an important role in innate immunity as well as in allergic reaction. However, regulatory mechanisms underlying mast cell-mediated innate immune responses remain largely unknown. Here we determined whether Smad3, a major signal transducer of TGF-beta, regulates innate immune response by mast cells against Gram-negative bacteria. Bone marrow-derived mast cells (BMMC) obtained from Smad3 null mutant mice showed augmented capacity to produce proinflammatory cytokines upon stimulation with a Gram-negative bacteria-associated product, LPS. In acute septic peritonitis model induced by cecal ligation and puncture, mast cell-deficient W/W(v) mice reconstituted with Smad3 null BMMC had significantly higher survival rate than W/W(v) mice reconstituted with wild-type BMMC, which was associated with higher production of proinflammatory cytokines in the peritoneal cavity. These in vitro and in vivo results suggest that Smad3 in mast cells functions as inhibitory for mast cell-mediated innate immune response against Gram-negative bacteria. Suppression of Smad3 expression in mast cells may thus have therapeutic potential for Gram-negative bacterial infection such as acute septic peritonitis by augmenting innate immune responses of mast cells.