Mouse Mutant Deficient in d-Amino Acid Oxidase Activity

D-Amino acid oxidase activity in the kidney homogenates of mice of seven strains was measured to search for a mutant for this enzyme. There was a consistent sex difference in the enzyme activity in these strains: male mice showed higher levels of the enzyme activity than females. In contrast to other strains, some mice of the ddY strain did not possess enzyme activity. This trait was inheritable, and a mouse stock without enzyme activity (DAO-) was established. The allele (Dao-1c) carried by the DAO- mice was recessive and behaved as a single autosomal gene in inheritance. Heterozygous mice for this gene (Dao-1+/Dao-1c) showed nearly half the enzyme activity of the wild-type homozygotes (Dao-1+/Dao-1+), suggesting that Dao-1c is a null allele and that there is a gene dosage effect on the enzyme activity.     

Neurospora Mutant Deficient in Tryptophanyl-Transfer Ribonucleic Acid Synthetase Activity

A tryptophan auxotroph of Neurospora crassa, trp-5, has been characterized as a mutant with a deficient tryptophanyl-transfer ribonucleic acid (tRNA) synthetase (EC 6.1.1.2) activity. When assayed by tryptophanyl-tRNA formation, extracts of the mutant have less than 5% of the wild-type specific activity. The adenosine triphosphate-pyrophosphate exchange activity is at about half the normal level. In the mutant derepressed levels of anthranilate synthetase and tryptophan synthetase were associated with free tryptophan pools equal to or higher than those found in the wild type. We conclude that a product of the normal tryptophanyl-tRNA synthetase, probably tryptophanyl-tRNA, rather than free tryptophan, participates in the repression of the tryptophan biosynthetic enzymes.     

Mutant of Escherichia coli Deficient in the Synthesis of cis-Vaccenic Acid

A temperature-sensitive unsaturated fatty acid (fabA) auxotroph of Escherichia coli was found also to be deficient in the elongation of palmitoleic acid to cis-vaccenic acid. Reversion and transductional analyses demonstrate that this second phenotype and the fabA mutation are independent in action and are not cotransduced. The deficiency in conversion of palmitoleic acid to cis-vaccenic acid was also demonstrated in vitro, and these results strongly suggest this phenotype is due to a deficiency in an elongation enzyme. We suggest that the phenotype may have been selected during growth because it can physiologically compensate for the fabA lesion. In fab(+) strains, the inability to synthesize cis-vaccenic acid is physiologically asymptomatic. Such strains grow normally at all temperatures tested and are not sodium sensitive. Although the parental strain has an increased amount of cis-vaccenic acid in cells grown at 15 C, the mutant does not. Since the mutant grows normally at 15 C, the data indicate that increased amounts of cis-vaccenic acid are not required for growth at 15 C.     

A Mutant of Arabidopsis with Increased Levels of Stearic Acid

A mutation at the fab2 locus of Arabidopsis caused increased levels of stearate in leaves. The increase in leaf stearate in fab2 varied developmentally, and the largest increase occurred in young leaves, where stearate accounted for almost 20% of total leaf fatty acids. The fatty acid composition of leaf lipids isolated from the fab2 mutant showed increased stearate in all the major glycerolipids of both the chloroplast and extrachloroplast membranes. Although the stearate content was increased, the fab2 mutant still contained abundant amounts of 18:1, 18:2, and 18:3 fatty acids. These results are consistent with the expectations for a mutation partially affecting the action of the stromal stearoyl-acyl carrier protein desaturase. Positional analysis indicated that the extra 18:0 is excluded with high specificity from the sn-2 position of both chloroplast and extrachloroplast glycerolipids. Although stearate content was increased in all the major leaf membrane lipids, the amount of increase varied considerably among the different lipids, from a high of 25% of fatty acids in phosphatidylcholine to a low of 2.9% of fatty acids in monogalactosyldiacylglycerol.     

Hormonal Studies of fass,an Arabidopsis Mutant That Is Altered in Organ Elongation

We have isolated an allele of fass, an Arabidopsis thaliana mutation that separates plant development and organ differentiation from plant elongation, and studied its hormonal regulation. Micro-surgically isolated fass roots elongate 2.5 times as much as the roots on intact mutant plants. Wild-type heart embryos, when cultured with a strong auxin, naphthaleneacetic acid, phenocopy fass embryos. fass seedlings contain variable levels of free auxin, which average 2.5 times higher than wild-type seedling levels, and fass seedlings evolve 3 times as much ethylene as wild-type seedlings on a per-plant basis over a 24-h period. The length-to-width ratios of fass seedlings can be changed by several compounds that affect their endogenous ethylene levels, but fass is epistatic to etr1, an ethylene-insensitive mutant. fass's high levels of free auxin may be inducing its high levels of ethylene, which may, in turn, result in the fass phenotype. We postulate that FASS may be acting as a negative regulator to maintain wild-type auxin levels and that the mutation may be in an auxin-conjugating enzyme.     

A Starch Deficient Mutant of Arabidopsis thaliana with Low ADPglucose Pyrophosphorylase Activity Lacks One of the Two Subunits of the Enzyme

A starch deficient mutant of Arabidopsis thaliana (L.) Heynh. has been isolated in which leaf extracts contain only about 5% as much activity of ADPglucose pyrophosphorylase (EC 2.7.7.27) as the wild type. A single, nuclear mutation at a previously undescribed locus designated adg2 is responsible for the mutant phenotype. Although the mutant contained only 5% as much ADPglucose pyrophosphorylase activity as the wild type, it accumulated 40% as much starch when grown in a 12 hour photoperiod. The mutant also contained about 40% as much starch as the wild type when grown in continuous light, suggesting that the rate of synthesis regulates its steady state accumulation. Immunological analysis of leaf extracts using antibodies against the spinach 54 and 51 kilodalton (kD) ADPglucose pyrophosphorylase subunits indicated that the mutant is deficient in a cross-reactive 54 kD polypeptide and has only about 4% as much as the wild type of a cross-reactive 51 kD polypeptide. This result and genetic studies suggested that adg2 is a structural gene which codes for the 54 kD polypeptide, and provides the first functional evidence that the 54 kD polypeptide is a required component of the native ADPglucose pyrophosphorylase enzyme.     

拟南芥草酸不敏感突变体的筛选与分析

草酸是多种真菌的致病因子.在含 1.2 mmol/L 草酸和 10 μmol/L 雌二醇的 MS 缺钙培养基上,从大约含 6000个独立株系的拟南芥化学诱导突变体库中筛选草酸不敏感的突变体.初筛获得的可能的草酸不敏感突变体单株收种后,进一步复筛获得 5 株较抗草酸的突变体 D33、D74、D154、D282 和 D630.对它们的 TAIL-PCR 的第三步产物回收、测序、比对的结果表明:D33 的 T-DNA 插入位点位于 At2g39720(Zinc finger)and At2g39730 (Rubisco activase) 之间,D74、D154、D282 和 D630 都插在 At5g10450 (14-3-3 protein GF14 lambda) 的第一个内含子上.突变体后继的遗传分析与分子分析正在进行中.     

拟南芥草酸不敏感突变体的筛选与分析

草酸是多种真菌的致病因子。在含1.2 mmol/L 草酸和10 mmol/L 雌二醇的MS缺钙培养基上, 从大约含6000个独立株系的拟南芥化学诱导突变体库中筛选草酸不敏感的突变体。初筛获得的可能的草酸不敏感突变体单株收种后, 进一步复筛获得5株较抗草酸的突变体D33、D74、D154、D282和D630。对它们的TAIL-PCR的第三步产物回收、测序、比对的结果表明：D33的T-DNA插入位点位于At2g39720 (Zinc finger ) and At2g39730 (Rubisco activase) 之间, D74、D154、D282和D630都插在At5g10450 (14-3-3 protein GF14 lambda) 的第一个内含子上。突变体后继的遗传分析与分子分析正在进行中。     

Enzymatic Formation of Hexadecenoic Acid from Palmitic Acid

Desaturation of palmitic acid was investigated in an enzyme system prepared from rat liver. 2-trans-Hexadecenoic acid as well as 9-cis-hexadecenoic acid (palmitoleic acid) were found to be formed as monoenoic acid in this system.     

Abscisic Acid Suppresses the Highly Occurred Somatic Homologous Recombination in Arabidopsis rfc1 Mutant

The phytohormone abscisic acid (ABA) regulates many aspects of plant growth, including seed germination, root growth and cell division. Previous study indicates that ABA treatment increases DNA damage and somatic homologous recombination (HR) in Arabidopsis abo4/pol ? (aba overly-sensitive 4 /DNA polymerase ?) mutants. DNA replication factor C (RFC) complex is required for loading PCNA (Proliferating Cell Nuclear Antigen) during DNA replication. The defect in RFC1, the largest subunit of RFC, causes the high HR and DNA damage sensitivity in Arabidopsis. Here we found that like pol ε/abo4, rfc1 is sensitive to ABA in both ABA-inhibiting seed germination and root growth. However, ABA treatment greatly reduces HR and also reduces the expression of the DNA-damaged marker genes in rfc1. These results suggest that RFC1 plays critical roles in ABA-mediated HR in Arabidopsis.     

单半乳糖甘油二脂缺失对烟草光合特性的影响

以单半乳糖甘油二脂(MGDG)相对含量比野生烟草显著降低的突变体(M18)及野生型烟草为材料,通过对转基因烟草叶绿体类囊体膜的低温荧光、放氧活性以及叶片的叶绿素荧光分析,研究MGDG部分缺失对烟草叶片光合特性的影响。结果表明,在低温下(77K)MGDG部分缺失并不影响烟草叶绿素荧光发射峰(F683和F730)的位置,但使光系统Ⅱ(PSⅡ)及光系统Ⅰ(PSⅠ)的荧光发射峰的强度减弱,F683/F730比值降低,直接影响激发能在PSⅡ和PSⅠ之间的均衡分配,使叶绿素a和叶绿素b之间的能量传递受阻,降低光能转化效率;MGDG部分缺失使PSⅡ放氧活性下降了72.9%;转基因烟草叶绿素荧光参数中最大光化学效率(Fv/Fm)、暗适应最大荧光(Fm)、实际光化学效率(Yield)、光化学猝灭系数(qP)比野生型烟草分别降低了7%、49%、32%和18%,并以Fm降幅最大。研究证明,MGDG在维持植物叶绿体类囊体膜的功能,特别是PSⅡ的功能方面起着重要的作用。     

A Mutant Escherichia coli Primase Defective in Elongation of Primer RNA Chains

Earlier we showed by affinity cross-linking of initiating substrates to Escherichia coli primase that one or more of the residues Lys211, Lys229, and Lys241 were involved in the catalytic center of the enzyme (A. A. Mustaev and G. N. Godson, J. Biol. Chem. 270:15711-15718, 1995). We now demonstrate by mutagenesis that only Lys241 but not Lys211 and Lys229 is part of the catalytic center. Primase with a mutation of Arg to Lys at position 241 (defined as K241R-primase) is almost unable to synthesize primer RNA (pRNA) on the single-stranded DNA-binding protein (SSB)/R199G4oric template. However, it is able to synthesize a pppApG dimer plus trace amounts of 8- to 11-nucleotide (nt) pRNA transcribed from the 5' CTG 3' pRNA initiation site on phage G4 oric DNA. The amount of dimer synthesized by K241R-primase is similar to that synthesized by the wild-type primase, demonstrating that the K241R mutant can initiate pRNA synthesis normally but is deficient in chain elongation. In the general priming system, the K241R-primase also can synthesize only the dimer and very small amounts of 11-nt pRNA. The results of gel retardation experiments suggested that this deficiency in pRNA chain elongation of the K241R mutant primase is unlikely to be caused by impairment of the DNA binding activity. The K241R mutant primase, however, can still prime DNA synthesis in vivo and in vitro.