Inflorescence architecture: A developmental genetics approach

We are characterizing a suiteof Pisum sativum mutants that alter inflorescence architecture to construct a model for the genetic regulation of inflorescence development in a plant with a compound raceme. Such a model, when compared with those created forAntirrhinum majus andArabidopsis thaliana, both of which have simple racemes, should provide insight into the evolution of the development of inflorescence architecture. The highly conserved nature of cloned genes that regulate reproductive development in plants and the morphological similarities among our mutants and those identified inA. majus andA. thaliana enhance the probability that a developmental genetics approach will be fruitful. Here we describe sixP. sativum mutants that affect morphologically and architecturally distinct aspects of the inflorescence, and we analyze interactions among these genes. Both vegetative and inflorescence growth of the primary axis is affected byUNIFOLIA TA, which is necessary for the function ofDETERMINATE (DET).DET maintains indeterminacy in the first-order axis. In its absence, the meristem differentiates as a stub covered with epidermal hairs.DET interacts withVEGETATIVE1 (VEG1).VEG1 appears essential for second-order inflorescence (I₂) development.veg1 mutants fail to flower or differentiate the I₂ meristem into a rudimentary stub,det veg1 double mutants produce true terminal flowers with no stubs, indicating that two genes must be eliminated for terminal flower formation inP. sativum, whereas elimination of a single gene accomplishes this inA. thaliana andA. majus. NEPTUNE also affects I₂ development by limiting to two the number of flowers produced prior to stub formation. Its role is independent ofDET, as indicated by the additive nature of the double mutantdet nep. UNI, BROC, and PIM all play roles in assigning floral meristem identity to the third-order branch.pim mutants continue to produce inflorescence branches, resulting in a highly complex architecture and aberrant flowers.uni mutants initiate a whorl of sepals, but floral organogenesis is aberrant beyond that developmental point, and the double mutantuni pim lacks identifiable floral organs. A wild-type phenotype is observed inbroc plants, butbroc enhancesthe pim phenotype in the double mutant, producing inflorescences that resemble broccoli. Collectively these genes ensure that only the third-order meristem, not higher- or lower-order meristems, generates floral organs, thus precisely regulating the overall architecture of the plant. Gene symbols used in this article: For clarity a common symbolization is used for genes of all species discussed in this article. Genes are symbolized with italicized capital letters. Mutant alleles are represented by lowercase, italicized letters. In both cases, the number immediately following the gene symbol differentiates among genes with the same symbol. If there are multiple alleles, a hyphen followed by a number is used to distinguish alleles. Protein products are represented by capital letters without italics.     

Testing the relationship between foldability and the early folding events of dihydrofolate reductase from Escherichia coli

A "folding element" is a contiguous peptide segment crucial for a protein to be foldable and is a new concept that could assist in our understanding of the protein-folding problem. It is known that the presence of the complete set of folding elements of dihydrofolate reductase (DHFR) from Escherichia coli is essential for the protein to be foldable. Since almost all of the amino acid residues known to be involved in the early folding events of DHFR are located within the folding elements, a close relationship between the folding elements and early folding events is hypothesized. In order to test this hypothesis, we have investigated whether or not the early folding events are preserved in circular permutants and topological mutants of DHFR, in which the order of the folding elements is changed but the complete set of folding elements is present. The stopped-flow circular dichroism (CD) measurements show that the CD spectra at the early stages of folding are similar among the mutants and the wild-type DHFR, indicating that the presence of the complete set of folding elements is sufficient to preserve the early folding events. We have further examined whether or not sequence perturbation on the folding elements by a single amino acid substitution affects the early folding events of DHFR. The results show that the amino acid substitutions inside of the folding elements can affect the burst-phase CD spectra, whereas the substitutions outside do not. Taken together, these results indicate that the above hypothesis is true, suggesting a close relationship between the foldability of a protein and the early folding events. We propose that the folding elements interact with each other and coalesce to form a productive intermediate(s) early in the folding, and these early folding events are important for a protein to be foldable.     

Variation of glycosaminoglycan in the growth of transplantable tumors derived from a spontaneous ddY mouse mammary tumor

The components and variations of glycosaminoglycan (GAG) in the growth of transplantable tumors derived from a spontaneous mouse mammary tumor were investigated. A 45-week-old ddY female mouse, obtained from Shizuoka Laboratory Animal Center, was found to have a bean-sized mass at the third mammary gland of the left side. The tumor mass was surgically excised and used for transplantation in the present study. This mammary tumor was histologically found to be Type B-adenocarcinoma. Transplantable mammary tumors consisted of the fibrous or edematous interstitium contained a large amount of GAG components, which was mainly hyaluronic acid (HA), dermatan sulfate (DS) and chondroitin sulfate A/C (ChS). In the analysis of GAG components, HA content was present in a large amount in logarithmic growth phase of transplanted mammary tumors, but it was markedly decreased in stationary phase. On the other hand, the contents of DS and ChS increased in stationary phase of the tumor growth, and these increases corresponded, histologically, with the propagation of the fibrous interstitial tissues.     

Development of near-infrared firefly luciferin analogue reacted with wild-type and mutant luciferases

Interestingly, only the D-form of firefly luciferin produces light by luciferin–luciferase (L–L) reaction. Certain firefly luciferin analogues with modified structures maintain bioluminescence (BL) activity; however, all L-form luciferin analogues show no BL activity. To this date, our group has developed luciferin analogues with moderate BL activity that produce light of various wavelengths. For in vivo bioluminescence imaging, one of the important factors for detection sensitivity is tissue permeability of the number of photons emitted by L–L reaction, and the wavelengths of light in the near-infrared (NIR) range (700–900 nm) are most appropriate for the purpose. Some NIR luciferin analogues by us had performance for in vivo experiments to make it possible to detect photons from deep target tissues in mice with high sensitivity, whereas only a few of them can produce NIR light by the L–L reactions with wild-type luciferase and/or mutant luciferase. Based on the structure–activity relationships, we designed and synthesized here a luciferin analogue with the 5-allyl-6-dimethylamino-2-naphthylethenyl moiety. This analogue exhibited NIR BL emissions with wild-type luciferase (λ_max = 705 nm) and mutant luciferase AlaLuc (λ_max = 655 nm).     

Sleep-related factors associated with industrial accidents among factory workers and sleep hygiene education intervention

Sleep and Biological Rhythms - This study was conducted to investigate the association between industrial accidents and sleep-related parameters in factory workers, and to examine the effectiveness...     

Background Factors of Reflux Esophagitis and Non-Erosive Reflux Disease: A Cross-Sectional Study of 10,837 Subjects in Japan

Background

Despite the high prevalence of gastroesophageal reflux disease (GERD), its risk factors are still a subject of controversy. This is probably due to inadequate distinction between reflux esophagitis (RE) and non-erosive reflux disease (NERD), and is also due to inadequate evaluation of adjacent stomach. Our aim is therefore to define background factors of RE and NERD independently, based on the evaluation of Helicobacter pylori infection and gastric atrophy.

Methods

We analyzed 10,837 healthy Japanese subjects (6,332 men and 4,505 women, aged 20–87 years) who underwent upper gastrointestinal endoscopy. RE was diagnosed as the presence of mucosal break, and NERD was diagnosed as the presence of heartburn and/or acid regurgitation in RE-free subjects. Using GERD-free subjects as control, background factors for RE and NERD were separately analyzed using logistic regression to evaluate standardized coefficients (SC), odds ratio (OR), and p-value.

Results

Of the 10,837 study subjects, we diagnosed 733 (6.8%) as RE and 1,722 (15.9%) as NERD. For RE, male gender (SC = 0.557, OR = 1.75), HP non-infection (SC = 0.552, OR = 1.74), higher pepsinogen I/II ratio (SC = 0.496, OR = 1.64), higher BMI (SC = 0.464, OR = 1.60), alcohol drinking (SC = 0.161, OR = 1.17), older age (SC = 0.148, OR = 1.16), and smoking (SC = 0.129, OR = 1.14) are positively correlated factors. For NERD, HP infection (SC = 0.106, OR = 1.11), female gender (SC = 0.099, OR = 1.10), younger age (SC = 0.099, OR = 1.10), higher pepsinogen I/II ratio (SC = 0.099, OR = 1.10), smoking (SC = 0.080, OR = 1.08), higher BMI (SC = 0.078, OR = 1.08), and alcohol drinking (SC = 0.076, OR = 1.08) are positively correlated factors. Prevalence of RE in subjects with chronic HP infection and successful HP eradication denotes significant difference (2.3% and 8.8%; p<0.0001), whereas that of NERD shows no difference (18.2% and 20.8%; p = 0.064).

Conclusions

Significantly associated factors of NERD are considerably different from those of RE, indicating that these two disorders are pathophysiologically distinct. Eradication of Helicobacter pylori may have disadvantageous effects on RE but not on NERD.     

DNA polymerase III holoenzyme of Escherichia coli. I. Purification and distinctive functions of subunits tau and gamma, the dnaZX gene products

Escherichia coli dnaZX, the gene which when mutant blocks DNA chain elongation, was cloned into a lambda PL promoter-mediated expression vector. In cells carrying this plasmid, the activity that complements a mutant dnaZ extract in replicating a primed single-stranded DNA circle was increased about 20-fold. Two polypeptides of 71 and 52 kDa were overproduced. Upon fractionation, two complementing activities were purified to homogeneity and proved to be the 71- and 52-kDa polypeptides. Immunoassays revealed their respective identities with the tau and gamma subunits of DNA polymerase III holoenzyme. The N-terminal amino acid sequences of the first 12 residues were identical in both subunits, as were their molar specific activities in dnaZ complementation. Thus, the tau subunit complements the defect in the mutant holoenzyme from the dnaZts strain as efficiently as does the gamma subunit. Inasmuch as the 71-kDa subunit (tau) can also overcome the enzymatic defect in a dnaX mutant strain, this polypeptide has dual replication functions, only one of which can be performed by the gamma subunit. Availability of pure tau and gamma subunits for study has provided the basis for proposing an asymmetry in the structure and function of a dimeric DNA polymerase III holoenzyme.