Molybdoenzymes in Drosophila. IV. Further characterization of the cinnamon phenotype

Summary Mutations at the cin gene display drastically lowered levels of the molybdoenzymes, xanthine dehydrogenase (XDH) and aldehyde oxidase (AO), and lack pyridoxal oxidase (PO) and sulfite oxidase (SO) activities. Certain mutations at cin also display varying degrees of female sterility, which is maternally affected. Here we characterize five new cin alleles with respect to the molybdoenzyme activities as well as the molybdenum cofactor, commonly required for molybdoenzyme activity. In complementing cin heterozygotes we find that, in addition to the previously reported unusually high levels of XDH and AO activities, there are unusually elevated levels of SO activity, as well as complementation for PO activity. The levels of immunologically crossreacting material in such heterozygotes indicate that the elevated levels of molybdoenzyme activities cannot be due to increases in the number of enzyme molecules. Measurements of the level of molybdenum cofactor activity normally present in XDH, AO, PO, and SO point to the possibility that a larger fraction of the enzyme molecules are active in these heterozygotes. The possible role of SO with respect to cinnamon's female sterility is also discussed.     

Inhomogeneity during deflation of excised canine lungs. I. Alveolar pressures

Factors both intrinsic and extrinsic to the lung may cause inhomogeneity of alveolar pressures during deflation. Wilson et al. (J. Appl. Physiol. 59: 1924-1928, 1985) predicted that any such inhomogeneity would be limited by interdependence of regional expiratory flows. To test this hypothesis and to explore how the pleural pressure gradient might affect inhomogeneity of alveolar pressures, we deflated at submaximal flows excised canine lobes that first were suspended in air and then were immersed in foams that simulated the vertical gradient of pleural pressure. Interregional inhomogeneity of regional transpulmonary pressures was measured with use of an alveolar capsule technique. Flow-dependent inhomogeneity of alveolar pressures was present, with differences in alveolar pressure quickly relaxing to a constant limiting value at each flow. Foam immersion increased inhomogeneity at a given flow. We conclude that factors intrinsic to the lung cause significant inhomogeneity of alveolar pressures at submaximal expiratory flows and that this inhomogeneity is enhanced by the extrinsic gradient of pleural pressure. These observations are consistent with the interdependence of flow proposed by Wilson et al.     

Isolation and mapping of polymorphic cosmid clones used for sublocalization of the multiple endocrine neoplasia type 1 (MEN1) locus

Summary Multiple endocrine neoplasia type 1 (MEN1) is characterized by neoplasia of the parathyroids, the pancreas, and the pituitary. Tumorigenesis involves unmasking of a recessive mutation at the MEN1 locus, which has been mapped to the centromeric part of chromosomal region 11q. In order to localize the MEN1 gene further and to make its isolation possible, a number of new markers were isolated. Two radiation-reduced somatic cell hybrids were identified that only contained markers close to and flanking the MEN1 region. DNA from these hybrids was used for the construction of a cosmid library, and clones containing human inserts were isolated. In addition, cosmid clones were isolated for locus expansion of 7 other markers that were mapped to the 11q12–13.2 region. The 33 newly isolated clones together with 25 previously published markers from this region were analyzed in a panel of radiation-reduced somatic cell hybrids. From the hybridization pattern, the region was divided into 11 parts. New restriction fragment length polymorphisms were identified in 7 of the newly isolated cosmid clones and in one plasmid. These were then used to sublocalize meiotic cross-overs more precisely in two MEN1 families, thus refining the mapping of the disease gene.     

Hydrotropism and Its Interaction with Gravitropism in Maize Roots

We have partially characterized root hydrotropism and its interaction with gravitropism in maize (Zea mays L.). Roots of Golden Cross Bantam 70, which require light for orthogravitropism, showed positive hydrotropism; bending upward when placed horizontally below a hydrostimulant (moist cheesecloth) in 85% relative humidity (RH) and in total darkness. However, the light-exposed roots of Golden Cross Bantam 70 or roots of a normal maize cultivar, Burpee Snow Cross, showed positive gravitropism under the same conditions; bending downward when placed horizontally below the hydrostimulant in 85% RH. Light-exposed roots of Golden Cross Bantam 70 placed at 70° below the horizontal plane responded positively hydrotropically, but gravitropism overcame the hydrotropism when the roots were placed at 45° below the horizontal. Roots placed vertically with the tip down in 85% RH bent to the side toward the hydrostimulant in both cultivars, and light conditions did not affect the response. Such vertical roots did not respond when the humidity was maintained near saturation. These results suggest that hydrotropic and gravitropic responses interact with one another depending on the intensity of one or both factors. Removal of the approximately 1.5 millimeter root tip blocked both hydrotropic and gravitropic responses in the two cultivars. However, removal of visible root tip mucilage did not affect hydrotropism or gravitropism in either cultivar.     

Normal mouse strains differ in the site of initiation of closure of the cranial neural tube

The scanning electron microscopic study of day 9 embryos reported here documents differences among normal mouse strains in morphology of cranial neural tube closure. The site of initiation of contact and fusion of the cranial neural folds, previously defined as Closure 2 (Macdonald et al., '89), is located in the region of the junction between the forebrain (prosencephalon) and midbrain (mesencephalon) in three normal strains: LM/Bc, AEJ/RkBc, and ICR/Bc. However in a fourth normal strain, SWV/Bc, Closure 2 is initiated much further rostral, in the prosencephalic region. In addition, the anterior neuropore, rostral to Closure 2, closes late in ICR/Bc embryos, relative to the posterior progress of development of the Closure 2 seam. Initiation of closure from the most rostral end of the neural tube (Closure 3) appears to be relatively delayed in ICR/Bc embryos. We hypothesize that the observed genetic polymorphism in location of the first site of fusion between the cranial neural folds in normal mouse embryos may be one basis for differences among normal strains in liability to exencephaly induced by teratogens.     

Studies of the effect of retinoic acid on anterior neural tube closure in mice genetically liable to exencephaly

Previously we have shown that all SELH/Bc mouse embryos close their anterior neural tubes by an abnormal mechanism and that 10-20% of SELH/Bc embryos are exencephalic. The purposes of these studies were (1) to observe the effects of retinoic acid on the frequency of exencephaly in SELH/Bc embryos; (2) to compare the SELH/Bc response with those of normal strains and of other neural tube mutants; and (3) to compare, between SELH/Bc and a normal strain (SWV/Bc), the effects of retinoic acid on morphology of the closing anterior neural tube. SELH/Bc was more liable to retinoic acid-induced exencephaly than were normal strains. After maternal treatment with 5 mg/kg retinoic acid on day 8.5 of gestation, 53% of SELH/Bc embryos had exencephaly, compared with 22% in ICR/Bc and 14% in SWV/Bc. When these results were transformed according to the assumptions of the developmental threshold model, the effects of genotype and retinoic acid appeared to be additive. Similar treatment on day 9 or 10 of gestation had little or no effect on the frequency of exencephaly in SELH/Bc mice. These results are similar to the reported responses of the curly-tail and Splotch mutants, where frequencies of spina bifida but not exencephaly were decreased. This pattern suggests that studies of effects of periconceptional vitamin treatment on risk of human neural tube defects should consider anencephaly and spina bifida separately. The study comparing the morphology of anterior neural tube closure in SELH/Bc and normal SWV/Bc embryos showed that retinoic acid delays the elevation of the mesencephalic neural folds. This results in a "stalling" of many embryos in the first steps of neural tube closure, with their neural folds remaining convex and splayed wide apart. The delay in fold elevation was superimposed on the different closure patterns of the two strains. The overall conclusion is that there is no nonadditive interaction in the parameters studied between retinoic acid treatment and the SELH/Bc genotype.     

Influence of mefluidide on growth,development, and cell wall digestibility of sorghum

Application of mefluidide (N-[2,4-dimethyl-5-([(trifluoromethyl)sulfonyl]amino) phenyl]acetamide) inhibits plant development in perennial grasses. This study examined the effect of mefluidide on the morphological development and digestibility of sorghum. In the greenhouse, 5.9 × 10^–5 g active ingredient (a.i.) plant^–1 applied at the seedling, eight-leaf and boot stages reduced mean plant height 70%, 59%, and 2%, respectively. Heights were also reduced 14%, 15% and 35% by 5.9 × 10^–8, 5.9 × 10^–7 and 5.9 × 10^–6 gram a.i. plant^–1 applied at the eight-leaf stage. Field application of 0.26 or 0.52 kg ha^–1 mefluidide at either the eight-leaf or flagleaf stage reduced mean plant height of all cultivars. Basal tiller numbers increased 319% 28 d, and dry matter production was reduced 65% 42 d following mefluidide application at the eight-leaf stage. Treated stems were 34% higher and treated leaves were 7% higher in cellulase dry matter digestibility than control plants following mefluidide application at the eight-leaf stage. These results indicate that mefluidide application to vegetative stages in sorghum may enhance the forage value of the plants while it inhibits normal plant growth.     

Stimulated indole alkaloid release from Catharanthus roseus immobilized cultures. Initial studies.

Vacuolar sequestration of valuable secondary metabolites remains the major limitation to the use of immobilization technology for large scale plant-cell-based bioprocesses, which otherwise may be a more efficient culture system than suspension for this biomass. In this initial study, the release of indole alkaloids produced by immobilized Catharanthus roseus cells cultured in Zenk's Alkaloid Production Medium was evaluated. Unstimulated alkaloid release in immobilized cultures reached levels of 10 to 50% of total production or 3 to 100% of known alkaloid content (30 to 4700 micrograms l-1), which was higher than that found for suspension cultures of the cell line used (10 to 25% of total production) without apparent cell lysis. Modifications of the medium pH value of immobilized cultures were explored in order to improve this release. Periodical additions of acid (HCl 0.1 N) or base (KOH 0.1 N) solutions (2% v/v) to different cultures resulted in rapid (less than 3 h) and transient variations in extracellular pH value from 5.5 to 4.3, and 5.8 to 8.5, respectively. In both cases, these variations provoked significant increase in total alkaloid (from approximately 5-10 mg l-1 to 15 mg l-1), ajmalicine (from 0 to approximately 0.29 mg l-1) and serpentine (from 0 to approximately 0.20 mg l-1) release, without apparent cell lysis or decrease in the culture viability. This product release was estimated to represent 100% of alkaloids produced.     

Mapping of the SLA complex of miniature swine: mapping of the SLA gene complex by pulsed field gel electrophoresis.

The overall order of the regions of the swine major histocompatibility complex (MHC), the SLA complex, was determined by pulsed field gel electrophoresis (PFGE). It was found that the order of the regions is class II-class III-class I. A class I probe hybridized to a 420 kb Mlu I and a 420 kb Not I fragment as did a class III probe for C2. None of the class II probes hybridized to these fragments. Thus, linkage of class I to class III was shown. The class III C2, Bf, and C4 genes were found to residue in a 190 kb Not I fragment. Linkage of class III and class II genes was shown when both the class III C4 and the class II DR probes hybridized to the same 195 kb Sac II and 340 kb Not I fragments. The class I probe did not hybridize to these fragments. The order of the regions, class II-class III-class I, is similar to that of human MHC genes and may have been conserved in evolution so that coordinated expression of MHC genes could be achieved.