Absolute configuration and tautomeric structure of xylindein, a blue-green pigment of Chlorociboria species

The S absolute configuration of both chiral centers of xylindein was assigned using X-ray crystallographic heavy atom analysis after its conversion to a synthetic derivative. Crystallographic analysis of xylindein crystallized with phenols revealed that the proposed structure is the proper tautomer in the crystals.     

Molecular cloning and expression of the mouse N-acetylneuraminic acid 9-phosphate synthase which does not have deaminoneuraminic acid (KDN) 9-phosphate synthase activity

A cDNA of the mouse homologue of Escherichia coli N-acetylneuraminic acid (Neu5Ac) synthase (neuB gene product) was cloned by the PCR-based method. The mouse homologue consists of 359 amino acids, and the cDNA sequence displays 33% identity to that of the E. coli Neu5Ac synthase. The recombinant mouse homologue which is transiently expressed in HeLa cells does not exhibit the Neu5Ac synthase activity, which catalyzes condensation of phosphoenolpyruvate (PEP) and N-acetylmannosamine (ManNAc) to synthesize Neu5Ac, but the Neu5Ac 9-phosphate (Neu5Ac-9-P) synthase activity, which catalyzes condensation of PEP and ManNAc 6-phosphate (ManNAc-6-P) to synthesize Neu5Ac-9-P. Thus, the mouse homologue of E. coli Neu5Ac synthase is the Neu5Ac-9-P synthase. The Neu5Ac-9-P synthase is a cytosolic enzyme and ubiquitously distributed in mouse various tissues. Notably, the Neu5Ac-9-P synthase can not catalyze the synthesis of deaminoneuraminic acid (KDN) or KDN-9-P from PEP and Man or ManNAc-6-P, thus suggesting that the enzyme is not involved in the synthesis of KDN. This is consistent with the previous observation that only a very low activity to synthesize KDN is found in mouse B16 cells [Angata, T., et al. (1999) Biochem. Biophys. Res. Commun. 261, 326-331].     

Diapause and survival in winter in two species of predatory bugs,Orius sauteri and O. minutus

The cause of differences in overwintering success between the sexes in Orius sauteri (Poppius) and O. minutus (Linnaeus) (Heteroptera: Anthocoridae) was investigated in the laboratory. The survival rate of adults was examined in a screen house outdoors during the winters of 1995–1996 and 1996–1997. None of the males of either species survived until spring in either year. However, 63.9% and 40.5% of the females in O. sauteri and O. minutus, respectively, survived the winter in 1995–1996, and 54.5% and 43.2%, respectively, in 1996–1997. Most of the males died by early winter. In both species, the adult females reared under short days (= L11:D13) survived for a long period at 5 or 0°C, while the males reared under the same photoperiod died shortly after transfer to 5 or 0°C from 22°C. The males and females kept at same temperature conditions under long days (= L16:D8) died early. When nymphs were kept under long days at 22°C, the lipid content in the female and male adults of O. sauteri was 27.9% and 17.7%, respectively. When nymphs were kept under short days, their lipid content was significantly higher (41.1%) than that of those reared under long days for females, but lipid content was comparable in the males regardless of photoperiod (15.6%). In O. minutus, the mean lipid content was 27.2% in females and 19.0% in males under long days at 22°C, and 40.8% in females and 19.8% in males under short days. Thus, a large amount of lipid was accumulated only in females kept under short days in both species. These results suggest that short days induced diapause only in adult females but not in males. Due to the lack of diapause and shortage of lipid accumulation, males may not be able to survive the winter.     

Effectiveness of weekly subcutaneous recombinant human erythropoietin administration for chemotherapy-induced anemia

The effects of weekly subcutaneousrecombinant human erythropoietin (r-hEPO)administration on anemia during chemotherapy includingcisplatin and 5-fluorouracil in patients with head andneck carcinomas were examined. Weekly subcutaneousr-hEPO administration in cancer patients has not beeninvestigated previously. Patients were treated withr-hEPO 100 IU/kg (2 patients), 200 IU/kg (6 patients),or 400 IU/kg (5 patients), or placebo, andeffectiveness was evaluated by monitoring hemoglobinconcentration changes after administration for 8weeks. Hemoglobin concentrations in all 3 r-hEPOdosage groups were higher than that in the controlgroup during chemotherapy. All r-hEPO doses producedimprovements in the anemia induced by chemotherapy;however, the 400 IU/kg dose was most effective. Therequirement for blood transfusions decreased inpatients receiving r-hEPO therapy, and no significantside-effects were associated with r-hEPOadministration. These results suggest thatchemotherapy-induced anemia can be prevented by weeklysubcutaneous r-hEPO administration.     

Growth and maturation of the 'autumn-fruiting type' of Sargassum horneri (Fucales, Phaeophyta) and comparisons with the 'spring-fruiting type'

The growth and maturation period of the autumn-fruiting population ot Sargassum horneri C. Agardh (Phaeophyta) was investigated in Hiroshima Bay, Seto Inland Sea. Surveyed traits were compared to those of the spring-fruiting type and ecological features of this species were discussed. The annual lifetime of the autumn-fruiting population could be divided into four phases according to the daily increase in thalius length:phase I from December to May (increase in length < 0.1 mm/ day), phase II from May to September (= 0.3–1.0 mm/ day), phase Ml (from September to December > 10 mm/day) and phase IV which was the senescence phase from December to March. Receptacle formation‘as observed in November and gamete release from November to February. Conversely, the spring-fruiting type germinated in April and exhibited swifter growth in its early stage of development than the autumn-fruiting type. Rapid increase in thalius iength in autumn was common in both fruiting types, although the spring-fruiting type continued to grow during winter. Receptacle formation of the spring-fruiting type started in February but gamete release was not observed until April and May. The difference in life-history patterns of both types of S. horneri was in the overwintering period. The autumn-fruiting type spent that season as germi-ings or as young plants exhibiting slow-paced growth, while the spring-fruiting type overvwintered as adult thal-li preparing for gamete release in spring.     

Inhaled nitric oxide prevents left ventricular impairment during endotoxemia

We evaluated theeffect of long-term inhalation of nitric oxide (NO) on cardiaccontractility after endotoxemia by using the end-systolicelastance of the left ventricle (LV) as a load-independent contractility index. Chronic instrumentation in 12 pigs included implantation of two pairs of endocardial dimension transducers tomeasure LV volume and a micromanometer to measure LV pressure. One weeklater, the animals were divided into a control group (n = 6) or a NO group(n = 6). All animals receivedintravenous Escherichia coliendotoxin (10 µg · kg¹ · h¹)and equivalent lactated Ringer solution. NO inhalation (20 parts/million) was begun 30 min after the initiation of endotoxemia andwas continued for 24 h. In both groups, tachycardia, pulmonaryhypertension, and systemic hyperdynamic changes were noted. Theend-systolic elastance in the control group was significantly decreasedbeyond 7 h. NO inhalation maintained the end-systolic elastance atbaseline levels and prevented its impairment. These findings indicatethat NO exerts a protective effect on LV contractility in this model of endotoxemia.

     

The MAP kinase kinase kinase MLK2 co-localizes with activated JNK along microtubules and associates with kinesin superfamily motor KIF3.

The MLK (mixed lineage) ser/thr kinases are most closely related to the MAP kinase kinase kinase family. In addition to a kinase domain, MLK1, MLK2 and MLK3 each contain an SH3 domain, a leucine zipper domain and a potential Rac/Cdc42 GTPase-binding (CRIB) motif. The C-terminal regions of the proteins are essentially unrelated. Using yeast two-hybrid analysis and in vitro dot-blots, we show that MLK2 and MLK3 interact with the activated (GTP-bound) forms of Rac and Cdc42, with a slight preference for Rac. Transfection of MLK2 into COS cells leads to strong and constitutive activation of the JNK (c-Jun N-terminal kinase) MAP kinase cascade, but also to activation of ERK (extracellular signal-regulated kinase) and p38. When expressed in fibroblasts, MLK2 co-localizes with active, dually phosphorylated JNK1/2 to punctate structures along microtubules. In an attempt to identify proteins that affect the activity and localization of MLK2, we have screened a yeast two-hybrid cDNA library. MLK2 and MLK3 interact with members of the KIF3 family of kinesin superfamily motor proteins and with KAP3A, the putative targeting component of KIF3 motor complexes, suggesting a potential link between stress activation and motor protein function.     

Stat3 links activated keratinocytes and immunocytes required for development of psoriasis in a novel transgenic mouse model

Here we report that epidermal keratinocytes in psoriatic lesions are characterized by activated Stat3. Transgenic mice with keratinocytes expressing a constitutively active Stat3 (K5.Stat3C mice) develop a skin phenotype either spontaneously, or in response to wounding, that closely resembles psoriasis. Keratinocytes from K5.Stat3C mice show upregulation of several molecules linked to the pathogenesis of psoriasis. In addition, the development of psoriatic lesions in K5.Stat3C mice requires cooperation between Stat3 activation in keratinocytes and activated T cells. Finally, abrogation of Stat3 function by a decoy oligonucleotide inhibits the onset and reverses established psoriatic lesions in K5.Stat3C mice. Thus, targeting Stat3 may be potentially therapeutic in the treatment of psoriasis.     

Quantification of self-renewal capacity in single hematopoietic stem cells from normal and Lnk-deficient mice

Despite being a hallmark of hematopoietic stem cells (HSCs), HSC self-renewal has never been quantitatively assessed. Establishment of a clonal and quantitative assay for HSC function permitted demonstration that adult mouse HSCs are significantly heterogeneous in degree of multilineage repopulation and that higher repopulating potential reflects higher self-renewal activity. An HSC with high repopulating potential could regenerate approximately 1000 HSCs, whereas the repopulating activity of regenerated HSCs on average was significantly reduced, indicating extensive but limited self-renewal capacity in HSCs. Comparisons of wild-type mice with mutant mice deficient in the signal adaptor molecule Lnk showed that not only HSC numbers but also the self-renewal capacity of some HSCs are markedly increased when Lnk function is lost. Lnk appears to control HSC numbers by negatively regulating HSC self-renewal signaling.     

Interaction analysis between tmRNA and SmpB from Thermus thermophilus

Small protein B, SmpB, is a tmRNA-specific binding protein essential for trans-translation. We examined the interaction between SmpB and tmRNA from Thermus thermophilus, using biochemical and NMR methods. Chemical footprinting analyses using full-length tmRNA demonstrated that the sites protected upon SmpB binding are located exclusively in the tRNA-like domain (TLD) of tmRNA. To clarify the SmpB binding sites, we constructed several segments derived from TLD. Optical biosensor interaction analyses and melting profile analyses with mutational studies showed that SmpB efficiently binds to only a 30-nt segment that forms a stem and loop, with the 5' and 3' extensions composed of the D-loop and variable-loop analogues. The conserved sequences, 16UCGA and 319GAC, in the extensions are responsible for the SmpB binding. These results agree with the those visualized by the cocrystal structure of TLD and SmpB from Aquifex aeolicus. In addition, NMR chemical shift mapping analyses, using the 30-nt segment and (15)N-labeled SmpB, revealed the characteristic RNA binding mode. The hydrogen bond pattern around beta2 changes, with the Gly in beta2, which acts as a hinge, showing the largest chemical shift change. It appears that SmpB undergoes structural changes indicating an induced fit upon binding to the specific region of TLD.