Involvement of MAK-1 and MAK-2 MAP kinases in cell wall integrity in Neurospora crassa.

Among three MAPK disruptants of Neurospora crassa, Δmak-1 was sensitive and Δmak-2 was hypersensitive to micafungin, a beta-1,3-glucan synthase inhibitor, than the wild-type or Δos-2 strains. We identified six micafungin-inducible genes that are involved in cell wall integrity (CWI) and found that MAK-1 regulated the transcription of non-anchored cell wall protein gene, ncw-1, and the beta-1,3-endoglucanase gene, bgt-2, whereas MAK-2 controlled the expression of the glycosylhydrolase-like protein gene, gh76-5, and the C4-dicarboxylate transporter gene, tdt-1. Western blotting analysis revealed that, in the wild-type strain, MAK-1 was constitutively phosphorylated from conidial germination to hyphal development.

New dipyridamole salt with improved dissolution and oral bioavailability under hypochlorhydric conditions.

The aim of this study was to develop new dipyridamole (DP) salts with pH-independent solubility for improving oral bioavailability under hypochlorhydria. Salt screening was carried out using nine counterions by the temperature gradient method. Six DP salts were obtained, and there was marked improvement in dissolution behavior for all DP salts in a neutral medium.

Deletion and expression analysis of beta-(1,3)-glucanosyltransferase genes in Neurospora crassa.

GPI(glycosylphosphatidylinositol)-anchored beta-(1,3)-glucanosyltransferases play an active role in cell wall biosynthesis in fungi. Neurospora crassa has 5 putative beta-(1,3)-glucanosyltransferase genes, namely, gel-1, gel-2, gel-3, gel-4, and gel-5, in its genome. Among them, the gel-3 gene is constitutively expressed at the highest level in growing hyphae, whereas gel-1 is expressed at the lowest level.

Novel formulations of dipyridamole with microenvironmental pH-modifiers for improved dissolution and bioavailability under hypochlorhydria.

This study was undertaken to develop new dipyridamole (DP) formulations with acidic microenvironmental pH-modifiers for improving dissolution and absorption under hypochlorhydric conditions. Dipyridamole granules (DPG) with ten acidic pH-modifiers were prepared with conventional wet granulation, and their manufacturability, stability and dissolution behavior were characterized. Pharmacokinetic profiling of the optimized DPG with acid was carried out in omeprazole-treated rats as a hypochlorhydric model.

Improved dissolution and pharmacokinetic behavior of dipyridamole formulation with microenvironmental pH-modifier under hypochlorhydria.

The present study aimed to develop and characterize new formulations of dipyridamole (DP), a pH-dependent poorly soluble drug, employing an acidic pH-modifier for improving dissolution and absorption under hypochlorhydric condition. Granule formulations of DP (DPG) with and without fumaric acid (FA) were prepared with wet granulation, physicochemical properties of which were characterized focusing on morphology, dissolution and stability. Pharmacokinetic profiling of orally dosed DPG or DPG with 60% loading of FA (DPG/FA60) was carried out in omeprazole-treated rats as a hypochlorhydric model.

An AP-1-like transcription factor, NAP-1, regulates expression of the glutathione S-transferase and NADH:flavin oxidoreductase genes in Neurospora crassa.

AP-1-like transcription factors play crucial roles in oxidative stress responses in yeast and filamentous fungi. The deletion of an AP-1-like transcription factor gene, nap-1, in Neurospora crassa slightly increased its sensitivity to oxidative stressors, including menadione. Microarray and quantitative real-time reverse transcriptase-PCR analyses were employed to identify menadione-inducible genes (migs) and the roles of NAP-1 in their regulation.

ATF-1 transcription factor regulates the expression of ccg-1 and cat-1 genes in response to fludioxonil under OS-2 MAP kinase in Neurospora crassa.

The ATF/CREB family transcriptional factors are regulated by stress-activated MAP kinase in yeast. The disruptants of the atf-1 gene, which encodes an ATF/CREB family transcriptional factor, were isolated and characterized in Neurospora crassa. The characteristic phenotypes in the os-2 MAP kinase strain, such as osmotic sensitivity and fludioxonil resistance, were not observed in the Deltaatf-1 strain; however, like the os-2 strain, up-regulation of the catalase gene cat-1 and the clock-controlled gene ccg-1 by treatment with fludioxonil (1 microg/mL) or 4% NaCl was almost completely abolished in the Deltaatf-1 strain.

OS-2 mitogen activated protein kinase regulates the clock-controlled gene ccg-1 in Neurospora crassa.

OS-2 MAP kinase is involved in osmoadaptation in Neurospora crassa. Clock-controlled genes ccg-1, bli-3, and con-10 were induced by osmotic stress in an OS-2 dependent manner. In contrast, osmotic stress did not affect the expression of clock genes frq, wc-1, and wc-2 or of clock-controlled genes ccg-2 and bli-4.

Involvement of OS-2 MAP kinase in regulation of the large-subunit catalases CAT-1 and CAT-3 in Neurospora crassa.

Neurospora crassa has four catalase genes--cat-1, cat-2, cat-3, and ctt-1/cat-4. cat-1 and cat-3 encode two fungal-specific large-subunit catalases CAT-1 and CAT-3 normally produced in conidia and growing hyphae, respectively. cat-2 encodes CAT-2 catalase-peroxidase normally produced in conidia.

Roles of putative His-to-Asp signaling modules HPT-1 and RRG-2, on viability and sensitivity to osmotic and oxidative stresses in Neurospora crassa.

Neurospora crassa has a putative histidine phosphotransfer protein (HPT-1) that transfers signals from 11 histidine kinases to two putative response regulators (RRG-1 and RRG-2) in its histidine-to-aspartate phosphorelay system. The hpt-1 gene was successfully disrupted in the os-2 (MAP kinase gene) mutant, but not in the wild-type strain in this study. Crossing the resultant hpt-1; os-2 mutants with the wild-type or os-1 (histidine kinase gene) mutant strains produced no progeny with hpt-1 or os-1; hpt-1 mutation, strongly suggesting that hpt-1 is essential for growth unless downstream OS-2 is inactivated.

Starch-branching enzyme I-deficient mutation specifically affects the structure and properties of starch in rice endosperm.

We have isolated a starch mutant that was deficient in starch-branching enzyme I (BEI) from the endosperm mutant stocks of rice (Oryza sativa) induced by the treatment of fertilized egg cells with N-methyl-N-nitrosourea. The deficiency of BEI in this mutant was controlled by a single recessive gene, tentatively designated as starch-branching enzyme mutant 1 (sbe1). The mutant endosperm exhibited the normal phenotype and contained the same amount of starch as the wild type.

Sildenafil for primary and secondary pulmonary hypertension.

Background: Sildenafil is a selective inhibitor of cyclic guanosine monophosphate-specific phosphodiesterase type 5, an enzyme that is abundant in both lung and penile tissues. Sildenafil is widely used to dilate penile arteries, suggesting that it may also dilate pulmonary arteries in patients with pulmonary hypertension. However, the long-term hemodynamic effects and safety of the drug in pulmonary hypertension are not known.