RB1CC1 together with RB1 and p53 predicts long-term survival in Japanese breast cancer patients

RB1-inducible coiled-coil 1 (RB1CC1) plays a significant role in the enhancement of the retinoblastoma tumor suppressor (RB1) pathway and is involved in breast cancer development. However, RB1CC1's role in clinical progression of breast cancer has not yet been evaluated, so, as a first step, it is necessary to establish its usefulness as a tool to evaluate breast cancer patients. In this report, we have analyzed the correlation between abnormalities in the RB1CC1 pathway and long-term prognosis, because disease-specific death in later periods (>5 years) of the disease is a serious problem in breast cancer. Breast cancer tissues from a large cohort in Japan were evaluated by conventional immunohistochemical methods for the presence of the molecules involved in the RB1CC1 pathway, including RB1CC1, RB1, p53, and other well-known prognostic markers for breast cancer, such as estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. The correlation between the immunohistochemical results and clinical outcomes of 323 breast cancer patients was analyzed using a Kaplan-Meier log-rank test and a multivariate Cox proportional hazards regression analysis. Absence of nuclear RB1CC1 expression was associated with the worst prognosis (Log-rank test, Chi-Square value = 17.462, p<0.0001). Dysfunction of either one of RB1CC1, RB1, or p53 was associated with the highest risk for cancer-specific death, especially related to survival lasting more than 5 years (multivariate Cox proportional hazard ratio = 3.951, 95% Confidence Interval =1.566-9.967, p = 0.0036). Our present data demonstrate that the combined evaluation of RB1CC1, RB1 and p53 by conventional immunohistochemical analysis provides an accurate prediction of the long-term prognoses of breast cancer patients, which can be carried out as a routine clinical examination.     

Adrenomedullin insufficiency increases allergen-induced airway hyperresponsiveness in mice.

Adrenomedullin (ADM), a newly identified vasodilating peptide, is reported to be expressed in lungs and have a bronchodilating effect. We hypothesized whether ADM could be involved in the pathogenesis of bronchial asthma. We examined the role of ADM in airway responsiveness using heterozygous ADM-deficient mice (AM+/-) and their littermate control (AM+/+). Here, we show that airway responsiveness is enhanced in ADM mutant mice after sensitization and challenge with ovalbumin (OVA). The immunoreactive ADM level in the lung tissue after methacholine challenge was significantly greater in the wild-type mice than that in the mutant. However, the impairment of ADM gene function did not affect immunoglobulins (OVA-specific IgE and IgG1), T helper 1 and 2 cytokines, and leukotrenes. Thus the conventional mechanism of allergen-induced airway responsiveness is not relevant to this model. Furthermore, morphometric analysis revealed that eosinophilia and airway hypersecretion were similarly found in both the OVA-treated ADM mutant mice and the OVA-treated wild-type mice. On the other hand, the area of the airway smooth muscle layer of the OVA-treated mutant mice was significantly greater than that of the OVA-treated wild-type mice. These results suggest that ADM gene disruption may be associated with airway smooth muscle hyperplasia as well as enhanced airway hyperresponsiveness. ADM mutant mice might provide novel insights to study the pathophysiological role of ADM in vivo.     

A cytokinin-repressed gene in cucumber for a bHLH protein homologue is regulated by light

CRR12 was identified as a cytokinin-repressed gene encoding a cucumber homologue for a basic region/helix-loop-helix protein. The level of CRR12 mRNA decreased in response to either cytokinins or light in etiolated cotyledons. The level was low in cotyledons and leaves of light-grown plants, but it increased during dark incubation.     

Identification of antigenic epitopes recognized by Mac-2 binding protein-specific cytotoxic T lymphocytes for use in cancer immunotherapy

We have previously reported that 90K/Mac-2 binding protein (M2BP) was highly expressed in lung cancer and that M2BP-specific immunity was observed in many of cancer patients. In this study, we analyzed the ability of 11 M2BP-derived oligopeptides with an HLA-A*0201-binding motif to induce M2BP-specific cytotoxic T lymphocytes (CTL) from peripheral blood lymphocytes of normal donors by in vitro stimulation. One of the CTLs that were induced using M2BP216-224 (RIDITLSSV) produced interferon-gamma in response to HLA-A2-positive T2 cells pulsed with the same peptide and lysed MDA-MB-231 cells expressing both M2BP and HLA-A2. The cytolytic activities were blocked by antibodies against HLA class I or CD8. These findings suggest that M2BP216-224 is naturally processed from the native M2BP in cancer cells and recognized by M2BP-specific CTLs in an HLA-A2 restriction. We first identified M2BP-derived CTL epitopes that may be useful as a target antigenic epitope in clinical immunotherapy of cancer.     

Structural and functional analysis of a lycopene beta-monocyclase gene isolated from a unique marine bacterium that produces myxol

A gene coding for lycopene beta-monocyclase, which metabolizes lycopene (psi,psi-carotene) to gamma-carotene (beta,psi-carotene), was isolated for the first time from a unique marine bacterium strain P99-3 that produces myxol (a gamma-carotene derivative). This lycopene beta-monocyclase gene (designated crtYm) was included in the gene cluster which contained carotenoid biosynthetic gene (crtI, crtB, crtZ, crtY, and crtA) homologs. CrtYm, the CrtY homolog, metabolized lycopene to gamma-carotene, which was confirmed by deletion/expression analysis of the crtYm and by subsequent analysis of the metabolites from lycopene based on the retention times on high-performance liquid chromatography, UV-visible absorption spectra, and mass spectrometry.     

Fungal cytochrome P450s catalyzing hydroxylation of substituted toluenes to form their hydroxymethyl derivatives

The degradation of a series of nitroaromatic compounds by the lignin-degrading fungus Phanerochaete chrysosporium was examined. From 4-nitrotoluene (4-NT), several metabolic intermediates were identified. Initially, 4-NT was converted to 4-nitrobenzyl alcohol (4-NBA), followed by the oxidation reactions to form 4-nitrobenzaldehyde and 4-nitrobenzoic acid, albeit slowly. Exogenously added 4-nitrobenzaldehyde and 4-nitrobenzoic acid were predominantly reduced to 4-NBA. The fungal formation of 4-NBA was inhibited by piperonyl butoxide, a cytochrome P450 inhibitor, suggesting the involvement of cytochrome P450 in the hydroxylation of the methyl group. Similarly, 2-, and 3-nitrotoluenes and 4-chlorotoluene were converted to the corresponding arylalcohols by P. chrysosporium. On the other hand, toluene and 4-methoxytoluene were not converted. Thus, P. chrysosporium possesses an alkyl hydroxylation activity against aromatic compounds substituted with a strong electron-withdrawing group.     

The light-harvesting complex of photosystem I in Chlamydomonas reinhardtii: protein composition, gene structures and phylogenic implications

Light-harvesting chlorophyll a/b-binding proteins (LHCI) associated with photosystem I (PSI) and the genes encoding these proteins have been characterized in the unicellular green alga Chlamydomonas reinhardtii, extending previous studies of the PSII-LHCII [Teramoto et al. (2001) Plant Cell Physiol. 42: 849]. In order to assign LHCI proteins in the thylakoid membranes, the PSI-LHCI supercomplex that retains all of the major LHCI proteins was purified. Seven distinct LHCI proteins were resolved from the purified supercomplex by a high-resolution SDS polyacrylamide gel electrophoresis, and their N-terminal amino acid sequences were determined. One LHCI protein (band e) was newly found, although the other six LHCI proteins corresponded to those previously reported. Genomic clones encoding these seven LHCI proteins were newly isolated and the nucleotide sequences were determined. A comprehensive characterization of all members of Lhc gene family in this alga revealed that LHCI proteins are more highly diverged than LHCII, suggesting functional differentiation of the protein components in LHCI. Neighbor joining trees were constructed for LHC proteins from C. reinhardtii and those of Arabidopsis thaliana or Galdieria sulphuraria to assess evolutionary relationships. Phylogenetic analysis revealed that (1). green algal LHCI and LHCII proteins are more closely related to one another than to LHCI proteins in red algae, (2). green algae and higher plants possess seven common lineages of LHC proteins, and (3). Type I and III LHCI proteins are conserved between green algae and higher plants, while Type II and IV are not. These findings are discussed in the context of evolution of multiple diverse antenna complexes.     

Spatial heterogeneity of TNF-alpha-induced T cell migration to colonic mucosa is mediated by MAdCAM-1 and VCAM-1

Relatively little is known about how recirculation of lymphocytes through the inflamed intestinal mucosa is regulated. The aim of this study was to investigate the dynamic process of T lymphocyte-endothelial cell adhesion in TNF-alpha-challenged murine colonic mucosa by intravital microscopy. T lymphocytes from spleen (SPL) and intestinal lamina propria (LPL) were fluorescence labeled, and their adhesion to microvessels in the colonic mucosa was observed. In TNF-alpha (25 microg/kg)-stimulated colonic venules, an enhanced adhesion of SPL and LPL was demonstrated, with dominant recruitment of LPLs. The magnitude of the increased LPL adhesion was more significant in the colon than in the small intestine. These T lymphocyte interactions in the colonic mucosa were significantly reduced by blocking MAbs against either mucosal addressin cell adhesion molecule-1 (MAdCAM-1), VCAM-1, alpha(4)-integrin, or beta(7)-integrin but not by anti-ICAM-1. Immunohistochemistry revealed significant MAdCAM-1 expression in the lamina propria and VCAM-1 expression in the submucosa of TNF-alpha-treated colon. Spatial heterogeneity of MAdCAM-1 and VCAM-1 activation following TNF-alpha challenge may promote specific T lymphocyte recruitment in the inflamed colonic mucosa.     

Cultivation of microalgae in the solution from the desalting process of soy sauce waste treatment and utilization of the algal biomass for ethanol fermentation

The growth of four marine microalgae was examined in a medium prepared from the effluent from the desalting process of soy sauce waste. A strain of Dunaliella that showed abundant growth on soy sauce waste extract was selected, and optimum cultivation conditions were determined. The algal cells produced were disrupted, and saccharified with glucoamylase. The saccharified solution was fermented using Saccharomyces cerevisiae IAM 4140. Stoichiometric study revealed that 11mg of ethanol was produced from 1g (dry cell weight) of Dunaliella cells. This work indicates a new method for removing waste products of the soy sauce production industry.