Identification of a second 16-hydroxytabersonine-O-methyltransferase suggests an evolutionary relationship between alkaloid and flavonoid metabolisms in Catharanthus roseus

Protoplasma - The medicinal plant Catharanthus roseus biosynthesizes many important drugs for human health, including the anticancer monoterpene indole alkaloids (MIAs) vinblastine and vincristine....     

The anionic protease inhibitor BWI-1 from buckwheat seeds. Kinetic properties and possible biological role

Kinetic characteristics and effects on the growth of filamentous fungi of one of the main anionic protease inhibitors, BWI-1, isolated from buckwheat seeds, have been studied. The inhibition constants of bovine trypsin, chymotrypsin and cathepsin G from human granulocytes with BWI-1 were found to be 1.1, 67 and 200 n M , respectively. Analysis of the amino acid sequence of BWI-1 in the vicinity of the reactive site revealed its homology to the potato proteinase inhibitor I family. It is suggested that the inability of BWI-1 to bind elastase of human granulocytes is due to the basic nature of the amino acid residue (Arg) at the P_j position in its reactive site. It was demonstrated that BWI-1 was able to suppress the germination of the spores and the growth of the mycelium of two filamentous fungi.     

Some Peculiarities of the Dynamics of the Immunoglobulin M Structure

Abstract

The isotropic mobility of separate regions of the intact molecule of immunoglobulin M (IgM) and its Fab and (Fc)₅ fragments was studied using spin-labeling of carbohydrate (2,2,6,6-tetramethyl-4-aminopiperidine-l-oxyl) and peptide (2,2,5,5-tetramethyl-3-dichloro-triazinylaminopyrrolidine-l-oxyl) moieties.

The spin-labeled oligosaccharide groups (OGs) in the Fab region are shown to have much more amplitude of anisotropic motion than those in the (Fc)₅ region. The spin label in the latter is evidently attached in the Cμ3 domain to one of its OGs which is probably stabilized by ionic contacts between terminal N-acetylneuraminic acid residue and the peptide moiety of the IgM molecule.

When the amount of the glycosidase-cleaved carbohydrate does not exceed 10–15%, most OGs affected are of the Fab region. Upon profound splitting (≥50%) the OGs of the (Fc)₅region are also affected; that results evidently in loosening the ionic contacts between the shortened OGs and the peptide moiety of IgM, and consequently in increasing mobility of the former.

The structure of the (Fc)₅ region of IgM is labile; after detaching this moiety from the intact IgM molecule, its structure is stabilized, but one of its domains (Cμ3) becomes more mobile than it is in the intact IgM molecule; at the same time the amplitude of anisotropic motion of OG bound here is decreased. In the latter case, this decrease depends on the sequence of spin-labeling and fragmentation.

The most probable cause of stabilization of the (Fc)₅ fragment is the heating of IgM solution to 56°C during fragmentation with trypsin. At this temperature the τ value for the (Fc)₅ fragment is unusually low, equaling 23 ns.

The spin-labeling in the peptide part of IgM occurs mostly in the Fab region which is a rather rigid moiety as expected.     

Expression of the interleukin-7 receptor alpha chain (CD127) on virus-specific CD8+ T cells identifies functionally and phenotypically defined memory T cells during acute resolving hepatitis B virus infection

Virus-specific CD8+ T cells play a central role in the outcome of several viral infections, including hepatitis B virus (HBV) infection. A key feature of virus-specific CD8+ T cells is the development of memory. The mechanisms resulting in the establishment of T-cell memory are still only poorly understood. It has been suggested that T-cell memory may depend on the survival of virus-specific CD8+ T cells in the contraction phase. Indeed, a population of effector cells that express high levels of the interleukin-7 receptor alpha chain (CD127) as the precursors of memory CD8+ T cells has recently been identified in mice. However, very little information is currently available about the kinetics of CD127 expression in an acute resolving viral infection in humans and its association with disease pathogenesis, viral load, and functional and phenotypical T-cell characteristics. To address these important issues, we analyzed the HBV-specific CD8+ T-cell response longitudinally in a cohort of six patients with acute HBV infection who spontaneously cleared the virus. We observed the emergence of CD127 expression on antigen-specific CD8+ memory T cells during the course of infection. Importantly, the up-regulation of CD127 correlated phenotypically with a loss of CD38 and PD-1 expression and acquisition of CCR7 expression: functionally with an enhanced proliferative capacity and clinically with the decline in serum alanine aminotransferase levels and viral clearance. These results suggest that the expression of CD127 is a marker for the development of functionally and phenotypically defined antigen-specific CD8+ memory T cells in cleared human viral infections.     

Loss of Heterozygosity and Copy Number Alterations in Flow-Sorted Bulky Cervical Cancer

Treatment choices for cervical cancer are primarily based on clinical FIGO stage and the post-operative evaluation of prognostic parameters including tumor diameter, parametrial and lymph node involvement, vaso-invasion, infiltration depth, and histological type. The aim of this study was to evaluate genomic changes in bulky cervical tumors and their relation to clinical parameters, using single nucleotide polymorphism (SNP)-analysis.Flow-sorted tumor cells and patient-matched normal cells were extracted from 81 bulky cervical tumors. DNA-index (DI) measurement and whole genome SNP-analysis were performed. Data were analyzed to detect copy number alterations (CNA) and allelic balance state: balanced, imbalanced or pure LOH, and their relation to clinical parameters.The DI varied from 0.92–2.56. Pure LOH was found in ≥40% of samples on chromosome-arms 3p, 4p, 6p, 6q, and 11q, CN gains in >20% on 1q, 3q, 5p, 8q, and 20q, and losses on 2q, 3p, 4p, 11q, and 13q. Over 40% showed gain on 3q. The only significant differences were found between histological types (squamous, adeno and adenosquamous) in the lesser allele intensity ratio (LAIR) (p = 0.035) and in the CNA analysis (p = 0.011). More losses were found on chromosome-arm 2q (FDR = 0.004) in squamous tumors and more gains on 7p, 7q, and 9p in adenosquamous tumors (FDR = 0.006, FDR = 0.004, and FDR = 0.029).Whole genome analysis of bulky cervical cancer shows widespread changes in allelic balance and CN. The overall genetic changes and CNA on specific chromosome-arms differed between histological types. No relation was found with the clinical parameters that currently dictate treatment choice.     

Two stable electron-conformational states of photoactivated reaction center and their observation in primary donor recovery kinetics

We investigated the peculiarity of primary donor recovery kinetics in the reaction centers from the purple bacteriaRb. Sphaeroides at low levels of their cw photoactivation. A pronounced biphasity of the relaxation kinetics was found for the total light activating intensity <5×10¹² quanta·cm^-2·s^-1. The effect was attributed to strong dependence of an electron transfer rate constant for the reactionP ⁺Q _infA ^pup- Q_B P ⁺Q_AQ _infB ^sup- upon the RC conformational state controlled by the light. We showed the existence of two different electron-conformational states for the photoexcited RC. The first reveals itself at low intensity of cw photoactivation while the second becomes actual under the intensity >5×10¹² quanta · cm^-2 · s^-1.     

Changes in free IAA level in the leaves of short-and long-day tobacco during flowering and the effect of applied IAA on the transition to flowering

Changes in free IAA level were studied in the leaves of the central stem zone of short-day tobacco (Mcotianatabacum, cv. Maryland Mammoth) and long-day tobacco(Nicotiana silvestris) in inductive photoperiodic regime after 10, 20, 30 and 40 d, respectively. The leaves of SD tobacco Mammoth showed a high free IAA level in vegetative plants kept under long days but it significantly decreased (by ca. 50 %) after 10, 30 and 40 short days, respectively. After 20 short days the IAA level was as high as in the leaves of plants at the beginning of inductive treatment. The changes of freeIAA level in the leaves of LD tobacco N.silvestris were similar to those of SD Mammoth, but the IAA level in this species was significantly lower than that of Mammoth throughout the investigated period. Consequently, the changes observed in N. silvestris were much less pronounced. Plants of both tobacco species were fully induced to flowering by 30 inductive days and this was associated with differentiation of the flower organs. Application of 10 -4 M IAA during the last 10 d of the inductive treatment of 30 d significantly reduced flowering in SD tobacco Mammoth without changing the stem length and apex width. Apex length was slightly reduced. IAA application elicited almost no effect inN. silvestris. The results are discussed with respect to the possible role of IAA in flower induction in SD and LD plants.     

Diagnostic value of nucleolar organizer regions (AgNORs) in brush biopsies of suspicious lesions of the oral cavity.

OBJECTIVE: The aim of this retrospective study was to report on the diagnostic accuracy of AgNOR-analysis as an adjunctive diagnostic tool of conventional oral exfoliative cytology taken from suspicious lesions in our clinic. STUDY DESIGN: Cytological diagnoses obtained from brush biopsies of macroscopically suspicious lesions of the oral mucosa from 75 patients (final diagnoses: 53 histologically proven squamous cell carcinomas, 11 leukoplakias and other inflammatory oral lesions) and from 11 patients with normal mucosa as a negative control group were compared with histological and/or clinical follow-ups. Five smears were doubtful and seven suspicious for tumor cells in the cytologic report. Number of AgNOR's were counted in 100 squamous epithelial cell-nuclei per slide after silver-restaining. RESULTS: Sensitivity of our cytological diagnosis alone on oral smears for the detection of squamous carcinomas was 92.5%, specificity 100%, positive predictive value was 100% and negative 84.6%. The best cut-off value of the mean number of AgNOR dots per nucleus distinguishing benign from malignant cells was 4.8. The percentage of nuclei with more than three AgNORs had a cut-off level of 70%. Applying these methods to twelve doubtful or suspicious cytological diagnoses we were able to correctly establish the diagnosis of malignancy in ten cases of histologically proven cancers and to reveal benignity in two histologically proven cases. Thus we achieved a positive and negative predictive value of 100% each. CONCLUSIONS: Smears from brushings of visible oral lesions, if clinically considered as suspicious for cancer, are an easily practicable, non-invasive, painless, safe and accurate screening method for detection of oral cancerous lesions. We conclude that AgNOR-analysis may be a useful adjunct to other methods in routine cytological diagnosis of oral cancer that can help to solve cytologically suspicious or doubtful cases.