Seasonal Changes in Nitrogen Fractions of Coptis japonica, a Temperate Forest Evergreen Chamaephyte, and their Ecological Implications

Seasonal changes in several forms of nitrogen were investigatedin Coptis japonica, an evergreen rosette hemicryptophyte intemperate deciduous forest. The concentration of total nitrogenin rhizomes and roots decreased during the period of new shootgrowth from winter to spring. In the rhizomes, total solubleprotein stored by early summer decreased gradually until winter,coupled with an increase in free amino acids. Nitrogen was largelystored in free amino acids in the roots, especially during summer.The total soluble protein in current-year leaves decreased fromspring to summer and then increased during winter. The seasonalchanges in nitrogen components were coincident with the changein light-saturated photosynthetic rates recorded in a previousstudy. The ratio of total soluble protein to total nitrogendecreased from spring to summer and then increased from latesummer to winter in the current-year leaves. In contrast, chlorophyllcontent and the ratio of chlorophyll to total nitrogen werehigher in summer than in other seasons. The results indicatethat nitrogen was used in a manner that better utilizes thevery weak light in summer and the higher light intensities inother seasons. The major component of the free amino acid poolwas asparagine, in every organ throughout the season, exceptfor the senescent leaves. Since asparagine has a high N:C ratio,we suppose that the asparagine-dominated amino acid pool isadvantageous in the carbon-limited environment of the forestfloor.Copyright 1994, 1999 Academic Press Free amino acid composition, total nitrogen, total soluble protein, photosynthesis, evergreen hemicryptophyte     

Adoptive transfer of H-2-incompatible lymphokine-activated killer (LAK) cells: an approach for successful cancer immunotherapy free from graft-versus-host disease (GVHD) using murine models

We investigated whether the adoptive transfer of H-2-incompatible lymphokine-activated killer (LAK) cells would efficiently demonstrate antitumor activity without damaging the normal host cells. Allogeneic LAK cells (5 X 10(7] did not cause graft-versus-host disease (GVHD) in irradiated recipients, whereas more than half of the mice transferred with the same dose of fresh allogeneic spleen cells developed GVHD. Repeated transfer (three times at 4-day intervals, 1.2 X 10(8) cells/mouse) did not result in GVHD. Graft-versus-host reaction (GVHR), which is detectable by spleen enlargement of recipients transferred with allogeneic lymphoid cells was also absent in LAK cell-transferred mice of all strain combinations tested. Host immune responses were not affected in these mice. Therefore, it is feasible to transfer allogeneic LAK cells. With the antitumor efficacy of allogeneic LAK cells, they preferentially lysed allogeneic tumor targets. Adoptive transfer of the allogeneic LAK cells led to a significant decrease in the lung-colonizing foci of intravenously inoculated B16 melanoma cells. Allogeneic LAK cells and syngeneic ones were equally active, in vivo. The use of allogeneic LAK cells may prove to be a valuable method for effective clinical antitumor immunotherapy.     

Legionella hijacks the host Golgi-to-ER retrograde pathway for the association of Legionella-containing vacuole with the ER

Legionella pneumophila (L. pneumophila) is a gram-negative bacterium that replicates in a compartment that resembles the host endoplasmic reticulum (ER). To create its replicative niche, L. pneumophila manipulates host membrane traffic and fusion machineries. Bacterial proteins called Legionella effectors are translocated into the host cytosol and play a crucial role in these processes. In an early stage of infection, Legionella subverts ER-derived vesicles (ERDVs) by manipulating GTPase Rab1 to facilitate remodeling of the Legionella-containing vacuole (LCV). Subsequently, the LCV associates with the ER in a mechanism that remains elusive. In this study, we show that L. pneumophila recruits GTPases Rab33B and Rab6A, which regulate vesicle trafficking from the Golgi to the ER, to the LCV to promote the association of LCV with the ER. We found that recruitment of Rab6A to the LCV depends on Rab33B. Legionella effector SidE family proteins, which phosphoribosyl-ubiquitinate Rab33B, were found to be necessary for the recruitment of Rab33B to the LCV. Immunoprecipitation experiments revealed that L. pneumophila facilitates the interaction of Rab6 with ER-resident SNAREs comprising syntaxin 18, p31, and BNIP1, but not tethering factors including NAG, RINT-1, and ZW10, which are normally required for syntaxin 18-mediated fusion of Golgi-derived vesicles with the ER. Our results identified a Rab33B-Rab6A cascade on the LCV and the interaction of Rab6 with ER-resident SNARE proteins for the association of LCV with the ER and disclosed the unidentified physiological role of SidE family proteins.     

Raman and infrared spectra of toxin gamma from the venom of the scorpion Tityus serrulatus

Toxin gamma is a basic, low-molecular-weight, neurotoxic protein, isolated from the venom of the Brazilian scorpion, Tityus serrulatus. Raman spectra (400-1800 cm-1 region) of this toxin in both the lyophilized state and in 0.1 M acetate buffer (pH 4.5) and the infrared spectrum (700-4000 cm-1 region) of a solid film were investigated. From the vibrational spectra, it can be concluded that the polypeptide backbone of toxin gamma consists of a mixture of the different secondary structures, with predominance of beta-sheet, followed by unordered structure and alpha-helix, with some evidence of beta-turn structures. The four disulfide bridges assume the gauche-gauche-gauche conformation of the CCSSCC fragments. The intensity ratio of the doublet at 853 and 828 cm-1 suggests that four out of the five tyrosine residues are exposed. The three tryptophan residues are exposed on the surface, and the single methionine residue assume the gauche-gauche conformation. Toxin gamma retains full activity in the pH 4.5-7.5 range, but is almost completely inactivated at pH 11.5.     

Regulatory Factors of Lymphocyte-Lymphocyte Interaction: II. Characterization of Human Mitogenic Factor Derived from the Culture Supernatant of a Mouse-Human Hybridoma

Cell hybridization of phytohemagglutinin (PHA)-stimulated human peripheral blood lymphocytes (PBL) with murine lymphoma (EL-4) provided three hybridomas (MHH-16, MHH-20, and MHH-22) which spontaneously produced human mitogenic factor (MF). MHH-16 was serially subcloned by limiting dilution procedures, which resulted in maintaining two subclones producing human MF spontaneously for more than one year (PQL-3 and PQL-5 subcloned lines). Human MF (MHH-MF) derived from supernatants of PQL-5 line cultures had a molecular weight (m.w.) of about 26,000–30,000 daltons (the major peak) with a minor peak with an m.w. of 15,000 daltons on Sephadex G-100 chromatography, and at a high concentration of NaCl (1 m), the activity of the 26,000–30,000-m.w. fraction became weak and that of the 15,000-m.w. fraction became predominant. MHH-MF had an isoelectric point of pH 5.0–6.5. On DEAE-cellulose chromatography, MHH-MF was eluted at a fairly low salt concentration (sodium phosphate buffer 0.02 M, pH 8.0, NaCl 10 mm). After periodate treatment of this MHH-MF, the mitogenic activity almost disappeared. MHH-MF was relatively unstable to heating at 56 C for 20 min. In the presence of tunicamycin (0.3μg/ml), an inhibitor of N-linked glycosylation, the synthesized MHH-MF showed a decrease in m.w. as follows: the major peak shifted from 26,000–30,000 to 23,000 daltons and the minor peak from 15,000 to 10,000 daltons on Sephadex G-100 chromatography. In internal labeling experiments with [³H]leucine, the ³H-labeled MF was partially purified, with mitogenic activity as a guide. This ³H-labeled MHH-MF fraction could be absorbed by PHA blasts but not by normal PBL. On SDS-PAGE under reducing conditions, only the radioactive peak of the 15,000-dalton fraction was recovered. MHH-MF obtained from the hybridoma culture supernatants may be a dimer of the 15,000-dalton fraction and a glycoprotein.     

Distribution and Numbers of Pygmies in Central African Forests

Pygmy populations occupy a vast territory extending west-to-east along the central African belt from the Congo Basin to Lake Victoria. However, their numbers and actual distribution is not known precisely. Here, we undertake this task by using locational data and population sizes for an unprecedented number of known Pygmy camps and settlements (n = 654) in five of the nine countries where currently distributed. With these data we develop spatial distribution models based on the favourability function, which distinguish areas with favourable environmental conditions from those less suitable for Pygmy presence. Highly favourable areas were significantly explained by presence of tropical forests, and by lower human pressure variables. For documented Pygmy settlements, we use the relationship between observed population sizes and predicted favourability values to estimate the total Pygmy population throughout Central Africa. We estimate that around 920,000 Pygmies (over 60% in DRC) is possible within favourable forest areas in Central Africa. We argue that fragmentation of the existing Pygmy populations, alongside pressure from extractive industries and sometimes conflict with conservation areas, endanger their future. There is an urgent need to inform policies that can mitigate against future external threats to these indigenous peoples’ culture and lifestyles.     

Cutting edge: inhibition of experimental tumor metastasis by dendritic cells pulsed with alpha-galactosylceramide.

A unique lymphoid lineage, Valpha14 NKT cells, bearing an invariant Ag receptor encoded by Valpha14 and Jalpha281 gene segments, play crucial roles in various immune responses, including protective immunity against malignant tumors. A specific ligand of Valpha14 NKT cells is determined to be alpha-galactosylceramide (alpha-GalCer) which is presented by the CD1d molecule. Here, we report that dendritic cells (DCs) pulsed with alpha-GalCer effectively induce potent antitumor cytotoxic activity by specific activation of Valpha14 NKT cells, resulting in the inhibition of tumor metastasis in vivo. Moreover, a complete inhibition of B16 melanoma metastasis in the liver was observed when alpha-GalCer-pulsed DCs were injected even 7 days after transfer of tumor cells to syngeneic mice where small but multiple metastatic nodules were already formed. The potential utility of DCs pulsed with alpha-GalCer for tumor immunotherapy is discussed.