The transport of class I major histocompatibility complex antigens is determined by sequences in the α1 and α2 protein domains

The transport of human-mouse hybrid class I histocompatibility antigens has been studied in a mutant human cell line, 174 × CEM.T2 (T2). T2, a somatic cell hybrid of human B- and T-lymphoblastoid cell lines (B-LCL and T-LCL, respectively), synthesizes HLA-A2 and HLA-B5 glycoproteins, but expresses only low levels of A2 and undetectable levels of B5 at the cell surface. We have previously shown that the products of human class I genes introduced into T2 by transfection behave like the endogenous HLA-B5 glycoproteins, while the products of mouse class I alleles similarly introduced are transported normally to the cell surface. We have now determined that the surface expression of class I glycoproteins in T2 depends on the origin of the ₁ and ₂ domains. Human (HLA-B7) and mouse (H-2D ^p ) hybrid class I genes, encoding the leader, ₁, and ₂ sequences of one species fused to the ₃, transmembrane, and cytoplasmic domains of the other, were transfected into T2. Normal surface expression of the hybrid class I molecule was observed in T2 only when the leader, ₁, and ₂-encoding exons were derived from the mouse gene. The reciprocal construct, encoding human leader, ₁, and ₂ domains fused to the mouse ₃, transmembrane, and cytoplasmic regions, resulted in biosynthesis of a hybrid glycoprotein which was not transported to the cell surface. The products of both constructs were expressed normally in control cells. The effects of glycosylation on class I antigen transport were also studied using mutant class I constructs with altered glycosylation sites. Two mutant B7 genes encoding either an extra glycosylation site at position 176 or no glycosylation sites were transfected into T2. These mutant products were expressed at the cell surface in control cells, but were synthesized and not surface-expressed in T2. These data demonstrate that the HLA/H-2 transport dichotomy in T2 is a function of the origin of the ₁ and/or ₂ domains of the class I glycoprotein, and is not a reflection of glycosylation differences between the human and mouse molecules. Offprint requests to: P. Cresswell.     

Development of Photosystem II in dark grown Chlamydomonas reinhardtii. A light-dependent conversion of PS IIβ, QB-nonreducing centers to the PS IIα, QB-reducing form

The green alga Chlamydomonas reinhardtii is a facultative heterotroph and, when cultured in the presence of acetate, will synthesize chlorophyll (Chl) and photosystem (PS) components in the dark. Analysis of the thylakoid membrane composition and function in dark grown C. reinhardtii revealed that photochemically competent PS II complexes were synthesized and assembled in the thylakoid membrane. These PS II centers were impaired in the electron-transport reaction from the primary-quinone electron acceptor, Q_A, to the secondary-quinone electron acceptor, Q_B (Q_B-nonreducing centers). Both complements of the PS II Chl a–b light harvesting antenna (LHC II-inner and LHC II-peripheral) were synthesized and assembled in the thylakoid membrane of dark grown C. reinhardtii cells. However, the LHC II-peripheral was energetically uncoupled from the PS II reaction center. Thus, PS II units in dark grown cells had a -type Chl antenna size with only 130 Chl (a and b) molecules (by definition, PS II units lack LHC II-peripheral). Illumination of dark grown C. reinhardtii caused pronounced changes in the organization and function of PS II. With a half-time of about 30 min, PS II centers were converted froma Q_B-nonreducing form in the dark, to a Q_B-reducing form in the light. Concomitant with this change, PS II units were energetically coupled with the LHC II-peripheral complement in the thylakoid membrane and were converted to a PS II form. The functional antenna of the latter contained more than 250 Chl(a+b) molecules. The results are discussed in terms of a light-dependent activation of the Q_A-Q_B electron-transfer reaction which is followed by association of the PS II unit with a LHC II-peripheral antenna and by inclusion of the mature form of PS II (PS II) in the membrane of the grana partition region.Abbreviations Chl chlorophyll - PS photosystem - Q_A primary quinone electron acceptor of PS II - Q_B secondary quinone electron acceptor of PS II - LHC light harvesting complex - F₀ non-variable fluorescence yield - F_plf intermediate fluorescence yield plateau leyel - F_max maximum fluorescence yield - F_i initial fluorescence yield increase from F₀ to F_pl (F_pl–F₀) - F_v total variable fluorescence yield (F_m–F0) - DCMU dichlorophenyl-dimethylurea     

Are natural lipids UV-screening agents?

Summary Isolated lipids from Deinococcus radiodurans were reconstituted at final concentrations of 1 mg/ml into dioleoyl phosphatidyl choline (DOPC) vesicles and assayed for the ability to protect cells of Escherichia coli against killing by UV light (254 nm). Values of D₃₇ (UV dose required to reduce the number of surviving cells to 37% of the original number) were calculated from killing curves. E. coli was afforded the greatest protection with an individual lipid, identified as vitamin MK8 (D₃₇=310 J//m², compared to D₃₇=67 J/m² for E. coli irradiated in the presence of DOPC alone). Liposome-mediated protection was dependent on UV₂₅₄ absorbance and not on turbidity-related light-scattering. BOth vitamin MK8 from D. radiodurans and vitamin K₁, which is available commercially, showed a similar degree of UV₂₅₄-protection for E. coli. The UV-protective properties of vitamin K₁ were also investigated on mammalian cells in comparison with other natural lipids and known sunscreens. Survival curves were obtained for mouse fibroblast (L) cells irradiated at UV₂₅₄ in the absence or presence of DOPC liposomes into which were incorporated various natural lipids or standard sunscreen ingredients, all at final concentrations of 1 mg/ml. Experimentally determined values of D₃₇ were as follows: Vitamin K₁, 73 J/m²; \-carotene, 44 J/m₂; -tocopherol, 20 J/m₂; sulisobenzone, 156 J/m²; p-aminobenzoic acid (PABA), 113 J/m²; benzophenone, 80 J/m²; oxybenzone, 61 J/m² and DOPC alone. 23 J/m². Vitamin K₁, the most protective lipid tested, was also compared with PABA and oxybenzone (all at concentrations of 20 mg/ml; applied topicall) for its ability to protec Skh-hairless mice from UV₂₅₄-induced erythema, yielding a UV₂₅₄ protection factor of 3.5. In addition, vitamin K₁ (at 100 mg/ml) was able to provide hairless mice with a small degree of UVB protection, as indicated by an experimentally determined Solar Protection Factor of 1.5–2.0. Although it is concluded that vitamin K is not likely to account for the extraordinarily high degree of UV-resistance of D. radiodurans, vitamin K does show characteristics worthy of its consideration as a UV-screening agent. Offprint requests to: R. Anderson     

Multiple fruiting of the edible mushroomCoprinus cinereus from explants of the stipe

Summary Carpophores were developed on defined medium from explants of stipes produced in the dark using an expansionless mutant (no. 190) ofCoprinus cinereus. Explants taken from stipes within 24 h of formation developed multiple fruit within 3–12 days without formation of mycelia. Fruiting levels were affected by stipe age (24 h), medium composition, explant size, and polarity of the explant on fruiting medium. This method offers a new tool for developmental studies and may also be of use to commercial mushroom growers.     

Expression of c-Fos in subcutaneous adipose tissue of the fetal pig

Calcium oxalate crystal growth and aggregation leads to the formation of renal calculi. It is known to be inhibited by several compounds both in vitro and in vivo conditions. The present study highlights the inhibitory potential of sodium pentosan polysulphate (SPP), a semi-synthetic glycosaminoglycan (GAG) on calcium oxalate crystal growth in vitro. Its efficacy was compared with those of known inhibitors like pyrophosphate, heparin and chondroitin-4-sulphate. Of the above compounds pyrophosphate was found to be the most potent inhibitor. Among the GAGs, SPP exhibited 80% inhibitory activity as compared to heparin. A lesser degree of inhibition was observed with chondroitin-4-sulphate.     

Characterization of two class II chitinase genes from peanut and expression studies in transgenic tobacco plants

Two different genes encoding class II chitinases from peanut (Arachis hypogaea L. cv. NC4), A.h.Chi2;1 and A.h.Chi2;2, have been cloned. In peanut cell suspension cultures, mRNA levels of A.h.Chi2;2 increased after ethylene or salicylate treatment and in the presence of conidia from Botrytis cinerea. The second gene, A.h.Chi2;1, was only expressed after treatment with the fungal spores. Transgenic tobacco plants containing the complete peanut A.h.Chi2;1 gene exhibited essentially the same expression pattern in leaves as observed in peanut cell cultures. Expression characteristics of transgenic tobacco carrying a promoter-GUS fusion of A.h.Chi2;1 are described.     

Feed-forward activation in a theoretical first-order biochemical pathway which contains an anticipatory model

This paper explores the consequences of the theoretical forward activation enzymatic pathway A ₀ A ₁ A ₂ A ₃ where E ₁ convents A ₀ to A ₁, E ₂ converts A ₁ to A ₂ and E ₃ converts A ₂ to A ₃. A ₀, which is environmentally determined, also serves to activate (or modulate) the activity of E ₃ in such a way as to keep the concentration of A ₂ ([A ₂]) constant at a particular set-point value. For mathematical simplicity, first order rate kinetics are used where k ₁, k ₂ and k ₃ are the rate constants for E ₁, E ₂, and E ₃ respectively. It is shown that if k ₃ is modulated appropriately so as keep [A ₂] at the setpoint value with a changing upstream [A ₀], then the modulation of k ₃ must be anticipatory of the dynamics of the biochemical pathway. In other words, the rate of change of k ₃, will be a function of [A ₀], k ₁, k ₂, k ₃, and the set-point value of [A ₂]. If the modulation of k ₃ does not perfectly model (anticipate) the reaction pathway of which it is a part, then the actual [A ₂] will deviate from the set-point value. This type of anticipatory feed-forward activation may represent an important aspect of biological organization.