Human RNaseP RNA and nucleolar 7-2 RNA share conserved ‘To’ antigen-binding domains

RNase P in both prokaryotes and eukaryotes is a ribonucleoprotein that cleaves tRNA precursors to generate the 5 termini of the mature tRNAs. Many patients with autoimmune diseases produce antibodies against a 40 kDa protein (designatedTo orTh antigen) which is an integral component of eukaryotic RNaseP as well as nucleolar 7-2 RNP which is identical to the mitochondrial RNA processing (MRP) RNP. Interestingly, theTo antigen found in human cells and the C5 protein, the only protein component ofE. coli RNaseP, are antigenically related. In this study, we show that a 56 nucleotide-long sequence, corresponding to nucleotides 20–75 near the 5 end of human RNaseP RNA, is sufficient to bind theTo antigen. We previously showed that the humanTo antigen binds to a short distinct structural domain near the 5 end of human 7-2/MRP RNA. There is no obvious primary sequence homology between theTo antigen binding sites in RNaseP RNA and 7-2/MRP RNA; however, these sequences are capable of assuming a similar secondary structure which corresponds to the recently proposed cage structure for RNaseP RNAs and 7-2/MRP RNA (Forster and Altman (1989) Cell 62: 407–409). These data are supportive of the idea that these two RNAs may have evolved from a common progenitor molecule.     

Effects of irradiation on neuropeptide expression in rat salivary gland and spinal cord

Summary It is well-known that a large number of factors can influence the expression of neuropeptides in the nervous system. In the present study, the effects of unilateral and bilateral irradiation to the rat head and neck on the expression of neuropeptides in the innervation of the submandibular gland and in the ganglionic cells of the submandibular ganglion was examined ten days and six months after treatment. Antisera directed against enkephalin and bombesin and immunohistochemical methods were used. The effects of bilateral irradiation on the staining pattern of various neuropeptides in the cervical spinal cord were also studied. In the submandibular gland and in the submandibular ganglionic cells, there was a markedly increased neuropeptide expression ten days after bilateral treatment, as seen after staining with both antisera used, while no changes occurred after unilateral treatment. Six months after treatment, the pattern of neuropeptide expression in the submandibular gland/ganglion corresponded to that seen in controls. Irradiation did not lead to any changes in the staining pattern of neuropeptides in the spinal cord. The observations show that there is a great complexity in the susceptibility of nervous tissues to radiotherapy with respect to influences on the expression of neuropeptides.     

Hypervariable region IV of Salmonella gene fliCd encodes a dominant surface epitope and a stabilizing factor for functional flagella.

To identify the major antigenic determinant of native Salmonella flagella of antigenic type d, we constructed a series of mutated fliCd genes with deletions and amino acid alterations in hypervariable region IV and in region of putative epitopes as suggested by epitope mapping with synthetic octameric peptides (T.M. Joys and F. Schödel, Infect. Immun. 59:3330-3332, 1991). The expressed product of most of the mutant genes, with deletions of up to 92 amino acids in region IV, assembled into functional flagella and conferred motility on flagellin-deficient hosts. Serological analysis of these flagella with different anti-d antibodies revealed that the peptide sequence centered at amino acids 229 to 230 of flagellin was a dominant B-cell epitope at the surface of d flagella, because replacement of these two amino acids alone or together with their flanking sequence by a tripeptide specified by a linker sequence eliminated most reactivity with antisera against wild-type d flagella as tested by enzyme-linked immunosorbent assay or by Western immunoblot. Functional analysis of the mutated flagellin genes with or without an insert suggested that amino acids 180 to 214 in the 5' part of hypervariable region IV (residues 181 to 307 of the total of 505) is important to the function of flagella. The hybrid proteins formed by insertion of peptide sequence pre-S1 12-47 of hepatitis B virus surface antigen into the deleted flagellins assembled into functional flagella, and antibody to the pre-S1 sequence was detected after immunization of mice with the hybrid protein. This suggests that such mutant flagellins containing heterologous epitopes have potential as vaccines.     

ISOLATION AND CHARACTERIZATION OF PHASE-SPECIFIC COMPLEMENTARY DNAs FROM SPOROPHYTES AND GAMETOPHYTES OF PORPHYRA PURPUREA (RHODOPHYTA) USING SUBTRACTED COMPLEMENTARY DNA LIBRARIES1

The red alga Porphyra purpurea (Roth) C. Agardh has a life cycle that alternates between shell-boring, filamentous sporophytes and free-living, foliose gametophytes. The significant morphological differences between these two phases suggest that many genes should be developmentally regulated and expressed in a phase-specific manner. In this study, we prepared and screened subtracted complementary DNA (cDNA) libraries specific for the sporophyte and gametophyte of P. purpurea. This involved the construction of cDNA libraries from each phase, followed by the removal of common clones through subtractive hybridization. Sampling of the subtracted libraries indicated that 8–10% of the recombinant colonies in each library were specific for the appropriate phase. Of 20 putative phase-specific cDNAs selected from each subtracted library, eight unique clones were obtained for the sporophyte and seven for the gametophyte. After confirming their phase-specificities by hybridization to gametophyte and sporophyte messenger RNA, these 15 phase-specific cDNAs were sequenced, and the deduced amino acid sequences were used to search protein databanks. Two proteins encoded by the sporophyte-specific cDNAs and two by the gametophyte-specific cDNAs were identified by their similarity to databank entries.     

Intracerebroventricular Administration of AT1 Receptor Antisense Oligonucleotides Inhibits the Behavioral Actions of Angiotensin II

Abstract: Antisense Oligonucleotides were developed to study the expression and function of angiotensin type 1 (AT₁) receptors in cultured cells and brain. In both liver epithelial WB and neuro-blastoma N1E-115 cells AT₁ antisense oligomers substantially decreased AT₁ receptor density, whereas angiotensin type 2 (AT₂) receptors remained unchanged. Similarly, repeated intracerebroventricular injections of AT₁ antisense oligomers in rats decreased AT₁ receptor density in hypothalamic-thalamic-septal tissue, and AT₂ receptors were unaffected. Intracerebroventricular antisense oligomers also attenuated drinking elicited by intra-cerebroventricular angiotensin II but not the cholinomimetic carbachol. Collectively, these results demonstrate that antisense Oligonucleotides attenuate angiotensin receptor expression and function in behaving animals.     

Protein synthesis is not required for the inhibitory effect of selenite on cell colony formation and RNA synthesis

Selenite has been shown to undergo intracellular metabolism that results in its conversion to other low molecular weight Secontaining species and also to its incorporation into a selenocysteine residue in selenoprotein. In order to investigate whether the incorporation into protein is required for the cytotoxic effects of selenite, we have examined whether inhibition of protein synthesis prevents the inhibitory effect of selenite on the ability of cells to form colonies or to synthesize RNA. We have found that treatment of HeLa cells with cycloheximide inhibited protein synthesis by >90% but had no effect on the inhibitory effect of selenite on cell colony formation or RNA synthesis. Since protein synthesis is not necessary for these cytotoxic effects of selenite they are unlikely to result from an increase in the synthesis of selenoproteins.     

Reconstitution of iron-sulfur center B of Photosystem I damaged by mercuric chloride

Incubation of thylakoid membranes from spinach with low concentrations of mercuric chloride induces the loss of one of the iron-sulfur centers, F_B, in Photosystem I (PS I) and inhibits the electron transfer from PS I to the soluble electron carrier, ferredoxin. Reconstitution of this damaged iron-sulfur center has been carried out by incubating treated thylakoid membranes with exogenous FeCl₃ and Na₂S in the presence of-mercaptoethanol under anaerobic conditions. Low temperature EPR measurements indicate that center F_B is largely restored. Kinetic experiments show that the restored F_B can be photoreduced from P700. However, these reconstituted thylakoid membranes are still incompetent in the photoreduction of ferredoxin and NADP⁺, even though ferredoxin binding to the modified membranes was not impaired, indicating additional changes in the structure of the PS I complex must have occurred.     

Leaf Developmental Age Controls Expression of Genes Encoding Enzymes of Chlorophyll and Heme Biosynthesis in Pea (Pisum sativum L.)

The effects of leaf developmental age on the expression of three nuclear gene families in pea (Pisum sativum L.) coding for enzymes of chlorophyll and heme biosynthesis have been examined. The steady-state levels of mRNAs encoding aminolevulinic acid (ALA) dehydratase, porphobilinogen (PBG) deaminase, and NADPH:protochlorophyllide reductase were measured by RNA gel blot and quantitative slot-blot analyses in the foliar leaves of embryos that had imbibed for 12 to 18 h and leaves of developing seedlings grown either in total darkness or under continuous white light for up to 14 d after imbibition. Both ALA dehydratase and PBG deaminase mRNAs were detectable in embryonic leaves, whereas mRNA encoding the NADPH:protochlorophyllide reductase was not observed at this early developmental stage. All three gene products were found to increase to approximately the same extent in the primary leaves of pea seedlings during the first 6 to 8 d after imbibition (postgermination) regardless of whether the plants were grown in darkness or under continuous white-light illumination. In the leaves of dark-grown seedlings, the highest levels of message accumulation were observed at approximately 8 to 10 d postgermination, and, thereafter, a steady decline in mRNA levels was observed. In the leaves of light-grown seedlings, steady-state levels of mRNA encoding the three chlorophyll biosynthetic enzymes were inversely correlated with leaf age, with youngest, rapidly expanding leaves containing the highest message levels. A corresponding increase in the three enzyme protein levels was also found during the early stages of development in the light or darkness; however, maximal accumulation of protein was delayed relative to peak levels of mRNA accumulation. We also found that although protochlorophyllide was detectable in the leaves immediately after imbibition, the time course of accumulation of the phototransformable form of the molecule coincided with NADPH:protochlorophyllide reductase expression. In studies in which dark-grown seedlings of various ages were subsequently transferred to light for 24 and 48 h, the effect of light on changes in steady-state mRNA levels was found to be more pronounced at later developmental stages. These results suggest that the expression of these three genes and likely those genes encoding other chlorophyll biosynthetic pathway enzymes are under the control of a common regulatory mechanism. Furthermore, it appears that not light, but rather as yet unidentified endogenous factors, are the primary regulatory factors controlling gene expression early in leaf development.     

The last 140 ka in the Afro-Asian arid/semi-arid transitional zone

During the last ten years, a great number of geological or other proxy records provided radiocarbon, U/Th or TL dated information on the past climatic oscillations in the arid/semi-arid zones extending between the Atlantic and the Pacific. Comparisons and a synthetic view of these results can now be attempted for the last 140 ka, and compared with global changes, as registered in oceanic or ice cores, and with palaeomonsson models: for this purpose, wide spatial and time scales have to be used. On the whole, arid/humid alternations roughly fit global changes, cold phases corresponding to an extension of the arid areas to the South, warm phases corresponding to the shrinking of the same to the North.

The last interglacial is associated with an increase of precipitation throughout the area considered. Isotopic stage 4 brings no evidence whatsoever of humid conditions. Two wetter episodes are registrated during stage 3. A major rainfall decrease is everywhere associated with the Last Glacial Maximum (21-15 ka in most regions), the arid or semi-arid zones extending several hundred kilometers southwards, relative to the present-day pattern. The two abrupt deglaciation steps and the Younger Dryas are recorded in all of the most sensitive regions, at the margins of the present-day monsoonal range. During the Holocene, the precipitation increases everywhere (by 100–400 mn, relative to the present-day values), the optimum being at 8.5-6.5 ka. A climatic deterioration follows with an irregular pattern of dry/wet episodes, according to the different geographic conditions. The humid phase terminates at 3.5-3 ka in the whole transitional zone.

The major southward shift of the monsoonal precipitation range since its optimum, some 8000 years ago, fits the astronomical neoglacial trend and may possibly be correlated with past stage 5d, although its rapidity and spatial importance may just reflect one of the sharp successive cold/warm variations registered by GRIP during the whole stage 5.