A possible role for the pcnB gene product of Escherichia coli in modulating RNA: RNA interactions.

Summary The sequence of the PcnB protein of Escherichia coli, a protein required for copy number maintenance of ColE1-related plasmids, was compared with the PIR sequence database. Strong local similarities to the sequence of the E. coli protein tRNA nucleotidyltransferase were found. Since a substrate of the latter protein, tRNA, structurally resembles the RNAs that control ColE1 copy number we believe that we may have identified a region in PcnB that interacts with these RNAs. Consistent with this idea is our observation that PcnB is required for the replication of R1, a plasmid whose replication is also regulated by a small RNA.     

Targeted snRNP depletion reveals an additional role for mammalian U1 snRNP in spliceosome assembly

HeLa cell nuclear splicing extracts have been prepared that are specifically and efficiently depleted of U1, U2, or U4/U6 snRNPs by antisense affinity chromatography using biotinylated 2'-OMe RNA oligonucleotides. Removal of each snRNP particle prevents pre-mRNA splicing but arrests spliceosome formation at different stages of assembly. Mixing extracts depleted for different snRNP particles restores formation of functional splicing complexes. Specific binding of factors to the 3' splice site region is still detected in snRNP-depleted extracts. Depletion of U1 snRNP impairs stable binding of U2 snRNP to the pre-mRNA branch site. This role of U1 snRNP in promoting stable preslicing complex formation is independent of the U1 snRNA-5' splice site interaction.     

SecA protein: Autoregulated initiator of secretory precursor protein translocation across theE. coli plasma membrane

Several classes ofsecA mutants have been isolated which reveal the essential role of this gene product forE. coli cell envelope protein secretion. SecA-dependent,in vitro protein translocation systems have been utilized to show that SecA is an essential, plasma membrane-associated, protein translocation factor, and that SecA's ATPase activity appears to play an essential but as yet undefined role in this process. Cell fractionation studies suggested that SecA protein is in a dynamic state within the cell, occurring in soluble, peripheral, and integral membraneous states. These data have been used to argue that SecA is likely to promote the initial insertion of secretory precursor proteins into the plasma membrane in a manner dependent on ATP hydrolysis. The protein secretion capability of the cell has been shown to translationally regulatesecA expression with SecA protein serving as an autogenous repressor, although the exact mechanism and purpose of this regulation need to be defined further.     

Acclimation of barley to changes in light intensity: photosynthetic electron transport activity and components

Barley seedlings (Hordeum vulgare L. Boone) were grown at 20°C with 16 h/8 h light/dark cycle of either high (H) intensity (500 mole m^-2 s^-1) or low (L) intensity (55 mole m^-2 s^-1) white light. Plants were transferred from high to low (H L) and low to high (L H) light intensity at various times from 4 to 8 d after leaf emergence from the soil. Primary leaves were harvested at the beginning of the photoperiod. Thylakoid membranes were isolated from 3 cm apical segments and assayed for photosynthetic electron transport, Photosystem II (PS II) atrazine-binding sites (Q_B), cytochrome f(Cytf) and the P-700 reaction center of Photosystem I (PS I). Whole chain, PS I and PS II electron transport activities were higher in H than in L controls. Q_B and Cytf were elevated in H plants compared with L plants. The acclimation of H L plants to low light occurred slowly over a period of 7 days and resulted in decreased whole chain and PS II electron transport with variable effects on PS I activity. The decrease in electron transport of H L plants was associated with a decrease in both Q_B and Cytf. In L H plants, acclimation to high light occurred slowly over a period of 7 days with increased whole chain, PS I and PS II activities. The increase in L H electron transport was associated with increased levels of Q_B and Cytf. In contrast to the light intensity effects on Q_B levels, the P-700 content was similar in both control and transferred plants. Therefore, PS II/PS I ratios were dependent on light environment.Abbreviations Asc ascorbate - BQ 2,5-dimethyl-p-benzoquinone - DBMIB 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone - DCIP 2,6-dichlorophenolindophenol - H control plants grown under high light intensity - H L plants transferred from high to low light intensity - L low control plants grown under low light intensity - L H plants transferred from low to high light intensity - MV methyl viologen - P-700 photoreaction center of Photosystem I - Q_B atrazine binding site - TMPD N,N,N,N-tetramethyl-p-phenylenediamine Cooperative investigations of the United States Department of Agriculture, Agricultural Research Service, and the North Carolina Agricultural Research Service, Raleigh, NC. Paper No. 11990 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, NC 27695-7643, USA.     

The Ultrastructure of the Spermatozoa of Squamata—I. Scincidae,Gekkonidae and Pygopodidae (Reptilia)

Abstract Squamate autapomorphies seen in sperm of the Scincidae (e.g. Ctenotus robustus, Carlia pectoralis, Cryptoblepharus virgatus, and Lampropholis delicata) are penetration of the fibrous sheath of the axoneme into the midpiece, and the paracrystalline subacrosomal cone. Sphenomorphus group spermatozoa (e.g. Ctenotus) and the Egernia group (Tiliqua) differ from the more derived Eugongylus group (C. virgatus, L. delicata and C. pectoralis) in that the acrosome is elongate and apically depressed; the perforatorium is strongly oblique; the midpiece is relatively short, with four dense ring structures in longitudinal succession; mitochondria are columnar; and enlarged peripheral fibres 3 and 8 do not show the gross anterior enlargement seen in Carlia and Lampropholis. Heteronotia binoei (Gekkonidae) sperm have no epinuclear electron-lucent region; nuclear shoulders are smooth, as in sphenomorph but not Eugongylus group skinks; mitochondria are columnar; unlike skinks, the median surfaces of the mitochondria are indented by triangular, sometimes longitudinally, interconnected dense bodies. In Lialis burtonis (Pygopodidae) sperm, the perforatorium extends virtually to the tip of the fore-shortened apically domed acrosome; nuclear shoulders are absent; the mitochondria alternate singly or in groups with one or more dense bodies which also form an interrupted collar around the distal centriole. Spermatozoal ultrastructure suggests that a common ancestry of snakes and pygopods deserves consideration.     

An Experimental Test of the Rhizopine Concept in Rhizobium meliloti

In some Rhizobium-legume symbioses, compounds known as rhizopines are synthesized by bacteroids and subsequently catabolized by free-living cells of the producing strain. It has been suggested than rhizopines act as proprietary growth substrates and enhance the competitive ability of the producing strain in its interactions with the diverse microbial community found within the rhizosphere. Wild-type, rhizopine-producing Rhizobium meliloti L5-30 and mutant L5-30 strains deficient for either rhizopine synthesis or catabolism were inoculated onto lucerne host plants in competition experiments. These experiments demonstrated that no apparent advantage resulted from the ability to synthesize a rhizopine, whereas the ability to catabolize rhizopine provided a clear advantage when an organism was in competition with a strain without this ability. The results suggest that when an organism is in competition with a catabolism-deficient mutant, the ability to catabolize rhizopine results in enhanced rates of nodulation. The results of the experiments were not consistent with the hypothesis that the sole role of rhizopines is to act as proprietary growth substrates for the free-living population of the producing strain.     

Detection of Vibrio vulnificus biotypes 1 and 2 in eels and oysters by PCR amplification.

DNA extraction procedures and PCR conditions to detect Vibrio vulnificus cells naturally occurring in oysters were developed. In addition, PCR amplification of V. vulnificus from oysters seeded with biotype 1 cells was demonstrated. By the methods described, V. vulnificus cells on a medium (colistin-polymyxin B-cellobiose agar) selective for this pathogen were detectable in oysters harvested in January and March, containing no culturable cells (< 67 CFU/g), as well as in oysters harvested in May and June, containing culturable cells. It was possible to complete DNA extraction, PCR, and gel electrophoresis within 10 h by using the protocol described for oysters. V. vulnificus biotype 2 cells were also detected in eel tissues that had been infected with this strain and subsequently preserved in formalin. The protocol used for detection of V. vulnificus cells in eels required less than 5 h to complete. Optimum MgCl2 concentrations for the PCR of V. vulnificus from oysters and eels were different, although the same primer pair was used for both. This is the first report on the detection of cells of V. vulnificus naturally present in shellfish and represents a potentially powerful method for monitoring this important human and eel pathogen.