Effect of organic and conventional farming on soil bacterial diversity of pecan tree (Carya illinoensis K. Kosh) orchard across two phenological stages

We described the bacterial diversity of walnut grove soils under organic and conventional farming. The bacterial communities of rhizospheric and nonrhizospheric soils of pecan tree (Carya illinoensis K. Koch) were compared considering two phenological stages (sprouting and ripening). Sixteen operational taxonomic units (OTUs) were identified significantly more abundant according to the plant development, only one according to the farming condition, and none according to the soil origin. The OTUs specificaly abundant according to plant development included Actinobateria (2) and Betaproteobacteria (1) related OTUs more abundant at the sprouting stage, while at the fruit ripening (FR) stage the more abundant OTUs were related to Actinobacteria (6), Alphaproteobacteria (6), and unclassified Bacteria (1). The Gaiellaceae OTU18 (Actinobacteria) was more abundant under conventional farming. Thus, our study revealed that the plant development stage was the main factor shaping the bacterial community structure, while less influence was noticed for the farming condition. The bacterial communities exhibited specific metabolic capacities, a large range of carbon sources being used at the FR stage. The identified OTUs specifically more abundant represent indicators providing useful information on soil condition, potential tools for the management of soil bacterial communities.     

Characterization of a centromere-linked recombination hot spot in Saccharomyces cerevisiae.

A 1.5-kilobase-pair SalI-HindIII (SH) restriction fragment from the region of Saccharomyces cerevisiae chromosome XIV immediately adjacent to the centromere appears to contain sequences that act as a hot spot for mitotic recombination. The presence of SH DNA on an autonomously replicating plasmid stimulates homologous genetic exchange between yeast genomic sequences and those present on the plasmid. In all recombinants characterized, exchange occurs in plasmid yeast sequences adjacent to rather than within the SH DNA. Hybridization analyses reveal that SH-containing plasmids are present in linear as well as circular form in S. cerevisiae and that linear forms are generated by cleavage at specific sites. Presumably, it is the linear form of the plasmid that is responsible for the stimulation of genetic exchange. Based on these observations, it is proposed that this DNA fragment contains a centromere-linked recombination hot spot and that SH-stimulated recombination occurs via a mechanism similar to double-strand-gap repair (J. W. Szostak, T. Orr-Weaver, J. Rothstein, and F. Stahl, Cell 33:25-35 1983).     

Structural organization and functional analysis of centromeric DNA in the fission yeast Schizosaccharomyces pombe.

Centromeric DNA in the fission yeast Schizosaccharomyces pombe was isolated by chromosome walking and by field inversion gel electrophoretic fractionation of large genomic DNA restriction fragments. The centromere regions of the three chromosomes were contained on three SalI fragments (120 kilobases [kb], chromosome III; 90 kb, chromosome II; and 50 kb, chromosome I). Each fragment contained several repetitive DNA sequences, including repeat K (6.4 kb), repeat L (6.0 kb), and repeat B, that occurred only in the three centromere regions. On chromosome II, these repeats were organized into a 35-kb inverted repeat that included one copy of K and L in each arm of the repeat. Site-directed integration of a plasmid containing the yeast LEU2 gene into K repeats at each of the centromeres or integration of an intact K repeat into a chromosome arm had no effect on mitotic or meiotic centromere function. The centromeric repeat sequences were not transcribed and possessed many of the properties of constitutive heterochromatin. Thus, S. pombe is an excellent model system for studies on the role of repetitive sequence elements in centromere function.     

An additional long-range interaction in human U1 snRNA.

We present evidence for the existence of an additional long-range interaction in vertebrate U1 snRNAs. By submitting human U1 snRNP, HeLa nuclear extracts, authentic human or X. laevis in vitro transcribed U1 snRNAs to RNase V1, a nuclease specific for double-stranded regions, cleavages occurred in the sequence psi psi ACC (positions 5-9) residing in the 5' terminal region of the RNA. The RNase V1 sensitive region is insensitive to single-stranded probes, something unexpected knowing that it was considered single-stranded in order to base-pair to pre-mRNA 5' splice site. We have identified the sequence GGUAG (positions 132-136) as the only possible 3' partner. Mutants, either abolishing or restoring the interaction between the partners, coupled to an RNase V1 assay, served to substantiate this base-pairing model. The presence of this additional helix, even detected in nuclear extracts under in vitro splicing conditions, implies that a conformational change must occur to release a free U1 snRNA 5' end.     

Identification and characterization of the centromere from chromosome XIV in Saccharomyces cerevisiae.

A functional centromere located on a small DNA restriction fragment from Saccharomyces cerevisiae was identified as CEN14 by integrating centromere-adjacent DNA plus the URA3 gene by homologous recombination into the yeast genome and then by localizing the URA3 gene to chromosome XIV by standard tetrad analysis. DNA sequence analysis revealed that CEN14 possesses sequences (elements I, II, and III) that are characteristic of other yeast centromeres. Mitotic and meiotic analyses indicated that the CEN14 function resides on a 259-base-pair (bp) RsaI-EcoRV restriction fragment, containing sequences that extend only 27 bp to the right of the element I to III region. In conjunction with previous findings on CEN3 and CEN11, these results indicate that the specific DNA sequences required in cis for yeast centromere function are contained within a region about 150 bp in length.     

A protein binds the selenocysteine insertion element in the 3'-UTR of mammalian selenoprotein mRNAs.

Several gene products are involved in co-translational insertion of selenocysteine by the tRNA(Sec). In addition, a stem-loop structure in the mRNAs coding for selenoproteins is essential to mediate the selection of the proper selenocysteine UGA codon. Interestingly, in eukaryotic selenoprotein mRNAs, this stem-loop structure, the selenocysteine insertion sequence (SECIS) element, resides in the 3'-untranslated region, far downstream of the UGA codon. In view of unravelling the underlying complex mechanism, we have attempted to detect RNA-binding proteins with specificity for the SECIS element. Using mobility shift assays, we could show that a protein, present in different types of mammalian cell extracts, possesses the capacity of binding the SECIS element of the selenoprotein glutathione peroxidase (GPx) mRNA. We have termed this protein SBP, for Secis Binding Protein. Competition experiments attested that the binding is highly specific and UV cross-linking indicated that the protein has an apparent molecular weight in the range of 60-65 kDa. Finally, some data suggest that the SECIS elements in the mRNAs of GPx and another selenoprotein, type I iodothyronine 5' deiodinase, recognize the same SBP protein. This constitutes the first report of the existence of a 3' UTR binding protein possibly involved in the eukaryotic selenocysteine insertion mechanism.     

Overexpression of the yeast MCK1 protein kinase suppresses conditional mutations in centromere-binding protein genes CBF2 and CBF5

We find that overexpression in yeast of the yeast MCK1 gene, which encodes a meiosis and centromere regulatory kinase, suppresses the temperature-sensitive phenotype of certain mutations in essential centromere binding protein genes CBF2 and CBF5. Since Mck1p is a known serine/threonine protein kinase, this suppression is postulated to be due to Mck1p-catalyzed in vivo phosphorylation of centromere binding proteins. Evidence in support of this model was provided by the finding that purified Mck1p phosphorylates in vitro the 110 kDa subunit (Cbf2p) of the multimeric centromere binding factor CBF3. This phosphorylation occurs on both serine and threonine residues in Cbf2p.