Storage of monokaryotic strains of Podospora anserina

Maintenance of Podospora anserina strains for experimental purposes is very time consuming (see Esser 1969 Neurospora Newsl. 15:27-31) and methods have been published that address this issue by freezing ascospores at -80 oC (Begel and Belcour 1991 Fungal Genet. Newsl. 38:67). Although the latter approach does reduce the amount of time required for yearly sexual crosses and ascospore isolation, it does not resolve the problem of the time required to rapidly generate monokaryotic hyphae, that are needed as a source for inoculum for many types of experiments

Substrate structural requirements of Schizosaccharomyces pombe RNase P

AbstractRNase P from Schizosaccharomyces pombe has been purified over 2000-fold. The apparent Km for two S. pombe tRNA precursors derived from the supS1 and sup3-e tRNASer genes is 20 nM; the apparent Vmax is 2.5 nM/min (supS1) and 1.1 nM/min (sup3-e). Processing studies with precursors of other mutants show that the structures of the acceptor stem and anticodon/intron loop of tRNA are crucial for S. pombe RNase P action

Georgia: Frozen Conflict and the Role of Displaced Persons

Though commonly overlooked, communities of displaced persons often play a complex and significant role in the emergence and perpetuation of ethnic conflict. This paper looks at the intersection of these themes in the conflict between the former Soviet Republic of Georgia and the separatist region of Abkhazia. In particular it looks at the nature of protracted or frozen conflict with particular attention to the role of the displaced community in the conflict\u27s entrenchment. Specifically, it seeks to answer the question: why do certain conflicts go unresolved for so long, and what role do refugees play in this resolution resistance? The paper is based on field research conducted in Georgia, including interviews with 45 Georgian internally displaced persons (IDPs) from Abkhazia. The results of the study suggest that various forces and motivations acting on the IDP community have the effect of entrenching it in the ambiguous state of neither returning to Abkhazia nor integrating into Georgian society that has become the status quo, and that this entrenchment plays a role in the factors that contribute to the frozen state of the conflict. In particular, the study suggests that power and identity play an unexpectedly large role in maintaining this population\u27s status quo

Group II intron mobility occurs by target DNA-primed reverse transcription

AbstractMobile group II introns encode reverse transcriptases and insert site specifically into intronless alleles (homing). Here, in vitro experiments show that homing of the yeast mtDNA group II intron a12 occurs by reverse transcription at a double-strand break in the recipient DNA. A site-specific endonuclease cleaves the antisense strand of recipient DNA at position +10 of exon 3 and the sense strand at the intron insertion site. Reverse transcription of al2-containing pre-mRNA is primed by the antisense strand cleaved in exon 3 and results in cotransfer of the intron and flanking exon sequences. Remarkably, the DNA endonuclease that initiates homing requires both the a12 reverse transcriptase protein and a12 RNA. Parallels in their reverse transcription mechanisms raise the possibility that mobile group II introns were ancestors of nuclear non-long terminal repeat retrotransposons and telomerases

A reverse transcriptase-related protein mediates phage resistance and polymerizes untemplated DNA in vitro

Reverse transcriptases (RTs) are RNA-dependent DNA polymerases that usually function in the replication of selfish DNAs such as retrotransposons and retroviruses. Here, we have biochemically characterized a RT-related protein, AbiK, which is required for abortive phage infection in the Grampositive bacterium Lactococcus lactis. In vitro, AbiK does not exhibit the properties expected for an RT, but polymerizes long DNAs of ‘random’ sequence, analogous to a terminal transferase. Moreover, the polymerized DNAs appear to be covalently attached to the AbiK protein, presumably because an amino acid serves as a primer. Mutagenesis experiments indicate that the polymerase activity resides in the RT motifs and is essential for phage resistance in vivo. These results establish a novel biochemical property and a non-replicative biological role for a polymerase

Database for bacterial group II introns

The Database for Bacterial Group II Introns (http://webapps2.ucalgary.ca/~groupii/index.html#) provides a catalogue of full-length, non-redundant group II introns present in bacterial DNA sequences in GenBank. The website is divided into three sections. The first section provides general information on group II intron properties, structures and classification. The second and main section lists information for individual introns, including insertion sites, DNA sequences, intron-encoded protein sequences and RNA secondary structure models. The final section provides tools for identification and analysis of intron sequences. These include a step-by-step guide to identify introns in genomic sequences, a local BLAST tool to identify closest intron relatives to a query sequence, and a boundary-finding tool that predicts 5′ and 3′ intron–exon junctions in an input DNA sequence. Finally, selected intron data can be downloaded in FASTA format. It is hoped that this database will be a useful resource not only to group II intron and RNA researchers, but also to microbiologists who encounter these unexpected introns in genomic sequences

A diversity of uncharacterized reverse transcriptases in bacteria

Retroelements are usually considered to be eukaryotic elements because of the large number and variety in eukaryotic genomes. By comparison, reverse transcriptases (RTs) are rare in bacteria, with only three characterized classes: retrons, group II introns and diversity-generating retroelements (DGRs). Here, we present the results of a bioinformatic survey that aims to define the landscape of RTs across eubacterial, archaeal and phage genomes. We identify and categorize 1021 RTs, of which the majority are group II introns (73%). Surprisingly, a plethora of novel RTs are found that do not belong to characterized classes. The RTs have 11 domain architectures and are classified into 20 groupings based on sequence similarity, phylogenetic analyses and open reading frame domain structures. Interestingly, group II introns are the only bacterial RTs to exhibit clear evidence for independent mobility, while five other groups have putative functions in defense against phage infection or promotion of phage infection. These examples suggest that additional beneficial functions will be discovered among uncharacterized RTs. The study lays the groundwork for experimental characterization of these highly diverse sequences and has implications for the evolution of retroelements

RNA Polymerase II CTD phosphatase Rtr1 fine-tunes transcription termination

RNA Polymerase II (RNAPII) transcription termination is regulated by the phosphorylation status of the C-terminal domain (CTD). The phosphatase Rtr1 has been shown to regulate serine 5 phosphorylation on the CTD; however, its role in the regulation of RNAPII termination has not been explored. As a consequence of RTR1 deletion, interactions within the termination machinery and between the termination machinery and RNAPII were altered as quantified by Disruption-Compensation (DisCo) network analysis. Of note, interactions between RNAPII and the cleavage factor IA (CF1A) subunit Pcf11 were reduced in rtr1Δ, whereas interactions with the CTD and RNA-binding termination factor Nrd1 were increased. Globally, rtr1Δ leads to decreases in numerous noncoding RNAs that are linked to the Nrd1, Nab3 and Sen1 (NNS) -dependent RNAPII termination pathway. Genome-wide analysis of RNAPII and Nrd1 occupancy suggests that loss of RTR1 leads to increased termination at noncoding genes. Additionally, premature RNAPII termination increases globally at protein-coding genes with a decrease in RNAPII occupancy occurring just after the peak of Nrd1 recruitment during early elongation. The effects of rtr1Δ on RNA expression levels were lost following deletion of the exosome subunit Rrp6, which works with the NNS complex to rapidly degrade a number of noncoding RNAs following termination. Overall, these data suggest that Rtr1 restricts the NNS-dependent termination pathway in WT cells to prevent premature termination of mRNAs and ncRNAs. Rtr1 facilitates low-level elongation of noncoding transcripts that impact RNAPII interference thereby shaping the transcriptome