Phosphorylation in vivo of a vaccinia-virus structural protein found associated with the ribosomes from infected cells.

When vaccinia-virus-infected cells were labeled with radioactive phosphate in the absence of viral gene expression an additional phosphoprotein, containing phosphoserine, was found specifically associated with the ribosomes. The phosphoprotein was removed from the ribosomes following a 0.5 M KCl washing or after EDTA treatment. This additional phosphoprotein was found in infected cells after either a long (3-4 h) or a short (30 min) labeling period; it was detected when the infected cells were incubated in the presence or absence of an inhibitor of RNA or protein synthesis. This phosphoprotein originated from the phosphorylation of vaccinia virion structural protein VP11b (Mr 11,000) at a specific site since only a single major phosphopeptide was obtained after trypsin digestion. This phosphoprotein was also present in purified vaccinia virions labeled with radioactive phosphate. VP11b protein was phosphorylated in vitro by the protein kinase associated with the cores. When the reaction was carried out at an alkaline pH the phosphorylation in vitro occurred at different sites in the protein; at neutral pH the phosphorylation of VP11b was more specific and, as judged by tryptic peptide analysis, occurred mainly at the same site as in the phosphorylation in vivo. A role for the involvement of phosphoprotein VP11b in the establishment of the shut off of host protein synthesis by vaccinia virus is suggested.     

Biosynthesis of Vitamins and Cofactors in Bacterium-Harbouring Trypanosomatids Depends on the Symbiotic Association as Revealed by Genomic Analyses

Some non-pathogenic trypanosomatids maintain a mutualistic relationship with a betaproteobacterium of the Alcaligenaceae family. Intensive nutritional exchanges have been reported between the two partners, indicating that these protozoa are excellent biological models to study metabolic co-evolution. We previously sequenced and herein investigate the entire genomes of five trypanosomatids which harbor a symbiotic bacterium (SHTs for Symbiont-Haboring Trypanosomatids) and the respective bacteria (TPEs for Trypanosomatid Proteobacterial Endosymbiont), as well as two trypanosomatids without symbionts (RTs for Regular Trypanosomatids), for the presence of genes of the classical pathways for vitamin biosynthesis. Our data show that genes for the biosynthetic pathways of thiamine, biotin, and nicotinic acid are absent from all trypanosomatid genomes. This is in agreement with the absolute growth requirement for these vitamins in all protozoa of the family. Also absent from the genomes of RTs are the genes for the synthesis of pantothenic acid, folic acid, riboflavin, and vitamin B_6. This is also in agreement with the available data showing that RTs are auxotrophic for these essential vitamins. On the other hand, SHTs are autotrophic for such vitamins. Indeed, all the genes of the corresponding biosynthetic pathways were identified, most of them in the symbiont genomes, while a few genes, mostly of eukaryotic origin, were found in the host genomes. The only exceptions to the latter are: the gene coding for the enzyme ketopantoate reductase (EC:1.1.1.169) which is related instead to the Firmicutes bacteria; and two other genes, one involved in the salvage pathway of pantothenic acid and the other in the synthesis of ubiquinone, that are related to Gammaproteobacteria. Their presence in trypanosomatids may result from lateral gene transfer. Taken together, our results reinforce the idea that the low nutritional requirement of SHTs is associated with the presence of the symbiotic bacterium, which contains most genes for vitamin production.     

Evolution under reversals: parsimony and conservation of common intervals

In comparative genomics, gene order data is often modeled as signed permutations. A classical problem for genome comparison is to detect common intervals in permutations, that is, genes that are colocalized in several species, indicating that they remained grouped during evolution. A second largely studied problem related to gene order is to compute a minimum scenario of reversals that transforms a signed permutation into another. Several studies began to mix the two problems and it was observed that their results are not always compatible: Often, parsimonious scenarios of reversals break common intervals. If a scenario does not break any common interval, it is called perfect. In two recent studies, Berard et al. defined a class of permutations for which building a perfect scenario of reversals sorting a permutation was achieved in polynomial time and stated as an open question whether it is possible to decide, given a permutation, if there exists a minimum scenario of reversals that is perfect. In this paper, we give a solution to this problem and prove that this widens the class of permutations addressed by the aforementioned studies. We implemented and tested this algorithm on gene order data of chromosomes from several mammal species and we compared it to other methods. The algorithm helps to choose among several possible scenarios of reversals and indicates that the minimum scenario of reversals is not always the most plausible     

Determinants of S. cerevisiae dynein localization and activation: implications for the mechanism of spindle positioning

BACKGROUND: During anaphase in budding yeast, dynein inserts the mitotic spindle across the neck between mother and daughter cells. The mechanism of dynein-dependent spindle positioning is thought to involve recruitment of dynein to the cell cortex followed by capture of astral microtubules (aMTs). RESULTS: We report the native-level localization of the dynein heavy chain and characterize the effects of mutations in dynein regulators on its intracellular distribution. Budding yeast dynein displays discontinuous localization along aMTs, with enrichment at the spindle pole body and aMT plus ends. Loss of Bik1p (CLIP-170), the cargo binding domain of Bik1p, or Pac1p (LIS1) resulted in diminished targeting of dynein to aMTs. By contrast, loss of dynactin or a mutation in the second P loop domain of dynein resulted in an accumulation of dynein on the plus ends of aMTs. Unexpectedly, loss of Num1p, a proposed dynein cortical anchor, also resulted in selective accumulation of dynein on the plus ends of anaphase aMTs. CONCLUSIONS: We propose that, rather than first being recruited to the cell cortex, dynein is delivered to the cortex on the plus ends of polymerizing aMTs. Dynein may then undergo Num1p-dependent activation and transfer to the region of cortical contact. Based on the similar effects of loss of Num1p and loss of dynactin on dynein localization, we suggest that Num1p might also enhance dynein motor activity or processivity, perhaps by clustering dynein motors.     

Current methods of gene prediction,their strengths and weaknesses

While the genomes of many organisms have been sequenced over the last few years, transforming such raw sequence data into knowledge remains a hard task. A great number of prediction programs have been developed that try to address one part of this problem, which consists of locating the genes along a genome. This paper reviews the existing approaches to predicting genes in eukaryotic genomes and underlines their intrinsic advantages and limitations. The main mathematical models and computational algorithms adopted are also briefly described and the resulting software classified according to both the method and the type of evidence used. Finally, the several difficulties and pitfalls encountered by the programs are detailed, showing that improvements are needed and that new directions must be considered.     

Algorithms for extracting structured motifs using a suffix tree with an application to promoter and regulatory site consensus identification.

This paper introduces two exact algorithms for extracting conserved structured motifs from a set of DNA sequences. Structured motifs may be described as an ordered collection of p > or = 1 "boxes" (each box corresponding to one part of the structured motif), p substitution rates (one for each box) and p - 1 intervals of distance (one for each pair of successive boxes in the collection). The contents of the boxes--that is, the motifs themselves--are unknown at the start of the algorithm. This is precisely what the algorithms are meant to find. A suffix tree is used for finding such motifs. The algorithms are efficient enough to be able to infer site consensi, such as, for instance, promoter sequences or regulatory sites, from a set of unaligned sequences corresponding to the noncoding regions upstream from all genes of a genome. In particular, both algorithms time complexity scales linearly with N2n where n is the average length of the sequences and N their number. An application to the identification of promoter and regulatory consensus sequences in bacterial genomes is shown.     

A stable microtubule array drives fission yeast polarity reestablishment upon quiescence exit

Cells perpetually face the decision to proliferate or to stay quiescent. Here we show that upon quiescence establishment, Schizosaccharomyces pombe cells drastically rearrange both their actin and microtubule (MT) cytoskeletons and lose their polarity. Indeed, while polarity markers are lost from cell extremities, actin patches and cables are reorganized into actin bodies, which are stable actin filament–containing structures. Astonishingly, MTs are also stabilized and rearranged into a novel antiparallel bundle associated with the spindle pole body, named Q-MT bundle. We have identified proteins involved in this process and propose a molecular model for Q-MT bundle formation. Finally and importantly, we reveal that Q-MT bundle elongation is involved in polarity reestablishment upon quiescence exit and thereby the efficient return to the proliferative state. Our work demonstrates that quiescent S. pombe cells assemble specific cytoskeleton structures that improve the swiftness of the transition back to proliferation.