Detection of hilA gene sequences in serovars of Salmonella enterica subspecies enterica

hilA gene promoter, component of the Salmonella Pathogenicity Island 1, has been found in Salmonella serovar Typhimurium, being important for the regulation of type III secretion apparatus genes. We detected hilA gene sequences in Salmonella serovars Typhi, Enteritidis, Choleraesuis, Paratyphi A and B, and Pullorum, by polymerase chain reaction (PCR) and hybridization techniques. The primers to carry out PCR were designed according to hilA sequence. A low stringency hybridization with the probe pVV441 (hilA open-reading-frame plasmid) was carried out. To find hilA gene sequences in other Salmonella sp. suggest that these serovars could have similar sequences of this kind of virulence genes.     

Fission yeast enters a joyful new era

A report on the Second International Fission Yeast Meeting, Kyoto, Japan, 25-30 March 2002.     

Decrease of cytochrome c oxidase protein in heart mitochondria of copper-deficient rats

Copper deficiency has been reported to be associated withdecreased cytochrome c oxidase activity, whichin turn may be responsible for theobserved mitochondrial impairment and cardiac failure. We isolatedmito-chondriafrom hearts of copper-deficient rats: cytochrome c oxidase activity was found to be lowerthan incopper-adequate mitochondria. The residual activity paralleled coppercontent of mitochondria and also corresponded with the heme amount associated with cytochromeaa3. In fact, lower absorption in thea-band region of cytochrome aa3 was foundfor copper-deficient rat heart mitochondria. Gel electrophoresisof protein extractedfrom mitochondrial membranes allowed measurements of protein content of thecomplexes ofoxidative phosphorylation, revealing a lower content of complex IV protein incopper-deficientrat heart mitochondria. The alterations caused by copper deficiency appear to bespecific forcytochrome c oxidase. Changes were not observed for F 0 F 1 ATP synthase activity,for heme contents ofcytochrome c and b, and for protein contents of complexes I, III and V.The present study demonstrates that the alteration of cytochrome c oxidase activityobserved in copper deficiency is due to a diminishedcontent of assembled protein and that shortnessof copper impairs heme insertion into cytochrome c oxidase.     

Meiotic abnormalities in sugarcane (Saccharum spp.) and parental species: Evidence for peri- and paracentric inversions

The modern cultivars of sugarcane (Saccharum spp.) are highly polyploid and accumulate aneuploidies due to their history of domestication, genetic improvement and interspecific hybrid origin involving the domesticated sweet species Saccharum officinarum (‘noble cane’) and the wild Saccharum spontaneum, both with an evolutionary history of polyploidy. The first hybrids were backcrossed with S. officinarum, and selection from progenies in subsequent generations established the genetic basis of modern cultivars. Saccharum genome complexity has inspired several molecular studies that have elucidated aspects of sugarcane genome constitution, architecture and cytogenetics. Herein, we conducted a comparative analysis of the meiotic behaviour of representatives of the parentals S. officinarum and S. spontaneum, and the commercial variety, SP80-3280. S. officinarum, an octoploid species, exhibited regular meiotic behaviour. In contrast, S. spontaneum and SP80-3280 exhibited several abnormalities from metaphase I to the end of division. We reported and typified, for the first time, the occurrence of peri- and paracentric inversions. Using in-situ hybridisation techniques, we were able to determine how pairing association occurred at diakinesis, the origin of lagging chromosomes and, in particular, the mitotic chromosome composition of SP80-3280. Interestingly, S. spontaneum and recombinant chromosomes showed the most marked tendency to produce laggards in both divisions. Future attempts to advance knowledge on sugarcane genetics and genomics should take meiotic chromosome behaviour information into account.     

Exploitation of the ribosomal protein L10 R98S mutation to enhance recombinant protein production in mammalian cells

Mammalian cells are commonly used to produce recombinant protein therapeutics, but suffer from a high cost per mg of protein produced. There is therefore great interest in improving protein yields to reduce production cost. We present an entirely novel approach to reach this goal through direct engineering of the cellular translation machinery by introducing the R98S point mutation in the catalytically essential ribosomal protein L10 (RPL10‐R98S). Our data support that RPL10‐R98S enhances translation levels and fidelity and reduces proteasomal activity in lymphoid Ba/F3 and Jurkat cell models. In HEK293T cells cultured in chemically defined medium, knock‐in of RPL10‐R98S was associated with a 1.7‐ to 2.5‐fold increased production of four transiently expressed recombinant proteins and 1.7‐fold for one out of two stably expressed proteins. In CHO‐S cells, eGFP reached a 2‐fold increased expression under stable but not transient conditions, but there was no production benefit for monoclonal antibodies. The RPL10‐R98S associated production gain thus depends on culture conditions, cell type, and the nature of the expressed protein. Our study demonstrates the potential for using a ribosomal protein mutation for pharmaceutical protein production gains, and further research on how various factors influence RPL10‐R98S phenotypes can maximize its exploitability for the mammalian protein production industry.     

Reconstruction of the Doradinae (Siluriformes-Doradidae) ancestral diploid number and NOR pattern reveals new insights about the karyotypic diversification of the Neotropical thorny catfishes

Doradinae (Siluriformes: Doradidae) is the most species-rich subfamily among thorny catfishes, encompassing over 77 valid species, found mainly in Amazon and Platina hydrographic basins. Here, we analyzed seven Doradinae species using combined methods (e.g., cytogenetic tools and Mesquite ancestral reconstruction software) in order to scrutinize the processes that mediated the karyotype diversification in this subfamily. Our ancestral reconstruction recovered that 2n=58 chromosomes and simple nucleolar organizer regions (NOR) are ancestral features only for Wertheimerinae and the most clades of Doradinae. Some exceptions were found in Trachydoras paraguayensis (2n=56), Trachydoras steindachneri (2n=60), Ossancora punctata (2n=66) and Platydoras hancockii whose karyotypes showed a multiple NOR system. The large thorny catfishes, such as Pterodoras granulosus, Oxydoras niger and Centrodoras brachiatus share several karyotype features, with subtle variations only regarding their heterochromatin distribution. On the other hand, a remarkable karyotypic variability has been reported in the fimbriate barbells thorny catfishes. These two contrasting karyoevolution trajectories emerged from a complex interaction between chromosome rearrangements (e.g., inversions and Robertsonian translocations) and mechanisms of heterochromatin dispersion. Moreover, we believe that biological features, such as microhabitats preferences, populational size, low vagility and migratory behavior played a key role during the origin and maintenance of chromosome diversity in Doradinae subfamily.     

Dynamic chromosomes

A report on the sixth biannual FASEB conference on Yeast Chromosome Structure, Replication and Segregation held in Snowmass, Colorado, 19-24 August, 2000.     

Mechanisms of glutamate release from astrocytes: gap junction "hemichannels", purinergic receptors and exocytotic release

Neuronal exocytotic release of glutamate at synapses involves a highly specialized vesicular apparatus, consisting of a variety of proteins connected to the vesicles or required for vesicular fusion to the presynaptic membrane. Astrocytes also release glutamate, and recent evidence indicates that this release can modify neuronal function. Several mechanisms have been proposed for astrocytic release of glutamate under pathological conditions, such as reversal of glutamate transporters and opening of volume sensitive ion channels. In this review we limit our discussion to findings supporting the exocytotic release of glutamate, as well as two new pathways implicated in this release, the ionotropic (P2X) purinergic receptors and gap junction hemichannels.