Erratum

Neurotransmitter receptor trafficking and the regulation of synaptic strength. Traffic 2001:2(7):437–448.     

Characterization of the Rac guanine nucleotide exchange factor P-Rex1 in platelets

Background

Blood platelets undergo a carefully regulated change in shape to serve as the primary mediators of hemostasis and thrombosis. These processes manifest through platelet spreading and aggregation and are dependent on platelet actin cytoskeletal changes orchestrated by the Rho GTPase family member Rac1. To elucidate how Rac1 is regulated in platelets, we captured Rac1-interacting proteins from platelets and identified Rac1-associated proteins by mass spectrometry.

Findings

Here, we demonstrate that Rac1 captures the Rac guanine nucleotide exchange factor P-Rex1 from platelet lysates. Western blotting experiments confirmed that P-Rex1 is expressed in platelets and associated with Rac1. To investigate the functional role of platelet P-Rex1, platelets from P-Rex1 ^-/- -deficient mice were treated with platelet agonists or exposed to platelet activating surfaces of fibrinogen, collagen and thrombin. Platelets from P-Rex1 ^-/- mice responded to platelet agonists and activating surfaces similarly to wild type platelets.

Conclusions

These findings suggest that P-Rex1 is not required for Rac1-mediated platelet activation and that the GEF activities of P-Rex1 may be more specific to GPCR chemokine receptor mediated processes in immune cells and tumor cells.     

Hybrid metabolic flux analysis: combining stoichiometric and statistical constraints to model the formation of complex recombinant products

Background  

Stoichiometric models constitute the basic framework for fluxome quantification in the realm of metabolic engineering. A recurrent bottleneck, however, is the establishment of consistent stoichiometric models for the synthesis of recombinant proteins or viruses. Although optimization algorithms for in silico metabolic redesign have been developed in the context of genome-scale stoichiometric models for small molecule production, still rudimentary knowledge of how different cellular levels are regulated and phenotypically expressed prevents their full applicability for complex product optimization.     

Cyclic GMP Regulates Nicotine-Induced Secretion from Cultured Bovine Adrenal Chromaffin Cells: Effects of 8–Bromo–Cyclic GMP, Atrial Natriuretic Peptide, and Nitroprusside (Nitric Oxide)

Methacholine, atrial natriuretic peptide (ANP), nitroprusside (nitric oxide), angiotensin II, and bradykinin raised cyclic GMP (cGMP) levels in cultured bovine adrenal chromaffin cells. The role of cGMP in secretion from chromaffin cells was examined using 8-bromo-cGMP. This analogue had no effect on basal secretion or secretion due to angiotensin II, bradykinin, or a high K+ level but potentiated secretion due to low doses of nicotine. At supramaximal doses of nicotine, 8-bromo-cGMP inhibited secretion. These effects of 8-bromo-cGMP were not due to changes in the nicotine-induced rise in cytosolic calcium concentration. A potentiation of secretion due to low doses of nicotine was also found following simultaneous addition of ANP or nitroprusside, a result suggesting that ANP and nitric oxide (endothelium-derived relaxing factor) could be important regulators of secretion from adrenal chromaffin cells.     

Reconstruction of cell population dynamics using CFSE

Background  

Quantifying cell division and death is central to many studies in the biological sciences. The fluorescent dye CFSE allows the tracking of cell division in vitro and in vivo and provides a rich source of information with which to test models of cell kinetics. Cell division and death have a stochastic component at the single-cell level, and the probabilities of these occurring in any given time interval may also undergo systematic variation at a population level. This gives rise to heterogeneity in proliferating cell populations. Branching processes provide a natural means of describing this behaviour.     

CXCR7 expression disrupts endothelial cell homeostasis and causes ligand-dependent invasion

The homeostatic function of endothelial cells (EC) is critical for a number of physiological processes including vascular integrity, immunity, and wound healing. Indeed, vascular abnormalities resulting from EC dysfunction contribute to the development and spread of malignancies. The alternative SDF-1/CXCL12 receptor CXCR7 is frequently and specifically highly expressed in tumor-associated vessels. In this study, we investigate whether CXCR7 contributes to vascular dysfunction by specifically examining the effect of CXCR7 expression on EC barrier function and motility. We demonstrate that CXCR7 expression in EC results in redistribution of CD31/PECAM-1 and loss of contact inhibition. Moreover, CXCR7+ EC are deficient in barrier formation. We show that CXCR7-mediated motility has no influence on angiogenesis but contributes to another motile process, the invasion of CXCR7+ EC into ligand-rich niches. These results identify CXCR7 as a novel manipulator of EC barrier function via alteration of PECAM-1 homophilic junctions. As such, aberrant expression of CXCR7 in the vasculature has the potential to disrupt vascular homeostasis and could contribute to vascular dysfunction in cancer systems.     

The effect of oligonucleotide microarray data pre-processing on the analysis of patient-cohort studies

Background  

Intensity values measured by Affymetrix microarrays have to be both normalized, to be able to compare different microarrays by removing non-biological variation, and summarized, generating the final probe set expression values. Various pre-processing techniques, such as dChip, GCRMA, RMA and MAS have been developed for this purpose. This study assesses the effect of applying different pre-processing methods on the results of analyses of large Affymetrix datasets. By focusing on practical applications of microarray-based research, this study provides insight into the relevance of pre-processing procedures to biology-oriented researchers.     

Complex phylogeographic history of central African forest elephants and its implications for taxonomy

Background

The mitochondrial genomes of snakes are characterized by an overall evolutionary rate that appears to be one of the most accelerated among vertebrates. They also possess other unusual features, including short tRNAs and other genes, and a duplicated control region that has been stably maintained since it originated more than 70 million years ago. Here, we provide a detailed analysis of evolutionary dynamics in snake mitochondrial genomes to better understand the basis of these extreme characteristics, and to explore the relationship between mitochondrial genome molecular evolution, genome architecture, and molecular function. We sequenced complete mitochondrial genomes from Slowinski's corn snake (Pantherophis slowinskii) and two cottonmouths (Agkistrodon piscivorus) to complement previously existing mitochondrial genomes, and to provide an improved comparative view of how genome architecture affects molecular evolution at contrasting levels of divergence.

Results

We present a Bayesian genetic approach that suggests that the duplicated control region can function as an additional origin of heavy strand replication. The two control regions also appear to have different intra-specific versus inter-specific evolutionary dynamics that may be associated with complex modes of concerted evolution. We find that different genomic regions have experienced substantial accelerated evolution along early branches in snakes, with different genes having experienced dramatic accelerations along specific branches. Some of these accelerations appear to coincide with, or subsequent to, the shortening of various mitochondrial genes and the duplication of the control region and flanking tRNAs.

Conclusion

Fluctuations in the strength and pattern of selection during snake evolution have had widely varying gene-specific effects on substitution rates, and these rate accelerations may have been functionally related to unusual changes in genomic architecture. The among-lineage and among-gene variation in rate dynamics observed in snakes is the most extreme thus far observed in animal genomes, and provides an important study system for further evaluating the biochemical and physiological basis of evolutionary pressures in vertebrate mitochondria.     

Cross-hybridizing snake satellite, Drosophila, and mouse DNA sequences may have arisen independently

Previous reports have interpreted hybridization between snake satellite DNA and DNA clones from a variety of distant taxonomic groups as evidence for evolutionary conservation, which implies common ancestry (homology) and/or convergence (analogy) to produce the cross- hybridizing sequences. We have isolated 11 clones from a genomic library of Drosophila melanogaster, using a cloned 2.5-kb snake satellite probe of known nucleotide sequence. We have also analysed published sequence data from snakes, mice, and Drosophila. These data show that (1) all of the cross-hybridization between the snake, fly, and mouse clones can be accounted for by the presence of either of two tandem repeats, [GATA]n and [GACA]n and (2) these tandem repeats are organized differently among the different species. We find no evidence that these sequences are homologous apart from the existence of the simple repeat itself, although their divergence from a common ancestral sequence cannot be ruled out. The sequences contain a variety of homogeneous clusters of tandem repeats of CATA, GA, TA, and CA, as well as GATA and GACA. We suggest that these motifs may have arisen by a self-accelerating process involving slipped-strand mispairing of DNA. Homogeneity of the clusters might simply be the result of a rate of accumulation of tandem repeats that exceeds that of other mutations.