A fresh look at the evolution and diversification of photochemical reaction centers

In this review, I reexamine the origin and diversification of photochemical reaction centers based on the known phylogenetic relations of the core subunits, and with the aid of sequence and structural alignments. I show, for example, that the protein folds at the C-terminus of the D1 and D2 subunits of Photosystem II, which are essential for the coordination of the water-oxidizing complex, were already in place in the most ancestral Type II reaction center subunit. I then evaluate the evolution of reaction centers in the context of the rise and expansion of the different groups of bacteria based on recent large-scale phylogenetic analyses. I find that the Heliobacteriaceae family of Firmicutes appears to be the earliest branching of the known groups of phototrophic bacteria; however, the origin of photochemical reaction centers and chlorophyll synthesis cannot be placed in this group. Moreover, it becomes evident that the Acidobacteria and the Proteobacteria shared a more recent common phototrophic ancestor, and this is also likely for the Chloroflexi and the Cyanobacteria. Finally, I argue that the discrepancies among the phylogenies of the reaction center proteins, chlorophyll synthesis enzymes, and the species tree of bacteria are best explained if both types of photochemical reaction centers evolved before the diversification of the known phyla of phototrophic bacteria. The primordial phototrophic ancestor must have had both Type I and Type II reaction centers

Expression profiling with arrays of randomly disrupted genes in mouse embryonic stem cells leads to in vivo functional analysis

DNA arrays are capable of profiling the expression patterns of many genes in a single experiment. After finding a gene of interest in a DNA array, however, labor-intensive gene-targeting experiments sometimes must be performed for the in vivo analysis of the gene function. With random gene trapping, on the other hand, it is relatively easy to disrupt and retrieve hundreds of genes/gene candidates in mouse embryonic stem (ES) cells, but one could overlook potentially important gene-disruption events if only the nucleotide sequences and not the expression patterns of the trapped DNA segments are analyzed. To combine the benefits of the above two experimental systems, we first created ≈900 genetrapped mouse ES cell clones and then constructed arrays of cDNAs derived from the disrupted genes. By using these arrays, we identified a novel gene predominantly expressed in the mouse brain, and the corresponding ES cell clone was used to produced mice homozygous for the disrupted allele of the gene. Detailed analysis of the knockout mice revealed that the gene trap vector completely abolished gene expression downstream of its integration site. Therefore, identification of a gene or novel gene candidate with an interesting expression pattern by using this type of DNA array immediately allows the production of knockout mice from an ES cell clone with a disrupted allele of the sequence of interest

Epigenetic silencing of serine protease HTRA1 drives polyploidy

Background: Increased numbers and improperly positioned centrosomes, aneuploidy or polyploidy, and chromosomal instability are frequently observed characteristics of cancer cells. While some aspects of these events and the checkpoint mechanisms are well studied, not all players have yet been identified. As the role of proteases other than the proteasome in tumorigenesis is an insufficiently addressed question, we investigated the epigenetic control of the widely conserved protease HTRA1 and the phenotypes of deregulation. Methods: Mouse embryonal fibroblasts and HCT116 and SW480 cells were used to study the mechanism of epigenetic silencing of HTRA1. In addition, using cell biological and genetic methods, the phenotypes of downregulation of HTRA1 expression were investigated. Results: HTRA1 is epigenetically silenced in HCT116 colon carcinoma cells via the epigenetic adaptor protein MBD2. On the cellular level, HTRA1 depletion causes multiple phenotypes including acceleration of cell growth, centrosome amplification and polyploidy in SW480 colon adenocarcinoma cells as well as in primary mouse embryonic fibroblasts (MEFs). Conclusions: Downregulation of HTRA1 causes a number of phenotypes that are hallmarks of cancer cells suggesting that the methylation state of the HtrA1 promoter may be used as a biomarker for tumour cells or cells at risk of transformation