Exploring the mode-of-action of bioactive compounds by chemical-genetic profiling in yeast

Discovering target and off-target effects of specific compounds is critical to drug discovery and development. We generated a compendium of "chemical-genetic interaction" profiles by testing the collection of viable yeast haploid deletion mutants for hypersensitivity to 82 compounds and natural product extracts. To cluster compounds with a similar mode-of-action and to reveal insights into the cellular pathways and proteins affected, we applied both a hierarchical clustering and a factorgram method, which allows a gene or compound to be associated with more than one group. In particular, tamoxifen, a breast cancer therapeutic, was found to disrupt calcium homeostasis and phosphatidylserine (PS) was recognized as a target for papuamide B, a cytotoxic lipopeptide with anti-HIV activity. Further, the profile of crude extracts resembled that of its constituent purified natural product, enabling detailed classification of extract activity prior to purification. This compendium should serve as a valuable key for interpreting cellular effects of novel compounds with similar activities.     

Prenatally identified case of mosaicism of chromosome 16 trisomy

We present the prenatally identified case of mosaicism of chromosome 16 trisomy. A patient with the pregnancy complicated in the first trimester by the threat of breaking was refered to the high risk group according to the results of the screening program. The ultrasonic research revealed a number of phenotypical pathologies in 19-weeks-old fetus such as congenital heart disease (ventricular septal defect), hyperechoic bowel, single umbilical artery and some other ones. Cytogenetical and FISH analyses of the placental villi revealed karyotype with chromosome 16 trisomy. The further research of amniotic fluid cells revealed the karyotype of fetus as mos47,XX,+16 / 46,XX. The pathologoanatomic research of the abortus has verified the multiple congenital malformations.     

Karyotyping results of the material of spontaneous abortions and miscarriages after using assisted reproductive technologies

The results of a molecular cytogenetic study of the material of spontaneous abortions and nonprogressive pregnancies in the first trimester in 43 couples who were treated with various ART methods are presented in this paper. Chromosomal pathologies (CPs) were present in 28 (65%) samples of chorionic villi. A comparative analysis of the frequency and types of CPs in groups composed according to the pathological states in the semen of males was performed. Recommendations to improve the efficiency of ART are developed based on the data.     

Linking the kinome and phosphorylome--a comprehensive review of approaches to find kinase targets

Protein phosphorylation is associated with most cell signaling and developmental processes in eukaryotes. Despite the vast extent of the phosphoproteome within the cell, connecting specific kinases with relevant targets remains a significant experimental frontier. The challenge of linking kinases and their substrates reflects the complexity of kinase function. For example, kinases tend to exert their biological effects through supernumerary, redundant phosphorylation, often on multiple protein complex components. Although these types of phosphorylation events are biologically significant, those kinases responsible are often difficult to identify. Recent methods for global analysis of protein phosphorylation promise to substantially accelerate efforts to map the dynamic phosphorylome. Here, we review both conventional methods to identify kinase targets and more comprehensive genomic and proteomic approaches to connect the kinome and phosphorylome.     

Activation of the Cdc42p GTPase by cyclin-dependent protein kinases in budding yeast

Cyclin-dependent kinases (CDKs) trigger essential cell cycle processes including critical events in G1 phase that culminate in bud emergence, spindle pole body duplication, and DNA replication. Localized activation of the Rho-type GTPase Cdc42p is crucial for establishment of cell polarity during G1, but CDK targets that link the Cdc42p module with cell growth and cell cycle commitment have remained largely elusive. Here, we identify the GTPase-activating protein (GAP) Rga2p as an important substrate related to the cell polarity function of G1 CDKs. Overexpression of RGA2 in the absence of functional Pho85p or Cdc28p CDK complexes is toxic, due to an inability to polarize growth. Mutation of CDK consensus sites in Rga2p that are phosphorylated both in vivo and in vitro by Pho85p and Cdc28p CDKs results in a loss of G1 phase-specific phosphorylation. A failure to phosphorylate Rga2p leads to defects in localization and impaired polarized growth, in a manner dependent on Rga2p GAP function. Taken together, our data suggest that CDK-dependent phosphorylation restrains Rga2p activity to ensure appropriate activation of Cdc42p during cell polarity establishment. Inhibition of GAPs by CDK phosphorylation may be a general mechanism to promote proper G1-phase progression.