Systematic search for putative new domain families in Mycoplasma gallisepticum genome

Background

Stem cell factor (SCF) receptor c-Kit is recognized as a key signaling molecule, which transduces signals for the proliferation, differentiation and survival of stem cells. Binding of SCF to its receptor triggers transactivation, leading to the recruitment of kinases and phosphatases to the docking platforms of c-Kit catalytic domain. Tyrosine phosphatase-1 (Shp-1) deactivates/attenuates 'Kit' kinase activity. Whereas, Asp816Val mutation in the Kit activation loop transforms kinase domain to a constitutively activated state (switch off-to-on state), in a ligand-independent manner. This phenomenon completely abrogates negative regulation of Shp-1. To predict the possible molecular basis of interaction between c-Kit and Shp-1, we have performed an in silico protein-protein docking study between crystal structure of activated c-Kit (phosphorylated c-Kit) and full length crystal structure of Shp-2, a close structural counterpart of Shp-1.

Findings

Study revealed a stretch of conserved amino acids (Lys818 to Ser821) in the Kit activation domain, which makes decisive H-bonds with N-sh2 and phosphotyrosine binding pocket residues of the phosphatase. These H-bonds may impose an inhibitory steric hindrance to the catalytic domain of c-Kit, there by blocking further interaction of the activation loop molecules with incoming kinases. We have also predicted a phosphotyrosine binding pocket in SH2 domains of Shp-1, which is found to be predominantly closer to a catalytic groove like structure in c-Kit kinase domain.

Conclusions

This study predicts that crucial hydrogen bonding between N-sh2 domain of Shp-1 and Kit activation loop can modulate the negative regulation of c-Kit kinase by Shp-1. Thus, this finding is expected to play a significant role in designing suitable gain-of-function c-Kit mutants for inducing conditional proliferation of hematopoietic stem cells.     

In vitro inhibition of human influenza A virus replication by chloroquine

Inhibition of the human cytomegalovirus UL97 kinase by maribavir is thought to be responsible for the antiviral activity of this compound. Some mutations that confer resistance to maribavir map to UL97, however additional mutations that also confer resistance to the drug were mapped to UL27. These open reading frames share a low level of homology, yet the function of pUL27 remains unknown. A recombinant virus with a deletion in the UL27 open reading frame was reported previously to exhibit a slight replication deficit, but a more important function in vivo was hypothesized given its homology to the UL97 kinase. The potential for an important function in vivo was investigated by determining if these knockout viruses could replicate in human tissue implanted in SCID mice. None of the AD169 derived viruses replicated well in the implanted thymus/liver tissue, and is consistent with previous observations, although all of the viruses replicated to some degree in retinal tissue implants. Replication of the parent viruses was observed at 7 days post inoculation, whereas no replication was detected with any of the recombinant viruses with deletions in UL27. By day 14, replication was detected in two of the three knockout viruses and in all of the viruses by day 42. These data are consistent with minimal defects observed in cell culture, but are not consistent with an important role for UL27 in vivo. We conclude that UL27 is not required for viral replication in vivo.     

Unusual pattern of bacterial ice nucleation gene evolution

Bacterial ice nucleation activity (INA+ phenotype) can be traced to the product of a single gene, ina. A remarkably sparse distribution of this phenotype within three bacterial genera indicates that the ina gene may have followed an unusual evolutionary path. Southern blot analyses, coupled with assays for ice-nucleating ability, revealed that within four bacterial species an ina gene is present in some strains but absent from others. Results of hybridization experiments using DNA fragments that flank the ina gene suggested that the genotypic dimorphism of ina may be anomalous. A phylogenetic analysis of 16S ribosomal RNA gene sequences from a total of 14 ina+ and ina- bacterial strains indicated that the ina+ bacteria are not monophyletic but instead phylogenetically interspersed among ina- bacteria. The relationships of ina+ bacteria inferred from ina sequence did not coincide with those inferred from the 16S data. These results suggest the possibility of horizontal transfer in the evolution of bacterial ina genes.      

β-Decay and the origin of optical purity

Improving Keszthelyis simple model the evolutionary appearance of concentration difference of enanthiomeric compounds due to their differential decomposition by -rays is investigated taking into account the racemization as well. It is shown that if the difference in the cross sections is very small then the resulting concentration difference will never exceed the statistical fluctuations, while in the case of a sufficiently large difference in the cross sections the concentration difference can overgrow the statistical fluctuations in an evolutionary reasonable period of time. The relative difference of the concentrations, however, will be generally much smaller than that of the cross sections. Therefore some other, amplifying mechanism must be postulated in order to explain the optical purity of living beings.     

Gene expression profiling of the preclinical scrapie-infected hippocampus

The molecular events that underlie prion disease neuropathology remain poorly defined. Within the hippocampus of the ME7/CV mouse scrapie model, profound CA1 neuronal loss occurs between 160 and 180 days post-infection (dpi). To elucidate the molecular events that may contribute to this neuronal loss, we have applied Affymetrix high-density oligonucleotide probe arrays to the study of ME7-infected hippocampal gene expression at 170 dpi. The study has identified 78 genes that are differentially expressed greater than 1.5-fold within the preclinical ME7-infected hippocampus prior to the profound late stage glial cell activation. The results indicate oxidative and endoplasmic reticulum (ER) stress, activated ER and mitochondrial apoptosis pathways, and activated cholesterol biosynthesis within the scrapie-infected hippocampus, and offer insight into the molecular events which underlie the neuropathology.     

The Drosophila embargoed gene is required for larval progression and encodes the functional homolog of schizosaccharomyces Crm1

The CRM1 (Exportin 1) protein is a receptor for leucine-rich nuclear export signal sequences. We have molecularly characterized the Drosophila melanogaster embargoed (emb) gene and find that it encodes a product with 49 and 71% sequence identity to the fission yeast Schizosaccharomyces pombe and human CRM1 proteins, respectively. We show that expression of the emb cDNA is sufficient to suppress the growth phenotype of both conditional-lethal and null S. pombe crm1(-) mutant strains, suggesting that emb encodes the functional homologue of the S. pombe Crm1 protein. Through mutagenesis screens we have recovered a series of recessive lethal emb mutations. There is a substantial maternal contribution of emb mRNA and animals hemizygous for our emb alleles can develop to second instar larvae but persist at this stage and consistently fail to undergo the molt to the third instar stage. We see a nuclear accumulation of endogenous actin in the intestinal epithelial cells of the emb mutant larvae, consistent with a role for the emb gene product in nuclear export of actin protein.     

Effects of ICI 182780 on estrogen receptor expression,fluid absorption and sperm motility in the epididymis of the bonnet monkey

Background  

The importance of estrogen in regulation of fluid absorption and sperm maturation in the rodent epididymis has been established from studies on estrogen receptor-alpha knockout mice. However, functional studies on the role of estrogen in primate epididymis have been few. The main objective of this study was therefore to extend these observations and systematically analyze the presence and function of estrogen receptors in modulating the function of the primate epididymis, using the bonnet monkey (Macaca radiata) as a model system.     

Oncogenic comparison of human papillomavirus type 58 E7 variants

Human papillomavirus 58 (HPV58) ranks the second or third in East Asian cervical cancers. Current studies on HPV58 are scarce and focus on the prototype. Previously, we identified the three most common circulating HPV58 E7 strains contained amino acid alterations: G41R/G63D (51%), T20I/G63S (22%) and T74A/D76E (14%) respectively. Among them, the T20I/G63S variant (V1) had a stronger epidemiological association with cervical cancer. We therefore suggested that V1 possessed stronger oncogenicity than the other two variants. Here, we performed phenotypic assays to characterize and compare their oncogenicities with HPV58 E7 prototype. Our results showed that overexpression of V1 conferred a higher colony‐forming ability to primary murine epithelial cells than prototype (P < 0.05) and other variants, implicating its higher immortalising potential. Further experiments showed that both V1 and prototype enhanced the anchorage‐independent growth of NIH/3T3 cells (P < 0.001), implicating their stronger transforming power than the two other variants. Moreover, they possessed an increased ability to degrade pRb (P < 0.001), which is a major effector pathway of E7‐driven oncogenesis. Our work represents the first study to compare the oncogenicities of HPV58 E7 prototype and variants. These findings deepened our understanding of HPV58 and might inform clinical screening and follow‐up strategy.     

Sequential colonization by river periphyton analysed by microscopy and molecular fingerprinting

1. An artificial glass substratum was incubated in the River Danube for a period of 28 days in order to detect the sequential colonization of microorganisms.
2. Light and fluorescent microscopy showed that microalgae and the picoalgal fraction on the slides increased rapidly over the first 2 weeks of colonization. Diatoms were numerically the most abundant component of the periphyton and their species richness and diversity increased rapidly in the early phase of colonization whereas diversity subsequently increased moderately.
3. Evenness of the diatom community was initially high, lower in the intermediate phase and again higher later on. Succession involving early, intermediate and late colonizer species was observed. Community composition during the first 5 days of colonization was very different from later stages whereas there were only minor changes subsequently.
4. Molecular community analysis by means of terminal restriction fragment length polymorphism analysis of PCR amplified 16S rRNA and 18S rRNA genes pointed to even larger differences between the composition of samples obtained early and late in the period.
5. The number of 18S rRNA and 16S rRNA terminal restriction fragments (T-RF-s) was variable over the colonization period and the fragment patterns of both the bacterial and eukaryotic portion of the microbial community were variable, with most T-RF-s unique to a single sample, suggesting a wide diversity and dynamic properties of periphytic organisms.