Characterization of the human SCF ubiquitin ligases - structure, function, and regulation

The SCF ubiquitin ligase complex of budding yeast triggers DNA replication by cata lyzi ng ubiquitination of the S phase CDK inhibitor SIC1. SCF is composed of several evolutionarily conserved proteins, including ySKP1, CDC53 (Cullin), and the F-box protein CDC4. We isolated hSKP1 in a two-hybrid screen with hCUL1, the human homologue of CDC53. We showed that hCUL1 associates with hSKP1 in vivo and directly interacts with hSKP1 and the human F-box protein SKP2 in vitro, forming an SCF-Iike particle. Moreover, hCUL1 complements the growth defect of yeast CDC53^(ts) mutants, associates with ubiquitination-promoting activity in human cell extracts, and can assemble into functional, chimeric ubiquitin ligase complexes with yeast SCF components. These data demonstrated that hCUL1 functions as part of an SCF ubiquitin ligase complex in human cells. However, purified human SCF complexes consisting of CUL1, SKP1, and SKP2 are inactive in vitro, suggesting that additional factors are required.

Subsequently, mammalian SCF ubiquitin ligases were shown to regulate various physiological processes by targeting important cellular regulators, like lĸBα, β-catenin, and p27, for ubiquitin-dependent proteolysis by the 26S proteasome. Little, however, is known about the regulation of various SCF complexes. By using sequential immunoaffinity purification and mass spectrometry, we identified proteins that interact with human SCF components SKP2 and CUL1 in vivo. Among them we identified two additional SCF subunits: HRT1, present in all SCF complexes, and CKS1, that binds to SKP2 and is likely to be a subunit of SCF5^(SKP2) complexes. Subsequent work by others demonstrated that these proteins are essential for SCF activity. We also discovered that COP9 Signalosome (CSN), previously described in plants as a suppressor of photomorphogenesis, associates with CUL1 and other SCF subunits in vivo. This interaction is evolutionarily conserved and is also observed with other Cullins, suggesting that all Cullin based ubiquitin ligases are regulated by CSN. CSN regulates Cullin Neddylation presumably through CSNS/JAB1, a stochiometric Signalosome subunit and a putative deneddylating enzyme. This work sheds light onto an intricate connection that exists between signal transduction pathways and protein degradation machinery inside the cell and sets stage for gaining further insights into regulation of protein degradation.

Maintaining internal representations: The role of the human superior parietal lobe

In sensorimotor integration, sensory input and motor output signals are combined to provide an internal estimate of the state of both the world and one's own body. Although a single perceptual and motor snapshot can provide information about the current state, computational models show that the state can be optimally estimated by a recursive process in which an internal estimate is maintained and updated by the current sensory and motor signals. These models predict that an internal state estimate is maintained or stored in the brain. Here we report a patient with a lesion of the superior parietal lobe who shows both sensory and motor deficits consistent with an inability to maintain such an internal representation between updates. Our findings suggest that the superior parietal lobe is critical for sensorimotor integration, by maintaining an internal representation of the body's state.

Y-chromosome evidence for no independent origin of modern human in China

East Asia is one of the few regions in the world where a large number of human fossils have been unearthed. The continuity of hominid fossils in East Asia, particularly in China has been presented as strong evidence supporting an independent origin of mod

Rapid evolution, genetic variations, and functional association of the human spermatogenesis-related gene NYD-SP12

NYD-SP12 is a recently identified spermatogenesis-related gene with a pivotal role in human testis development. In this study, we analyzed between-species divergence and within-species variation of NYD-SP12 in seven representative primate species, four wo

Functional characterization of the human-specific (type II) form of kallikrein 8, a gene involved in learning and memory

Kallikrein 8 (KLK8) is a serine protease functioning in the central nervous system, and essential in many aspects of neuronal activities. Sequence comparison and gene expression analysis among diverse primate species identified a human-specific splice for

Origination and evolution of a human-specific transmembrane protein gene, c1orf37-dup

A transmembrane protein gene, c1orf37-dup, was identified as a young gene specific to humans. It was derived from the conserved c1orf37 gene through retroposition after the divergence of human and chimpanzee. This gene has evolved rapidly driven by positi

A novel high molecular weight metalloproteinase cleaves fragment F1 of activated human prothrombin

A hemorrhagic proteinase, jerdohagin, was purified from Trimeresurus jerdonii venom by gel filtration and ion-exchange chromatographies. It was a single chain polypeptide with an apparent molecular weight of 96 kDa as estimated by SDS-PAGE under the non-reducing and reducing conditions. Internal peptide sequencing indicated that it consisted of metalloproteinase, disintegrin-like and cysteine-rich domains and belonged to the class III snake venom metalloproteinases (class P-III SVMPs). Like other typical metalloproteinases, hemorrhagic activities of jerdohagin were completely inhibited by EDTA, but not by PMSF. Jerdohagin preferentially degraded a-chain of human fibrinogen. Interestingly, jerdohagin did not activate human prothrombin, whereas it cleaved human prothrombin and fragment F1 of activated human prothrombin. (C) 2004 Elsevier Ltd. All rights reserved.

Pitfalls in the analysis of ancient human mtDNA

The retrieval of DNA from ancient human specimens is not always successful owing to DNA deterioration and contamination although it is vital to provide new insights into the genetic structure of ancient people and to reconstruct the past history. Normally, only short DNA fragments can be retrieved from the ancient specimens. How to identify the authenticity of DNA obtained and to uncover the information it contained are difficult. We employed the ancient mtDNAs reported from Central Asia (including Xinjiang, China) as an example to discern potentially extraneous DNA contamination based on the updated mtDNA phylogeny derived from mtDNA control region, coding region, as well as complete sequence information. Our results demonstrated that many mtDNAs reported are more or less problematic. Starting from a reliable mtDNA phylogeney and combining the available modern data into analysis, one can ascertain the authenticity of the ancient DNA, distinguish the potential errors in a data set, and efficiently decipher the meager information it harbored. The reappraisal of the mtDNAs with the age of more than 2000 years from Central Asia gave support to the suggestion of extensively (pre)historical gene admixture in this region.

Effects of dietary supplementation of commercial human probiotic and antibiotic on the growth rate and content of intestinal microflora in ornamental fishes

The effects of dietary supplementation of commercial human probiotic, Lactobacil and antibiotic, oxytetracycline on the growth, survival, disease resistance and content of intestinal microflora in two ornamental fishes, viz., goldfish, Carassius auratus and swordtail, Xiphophorus helleri were studied. The total wet weight gain, food conversion ratio and specific growth rate of C. auratus did not vary significantly (p>0.05) among treatments. While in X. helleri, significant differences existed in the total wet weight gain, survival, food conversion ratio and specific growth rate among treatment groups (p<0.05). The counts of antibiotic resistant bacteria in fish gut increased with days of culture in all the treatments and the increase was more in antibiotic fed fishes. A reduction in the development of antibiotic resistance among the bacterial flora of fish gut was noticed in probiotic fed groups of C auratus and X. helleri. The results of the present study revealed that the effects of human probiotic on the growth, survival and disease resistance of ornamental fishes are variable and difficult to reproduce the similar effect on different species.

Structural and functional characterization of the human CCR5 receptor in complex with HIV gp120 envelope glycoprotein and CD4 receptor by molecular modeling studies

The entry of human immunodeficiency virus (HIV) into cells depends on a sequential interaction of the gp120 envelope glycoprotein with the cellular receptors CD4 and members of the chemokine receptor family. The CC chemokine receptor CCR5 is such a receptor for several chemokines and a major coreceptor for the entry of R5 HIV type-1 (HIV-1) into cells. Although many studies focus on the interaction of CCR5 with HIV-1, the corresponding interaction sites in CCR5 and gp120 have not been matched. Here we used an approach combining protein structure modeling, docking and molecular dynamics simulation to build a series of structural models of the CCR5 in complexes with gp120 and CD4. Interactions such as hydrogen bonds, salt bridges and van der Waals contacts between CCR5 and gp120 were investigated. Three snapshots of CCR5-gp120-CD4 models revealed that the initial interactions of CCR5 with gp120 are involved in the negatively charged N-terminus (Nt) region of CCR5 and positively charged bridging sheet region of gp120. Further interactions occurred between extracellular loop2 (ECL2) of CCR5 and the base of V3 loop regions of gp120. These interactions may induce the conformational changes in gp120 and lead to the final entry of HIV into the cell. These results not only strongly support the two-step gp120-CCR5 binding mechanism, but also rationalize extensive biological data about the role of CCR5 in HIV-1 gp120 binding and entry, and may guide efforts to design novel inhibitors.