Immunochemical and amino-terminal sequence comparison of two cytoadhesins indicates they contain similar or identical beta subunits and distinct alpha subunits

Platelet membrane glycoprotein (GP) IIb-IIIa is functionally and antigenically related to proteins present on many cell types, suggesting that it is a member of the proposed cytoadhesin family of membrane proteins. We have compared the purified tissue vitronectin receptor (VnR) with GP IIb-IIIa. Anti-VnR immunoprecipitated GP IIb-IIIa and a related endothelial cell protein. In immunoblots, GP IIIa reacted with anti-VnR and the beta subunit of the VnR reacted with poly and monoclonal anti-GP IIIa. In contrast, the alpha subunit of the VnR failed to react either with a polyclonal anti-GP IIb or with monoclonal anti-GP IIb. Furthermore, the amino-terminal sequence of GP IIIa and the beta subunit of VnR were identical at determined residues while the alpha subunit and the GP IIb were different, but showed 33% identity. These data indicate the identity or close homology of GP IIIa and the beta subunit of the VnR. In contrast, the alpha subunit and GP IIb are distinct polypeptides which may be homologous. Since GP IIb-IIIa and the VnR differ in ligand recognition specificity, the data also suggest that this specificity may be governed by the alpha subunit of cytoadhesins.     

Occupancy of an adhesive glycoprotein receptor modulates expression of an antigenic site involved in cell adhesion

Binding of ligands that contain Arg-Gly-Asp to adhesion receptors induces cell spreading and aggregation and alters gene expression, possibly due to conformational changes within occupied adhesion receptors. PMI-1 is a monoclonal antibody which reacts with the platelet fibrinogen receptor, glycoprotein IIb-IIIa, and reports such a conformational change. ADP stimulation of platelets results in a fibrinogen-dependent increase in binding of the PMI-1 antibody. Peptides containing Arg-Gly-Asp also reversibly increase the binding of this antibody to cells and to purified glycoprotein IIb-IIIa. The PMI-1 antibody inhibits platelet adhesion and spreading on certain substrata (Shadle, P. J., Ginsberg, M. H., Plow, E. F., and Barondes, S. H. (1984) J. Cell Biol. 99, 2056-2060); thus this occupancy-modulated site may participate in adhesive function.     

A YAC contig encompassing the Treacher Collins syndrome critical region at 5q31.3-32.

Treacher Collins syndrome (TCOF1) is an autosomal dominant disorder of craniofacial development the features of which include conductive hearing loss and cleft palate. Previous studies have localized the TCOF1 locus between D5S519 (proximal) and SPARC (distal), a region of 22 centirays as estimated by radiation hybrid mapping. In the current investigation we have created a contig across the TCOF1 critical region, using YAC clones. Isolation of a novel short tandem repeat polymorphism corresponding to the end of one of the YACs has allowed us to reduce the size of the critical region to approximately 840 kb, which has been covered with three nonchimeric YACs. Restriction mapping has revealed that the region contains a high density of clustered rare-cutter restriction sites, suggesting that it may contain a number of different genes. The results of the present investigation have further allowed us to confirm that the RPS14 locus lies proximal to the critical region and can thereby be excluded from a role in the pathogenesis of TCOF1, while ANX6 lies within the TCOF1 critical region and remains a potential candidate for the mutated gene.     

The Src Homology 3 Domain-containing Guanine Nucleotide Exchange Factor Is Overexpressed in High-grade Gliomas and Promotes Tumor Necrosis Factor-like Weak Inducer of Apoptosis-Fibroblast Growth Factor-inducible 14-induced Cell Migration and Invasion via Tumor Necrosis Factor Receptor-associated Factor 2

Glioblastoma (GB) is the highest grade of primary adult brain tumors, characterized by a poorly defined and highly invasive cell population. Importantly, these invading cells are attributed with having a decreased sensitivity to radiation and chemotherapy. TNF-like weak inducer of apoptosis (TWEAK)-Fn14 ligand-receptor signaling is one mechanism in GB that promotes cell invasiveness and survival and is dependent upon the activity of multiple Rho GTPases, including Rac1. Here we report that Src homology 3 domain-containing guanine nucleotide exchange factor (SGEF), a RhoG-specific guanine nucleotide exchange factor, is overexpressed in GB tumors and promotes TWEAK-Fn14-mediated glioma invasion. Importantly, levels of SGEF expression in GB tumors inversely correlate with patient survival. SGEF mRNA expression is increased in GB cells at the invasive rim relative to those in the tumor core, and knockdown of SGEF expression by shRNA decreases glioma cell migration in vitro and invasion ex vivo. Furthermore, we showed that, upon TWEAK stimulation, SGEF is recruited to the Fn14 cytoplasmic tail via TRAF2. Mutation of the Fn14-TRAF domain site or depletion of TNF receptor-associated factor 2 (TRAF2) expression by siRNA oligonucleotides blocked SGEF recruitment to Fn14 and inhibited SGEF activity and subsequent GB cell migration. We also showed that knockdown of either SGEF or RhoG diminished TWEAK activation of Rac1 and subsequent lamellipodia formation. Together, these results indicate that SGEF-RhoG is an important downstream regulator of TWEAK-Fn14-driven GB cell migration and invasion.     

A curated gene list for expanding the horizons of pigmentation biology

Over the past century, studies of human pigmentary disorders along with mouse and zebrafish models have shed light on the many cellular functions associated with visible pigment phenotypes. This has led to numerous genes annotated with the ontology term “pigmentation” in independent human, mouse, and zebrafish databases. Comparisons among these datasets revealed that each is individually incomplete in documenting all genes involved in integument‐based pigmentation phenotypes. Additionally, each database contained inherent species‐specific biases in data annotation, and the term “pigmentation” did not solely reflect integument pigmentation phenotypes. This review presents a comprehensive, cross‐species list of 650 genes involved in pigmentation phenotypes that was compiled with extensive manual curation of genes annotated in OMIM, MGI, ZFIN, and GO. The resulting cross‐species list of genes both intrinsic and extrinsic to integument pigment cells provides a valuable tool that can be used to expand our knowledge of complex, pigmentation‐associated pathways.     

Differential role of proline-rich tyrosine kinase 2 and focal adhesion kinase in determining glioblastoma migration and proliferation

The propensity of malignant gliomas to invade surrounding brain tissue contributes to poor clinical outcome. Integrin-mediated adhesion to extracellular matrix regulates the migration and proliferation of many cell types, but its role in glioma progression is undefined. We investigated the role of the cytoplasmic tyrosine kinases FAK and Pyk2, potential integrin effectors, in the phenotypic determination of four different human glioblastoma cell lines. While FAK expression was similar between the four cell lines, increased FAK activity correlated with high proliferation and low migratory rates. In contrast, Pyk2 activity was significantly increased in migratory cell lines and depressed in proliferative cell lines. Overexpression of Pyk2 stimulated migration, whereas FAK overexpression inhibited cell migration and stimulated cellular proliferation. These data suggest that FAK and Pyk2 function as important signaling effectors in gliomas and indicate that their differential regulation may be determining factors in the temporal development of proliferative or migrational phenotypes.     

Construction of a BAC library and generation of BAC end sequence-tagged connectors for genome sequencing of the African malaria mosquito Anopheles gambiae

A Bacterial Artificial Chromosome (BAC) genomic DNA library of Anopheles gambiae, the major human malaria vector in sub-Saharan Africa, was constructed and characterized. This library (ND-TAM) is composed of 30,720 BAC clones in eighty 384-well plates. The estimated average insert size of the library is 133 kb, with an overall genome coverage of approximately 14-fold. The ends of approximately two-thirds of the clones in the library were sequenced, yielding 32,340 pair-mate ends. A statistical analysis (G-test) of the results of PCR screening of the library indicated a random distribution of BACs in the genome, although one gap encompassing the white locus on the X-chromosome was identified. Furthermore, combined with another previously constructed BAC library (ND-1), ~2,000 BACs have been physically mapped by polytene chromosomal in situ hybridization. These BAC end pair mates and physically mapped BACs have been useful for both the assembly of a fully sequenced A. gambiae genome and for linking the assembled sequence to the three polytene chromosomes. This ND-TAM library is now publicly available at both http://www.malaria.mr4.org/mr4pages/index.html/ and http://hbz.tamu.edu/, providing a valuable resource to the mosquito research community.     

Our changeable memories: legal and practical implications

The malleability of memory is becoming increasingly clear. Many influences can cause memories to change or even be created anew, including our imaginations and the leading questions or different recollections of others. The knowledge that we cannot rely on our memories, however compelling they might be, leads to questions about the validity of criminal convictions that are based largely on the testimony of victims or witnesses. Our scientific understanding of memory should be used to help the legal system to navigate this minefield.