Pollen tube growth and guidance is regulated by POP2, an Arabidopsis gene that controls GABA levels

During angiosperm reproduction, pollen grains form a tube that navigates through female tissues to the micropyle, delivering sperm to the egg; the signals that mediate this process are poorly understood. Here, we describe a role for gamma-amino butyric acid (GABA) in pollen tube growth and guidance. In vitro, GABA stimulates pollen tube growth, although vast excesses are inhibitory. The Arabidopsis POP2 gene encodes a transaminase that degrades GABA and contributes to the formation of a gradient leading up to the micropyle. pop2 flowers accumulate GABA, and the growth of many pop2 pollen tubes is arrested, consistent with their in vitro GABA hypersensitivity. Some pop2 tubes continue to grow toward ovules, yet they are misguided, presumably because they target ectopic GABA on the ovule surface. Interestingly, wild-type tubes exhibit normal growth and guidance in pop2 pistils, perhaps by degrading excess GABA and sharpening the gradient leading to the micropyle.     

A single nucleotide polymorphism in the<Emphasis Type="Italic"> Emp3</Emphasis> gene defines the H4 minor histocompatibility antigen

Minor histocompatibility antigens (minor H antigen) elicit strong T-cell-mediated responses during both graft rejection and graft versus leukemia (GvL) among MHC-matched individuals (where MHC is major histocompatibility complex). Employing expression-cloning methodology, we have identified a cDNA clone, MI-35, encoding the immunodominant H4^b minor H antigen within the classical mouse H4 complex. The minimal antigenic epitope derived from H4^b presented on K^b class I MHC is SGIVYIHL (SYL8) and the polymorphism is due to CT nucleotide modification in p3 resulting in the change of threonine (ACT) to isoleucine (ATT). The results presented here demonstrate that amino acid variation in the allelic epitopes results in the low abundance of H4^a peptide. The differential peptide copy number resulted in an immunodominant cytotoxic T cells (CTL) response directed against H4^b while the anti-B6 response directed against H4^a was easily dominated. These results provide a molecular mechanism for the H4 minor H antigen and suggest a novel mechanism by which alloantigenic disparity caused by conservative amino acid changes can be augmented by posttranslational antigen processing events.     

Investigation of protein export in Bifidobacterium breve UCC2003

The molecular interactions between the bifidobacterial cell and its natural environment, namely, the gastrointestinal tract of its host, are particularly important in understanding the presumed positive effects of Bifidobacterium on the health status of the host. In this study an export-specific reporter system, designed for use in gram-positive organisms and based on the use of the staphylococcal nuclease (Nuc) as a reporter, was employed to identify exported proteins in Bifidobacterium breve UCC2003. A B. breve genomic library of translational fusions to the Nuc-encoding gene devoid of its own export signal was established in the shuttle vector pFUN (I. Poquet, S. D. Ehrlich, and A. Gruss, J. Bacteriol. 180:1904-1912, 1998) and screened for bifidobacterial export signals. Sequence analysis of the fusion proteins obtained that displayed a nuclease-producing phenotype in both Lactococcus lactis and B. breve predicted the presence of a classical signal peptide and/or single or multiple transmembrane domains, thus indicating that some of the export signals in B. breve are comparable to those used in L. lactis. Cell fractionation studies, zymograms, nuclease assays, and Western blotting were employed to confirm the function of the predicted signals and to determine the location and activity of the exported fusion proteins in B. breve and/or L. lactis.     

Role for mannose-sensitive hemagglutinin in promoting interactions between Vibrio cholerae El Tor and mussel hemolymph

The role of mannose-sensitive hemagglutinin (MSHA) in Vibrio cholerae O1 El Tor interactions with hemolymph of the mussel Mytilus galloprovincialis was studied. Bacterial adherence to and association with hemocytes were evaluated at 4 and 18 degrees C, respectively. In hemolymph serum, the wild-type strain N16961 adhered to and associated with hemocytes about twofold more efficiently than its mutant lacking MSHA. In artificial seawater (ASW), no significant differences between the two strains were observed. N16961 was also more sensitive to hemocyte bactericidal activity than its MSHA mutant; in fact, the percentages of killed bacteria after 120 min of incubation were 60 and 34%, respectively. The addition of D-mannose abolished the serum-mediated increase in adherence, association, and sensitivity to killing of the wild-type strain without affecting the interactions of the mutant. A similar increase in N16961 adherence to hemocytes was observed when serum was adsorbed with MSHA-deficient bacteria. In contrast, serum adsorbed with either wild-type V. cholerae El Tor or wild-type Escherichia coli carrying type 1 fimbriae was no longer able to increase adherence of N16961 to hemocytes. The results indicate that hemolymph-soluble factors are involved in interactions between hemocytes and mannose-sensitive adhesins.     

Self-regulating model for control of replication origin firing in budding yeast

A major research area concentrates on understanding the regulation of replication origin firing. It is now appreciated that checkpoint signaling participates in this controlled process and that defects in such signaling systems affect genome integrity. Inhibition of replication origin firing is most obviously apparent under conditions of replication stress, but origin firing must also be regulated on a minute-by-minute basis as cells progress normally through an unabated S-phase. Here we summarize a straightforward model to account for how origin firing could be controlled by a self-regulating system.     

Selection of DNA aptamers that bind the RNA-dependent RNA polymerase of hepatitis C virus and inhibit viral RNA synthesis in vitro

The RNA-dependent RNA polymerase (NS5B) of the hepatitis C virus (HCV) plays a key role in the life cycle of the virus. In order to find inhibitors of the HCV polymerase, we screened a library of 81 nucleotide (nt)-long synthetic DNA containing 35 random nucleotides by the Systematic Evolution of Ligands by Exponential enrichment (SELEX) approach. Thirty ligands selected for their binding affinity to the NS5B were classified into four groups on the basis of their sequence homologies. Among the selected molecules, two were able to inhibit in vitro the polymerase activity of the HCV NS5B. These aptamers appeared to be specific for HCV polymerase, as no inhibition of poliovirus 3D polymerase activity was observed. The binding and inhibitory potential of one aptamer (27v) was associated with the 35 nt-long variable region. This oligonucleotide displayed an apparent dissociation constant (K(d)) in the nanomolar range. Our results showed that it was able to compete with RNA templates corresponding to the 3'-ends of the (+) and the (-) HCV RNA for binding to the polymerase. The fact that a DNA aptamer could interfere with the binding of natural templates of the enzyme could help in performing structure-function analysis of the NS5B and might constitute a basis for further structure-based drug design of this crucial enzyme of HCV replication.     

Supporting persons with developmental disability--a new model

The way we think about and care for people with developmental disability has changed. Twenty-five years ago, society believed that caregivers always knew what was best for their individual and that he or she must be shielded, even shut away from, the harms that could occur in society. Now, people with disability participate in all aspects of community life. They are educated in local schools, live at home or in their own home, and compete with others in the job market. Caregiving for people with developmental disability is no longer modeled on medicine or stigmatizing labels. Instead, caregivers have become support persons who focus on identifying community resources and making the environment friendlier and safer for persons with disability.