Messages are received both near and far, they show, to create transient and sustained responses. The cone presynaptic terminal is highly invaginated, with ribbons of glutamate-containing vesicles above each invagination. Cones respond to changes in light with graded changes in membrane potential. Decreases in light intensity depolarize cones and increase glutamate release, which then activates a class of cells known as Off bipolar cells. In the new report, DeVries et al. show that Off bipolar cell dendrites contact cone terminals at two sites. Most subtypes of Off bipolar cells contact the base of the cone terminal, 300 nm away from the vesicle fusion sites. The group found, however, that one Off cell subtype extended its dendrites up into each invagination to end close to fusion sites. These contacts within invaginations experienced large, rapid fl uctuations in glutamate levels when a cone was depolarized. Glutamate then spilled out of the invaginations to the basal contacts. In spite of their distance from release sites, even a single vesicle's worth of glutamate was able to reach and activate these cells. Distance exacted a toll, however, as the glutamate concentrations sensed by these cells fl uctuated more slowly and at much lower levels. The invaginating cell senses glutamate via AMPA receptors, which recover rapidly from glutamate-induced desensitization and can thus decode rapid consecutive pulses. Basal cells instead use kainate receptors, which have much slower recovery times and produce responses that average over rapid fl uctuations in glutamate concentration. The basally located Off bipolar cells thus generate more sustained responses. The steady signal conveys the basic sight information of change magnitude and duration. The transient signal saying just that there was a change "is probably very important," says DeVries, "because it can help an animal avoid predators or moving objects."

SUMMARY The cell wall envelope of gram-positive bacteria is a macromolecular, exoskeletal organelle that is assembled and turned over at designated sites. The cell wall also functions as a surface organelle that allows gram-positive pathogens to interact with their environment, in particular the tissues of the infected host. All of these functions require that surface proteins and enzymes be properly targeted to the cell wall envelope. Two basic mechanisms, cell wall sorting and targeting, have been identified. Cell well sorting is the covalent attachment of surface proteins to the peptidoglycan via a C-terminal sorting signal that contains a consensus LPXTG sequence. More than 100 proteins that possess cell wall-sorting signals, including the M proteins of Streptococcus pyogenes, protein A of Staphylococcus aureus, and several internalins of Listeria monocytogenes, have been identified. Cell wall targeting involves the noncovalent attachment of proteins to the cell surface via specialized binding domains. Several of these wall-binding domains appear to interact with secondary wall polymers that are associated with the peptidoglycan, for example teichoic acids and polysaccharides. Proteins that are targeted to the cell surface include muralytic enzymes such as autolysins, lysostaphin, and phage lytic enzymes. Other examples for targeted proteins are the surface S-layer proteins of bacilli and clostridia, as well as virulence factors required for the pathogenesis of L. monocytogenes (internalin B) and Streptococcus pneumoniae (PspA) infections. In this review we describe the mechanisms for both sorting and targeting of proteins to the envelope of gram-positive bacteria and review the functions of known surface proteins.

Many surface proteins are thought to be anchored to the cell wall of Gram-positive bacteria via their C-terminus. Cell wall anchoring requires a specific sorting signal, normally located at the predicted C-terminus of surface proteins. Here we show that when placed into the middle of a polypeptide chain, the sorting signal causes the specific cleavage of the precursor as well as the cell wall anchoring of its N-terminal fragment, while the C-terminal fragment remains within the cytoplasm. N-terminal sequencing of the C-terminal cleavage fragment suggests that the cleavage site is located between threonine (T) and glycine (G) of the LPXTG motif, the signature sequence of cell wall sorting signals. All surface proteins harbouring an LPXTG sequence motif may therefore be cleaved and anchored by a universal mechanism. We also propose a novel hypothesis for the cell wall linkage of surface proteins in Gram-positive bacteria.

H-NS and Lsr2 are nucleoid-associated proteins from Gram-negative bacteria and Mycobacteria , respectively, that play an important role in the silencing of horizontally acquired foreign DNA that is more AT-rich than the resident genome. Despite the fact that Lsr2 and H-NS proteins are dissimilar in sequence and structure, they serve apparently similar functions and can functionally complement one another. The mechanism by which these xenogeneic silencers selectively target AT-rich DNA has been enigmatic. We performed high-resolution protein binding microarray analysis to simultaneously assess the binding preference of H-NS and Lsr2 for all possible 8-base sequences. Concurrently, we performed a detailed structure-function relationship analysis of their C-terminal DNA binding domains by NMR. Unexpectedly, we found that H-NS and Lsr2 use a common DNA binding mechanism where a short loop containing a “Q/RGR” motif selectively interacts with the DNA minor groove, where the highest affinity is for AT-rich sequences that lack A-tracts. Mutations of the Q/RGR motif abolished DNA binding activity. Netropsin, a DNA minor groove-binding molecule effectively outcompeted H-NS and Lsr2 for binding to AT-rich sequences. These results provide a unified molecular mechanism to explain findings related to xenogeneic silencing proteins, including their lack of apparent sequence specificity but preference for AT-rich sequences. Our findings also suggest that structural information contained within the DNA minor groove is deciphered by xenogeneic silencing proteins to distinguish genetic material that is self from nonself.