Slits are large multidomain leucine-rich repeat (LRR)-containing proteins that provide crucial guidance cues in neuronal and vascular development. More recently, Slits have been implicated in heart morphogenesis, angiogenesis, and tumor metastasis. Slits are ligands for the Robo (Roundabout) receptors, which belong to the Ig superfamily of transmembrane signaling molecules. The Slit-Robo interaction is mediated by the second LRR domain of Slit and the two N-terminal Ig domains of Robo, but the molecular details of this interaction and how it induces signaling remain unclear. Here we describe the crystal structures of the second LRR domain of human Slit2 (Slit2 D2), the first two Ig domains of its receptor Robo1 (Ig1-2), and the minimal complex between these proteins (Slit2 D2-Robo1 Ig1). Slit2 D2 binds with its concave surface to the side of Ig1 with electrostatic and hydrophobic contact regions mediated by residues that are conserved in other family members. Surface plasmon resonance experiments and a mutational analysis of the interface confirm that Ig1 is the primary domain for binding Slit2. These structures provide molecular insight into Slit-Robo complex formation and will be important for the development of novel cancer therapeutics.guidance cues ͉ neurons ͉ signaling B ilaterally symmetric nervous systems, such as those of insects and vertebrates, possess a special midline structure that establishes a partition between the left and right mirror-image halves. To connect and coordinate both sides, a subset of so-called commissural axons has to cross the midline. Developing commissural axons navigate through the embryo by processing and responding to a number of different signals in their immediate environment. Both Slit and Netrin and their receptors, Roundabout (Robo) and Deleted in Colorectal Carcinoma (DCC), provide key ligand-receptor interactions for this process during neuronal development, especially at the midline of the central nervous system of vertebrates and invertebrates (1, 2). The Slit-Robo signaling complex is also central to the development of blood vessels (3, 4) and some organs, for example, the heart (5, 6). In addition, Slit2 has been implicated in breast cancer metastasis (7) and Robo1 in heptacellular carcinoma (8).Three Slit proteins (Slit1-3) (9) and four Robo proteins (Robo1, Robo2, Robo3/Rig-1, and the vascular-specific Robo4/ magic Roundabout) (10-12) have been identified in mammals. Netrin and Slit1-3 are secreted by the midline cells, whereas DCC and Robo1-3 are expressed on the surface of growing axons (2). During neuronal development, the commissural axons are initially attracted to the midline through a Netrin-DCCmediated interaction (2). This attractive signal is subsequently silenced near the midline to allow crossing through a Slitmediated interaction between DCC and Robo (13). Slit-Robo signaling then induces repulsion, expelling the axons from the midline and preventing recrossing (1). In vertebrates, Robo3 is also proposed to have a role in midline crossing by antagon...

Sporulation in Bacillus subtilis involves two cells that follow separate but coordinately regulated developmental programs. Late in sporulation, the developing spore (the forespore) resides within a mother cell. The regulation of the forespore transcription factor σG that acts at this stage has remained enigmatic. σG activity requires eight mother-cell proteins encoded in the spoIIIA operon and the forespore protein SpoIIQ. Several of the SpoIIIA proteins share similarity with components of specialized secretion systems. One of them resembles a secretion ATPase and we demonstrate that the ATPase motifs are required for σG activity. We further show that the SpoIIIA proteins and SpoIIQ reside in a multimeric complex that spans the two membranes surrounding the forespore. Finally, we have discovered that these proteins are all required to maintain forespore integrity. In their absence, the forespore develops large invaginations and collapses. Importantly, maintenance of forespore integrity does not require σG. These results support a model in which the SpoIIIA-SpoIIQ proteins form a novel secretion apparatus that allows the mother cell to nurture the forespore, thereby maintaining forespore physiology and σG activity during spore maturation.

SummaryThe bacterial peptidoglycan, the main component of the cell wall, is synthesized by the penicillin-binding proteins (PBPs). We used immunofluorescence microscopy to determine the cellular localization of all the high molecular weight PBPs of the human pathogen Streptococcus pneumoniae , for a wild type and for several PBP-deficient strains. Progression through the cell cycle was investigated by the simultaneous labelling of DNA and the FtsZ protein. Our main findings are: (i) the temporal dissociation of cell wall synthesis, inferred by the localization of PBP2x and PBP1a, from the constriction of the FtsZ-ring; (ii) the localization of PBP2b and PBP2a at duplicated equatorial sites indicating the existence of peripheral peptidoglycan synthesis, which implies a similarity between the mechanism of cell division in bacilli and streptococci; (iii) the abnormal localization of some class A PBPs in PBP-defective mutants which may explain the apparent redundancy of these proteins in S. pneumoniae .