The initial stage of invasion by apicomplexan parasites involves the exocytosis of the micronemes-containing molecules that contribute to host cell attachment and penetration. MIC4 was previously described as a protein secreted by Toxoplasma gondii tachyzoites upon stimulation of micronemes exocytosis. We have microsequenced the mature protein, purified after discharge from micronemes and cloned the corresponding gene. The deduced amino acid sequence of MIC4 predicts a 61-kDa protein that contains 6 conserved apple domains. Apple domains are composed of six spacely conserved cysteine residues which form disulfide bridges and are also present in micronemal proteins from two closely related apicomplexan parasites, Sarcocystis muris and Eimeria species, and several mammalian serum proteins, including kallikrein. Here we show that MIC4 localizes in the micronemes of all the invasive forms of T. gondii, tachyzoites, bradyzoites, sporozoites, and merozoites. The protein is proteolytically processed both at the N and the C terminus only upon release from the organelle. MIC4 binds efficiently to host cells, and the adhesive motif maps in the most C-terminal apple domain.

Cells use γ-tubulin complex to nucleate microtubules. The assembly of active microtubule nucleator is spatially and temporally regulated through the cell cycle. Lin et al. show that the protein Mzt1/MOZART1 and γ-tubulin complex receptors directly interact and act together to assemble the γ-tubulin small complex into an active microtubule nucleation template and that such interaction is conserved between Candida albicans and human cells.

Most transport pathways between cell nucleus and cytoplasm are mediated by nuclear transport receptors of the importin b family. These receptors are in continuous circulation between the two compartments and transfer cargo molecules from one side of the nuclear envelope to the other. RanBP16 is a family member from higher eukaryotes of so far unknown function. We now show that it exports p50RhoGAP from the nucleus and thereby confines this activity to the cytoplasm. It also accounts for nuclear exclusion of 14-3-3r, which in turn is known to anchor, for example, cyclin-dependent kinases in the cytoplasm. Our data further suggest that RanBP16 exports several additional cargoes. It thus appears to be a nuclear export mediator with broad substrate specificity and we will therefore refer to it as exportin 7 (Exp7). Finally, we demonstrate that Exp7-dependent nuclear export signals differ fundamentally from the leucine-rich, CRM1-dependent ones: First, they are not just short linear sequences, but instead include folded motifs. Second, basic residues are critical for Exp7 recruitment.

Stu2/XMAP215/ZYG-9/Dis1/Alp14/Msps/ch-TOG family members in association with with γ-tubulin complexes nucleate microtubules, but we know little about the interplay of these nucleation factors. Here, we show that the budding yeast Stu2 in complex with the γ-tubulin receptor Spc72 nucleates microtubules in vitro without the small γ-tubulin complex (γ-TuSC). Upon γ-TuSC addition, Stu2 facilitates Spc72–γ-TuSC interaction by binding to Spc72 and γ-TuSC. Stu2 together with Spc72–γ-TuSC increases microtubule nucleation in a process that is dependent on the TOG domains of Stu2. Importantly, these activities are also important for microtubule nucleation in vivo. Stu2 stabilizes Spc72–γ-TuSC at the minus end of cytoplasmic microtubules (cMTs) and an in vivo assay indicates that cMT nucleation requires the TOG domains of Stu2. Upon γ-tubulin depletion, we observed efficient cMT nucleation away from the spindle pole body (SPB), which was dependent on Stu2. Thus, γ-TuSC restricts cMT assembly to the SPB whereas Stu2 nucleates cMTs together with γ-TuSC and stabilizes γ-TuSC at the cMT minus end.