Insights into centriole geometry revealed by cryotomography of doublet and triplet centrioles.

Centrioles are cylindrical assemblies comprised of 9 singlet, doublet, or triplet microtubules, essential for the formation of motile and sensory cilia. While the structure of the cilium is being defined at increasing resolution, centriolar structure remains poorly understood. Here, we used electron cryo-tomography to determine the structure of mammalian (triplet) and Drosophila (doublet) centrioles. Mammalian centrioles have two distinct domains: a 200 nm proximal core region connected by A-C linkers, and a distal domain where the C-tubule is incomplete and a pair of novel linkages stabilize the assembly producing a geometry more closely resembling the ciliary axoneme. Drosophila centrioles resemble the mammalian core, but with their doublet microtubules linked through the A tubules. The commonality of core-region length, and the abrupt transition in mammalian centrioles, suggests a conserved length-setting mechanism. The unexpected linker diversity suggests how unique centriolar architectures arise in different tissues and organisms

Longevity and Highest-Energy Cosmic Rays

It is proposed that the highest energy $\sim 10^{20}$eV cosmic ray primaries are protons, decay products of a long-lived progenitor which has propagated from typically $\sim 100$Mpc. Such a scenario can occur in e.g. SU(15) grand unification and in some preon models, but is more generic; if true, these unusual cosmic rays provide a window into new physics.Comment: 8 pages, RevTe

NEUTRINOS FROM PRIMORDIAL BLACK HOLES

The emission of particles from black holes created in the early Universe has detectable astrophysical consequences. The most stringent bound on their abundance has been obtained from the absence of a detectable diffuse flux of 100 MeV photons. Further scrutiny of these bounds is of interest as they, for instance, rule out primordial black holes as a dark matter candidate. We here point out that these bounds can, in principle, be improved by studying the diffuse cosmic neutrino flux. Measurements of near-vertical atmospheric neutrino fluxes in a region of low geomagnetic latitude can provide a competitive bound. The most favorable energy to detect a possible diffuse flux of primordial black hole origin is found to be a few MeV. We also show that measurements of the diffuse $\nu _\tau$ flux is the most promising to improve the existing bounds deduced from gamma-ray measurements. Neutrinos from individual black hole explosions can be detected in the GeV-TeV energy region. We find that the kilometer-scale detectors, recently proposed, are able to establish competitive bounds.Comment: 19 pages plus 9 uuencoded and compressed postscript figure

The centrosomin CM2 domain is a multi-functional binding domain with distinct cell cycle roles.

The centrosome serves as the main microtubule-organizing center in metazoan cells, yet despite its functional importance, little is known mechanistically about the structure and organizational principles that dictate protein organization in the centrosome. In particular, the protein-protein interactions that allow for the massive structural transition between the tightly organized interphase centrosome and the highly expanded matrix-like arrangement of the mitotic centrosome have been largely uncharacterized. Among the proteins that undergo a major transition is the Drosophila melanogaster protein centrosomin that contains a conserved carboxyl terminus motif, CM2. Recent crystal structures have shown this motif to be dimeric and capable of forming an intramolecular interaction with a central region of centrosomin. Here we use a combination of in-cell microscopy and in vitro oligomer assessment to show that dimerization is not necessary for CM2 recruitment to the centrosome and that CM2 alone undergoes significant cell cycle dependent rearrangement. We use NMR binding assays to confirm this intramolecular interaction and show that residues involved in solution are consistent with the published crystal structure and identify L1137 as critical for binding. Additionally, we show for the first time an in vitro interaction of CM2 with the Drosophila pericentrin-like-protein that exploits the same set of residues as the intramolecular interaction. Furthermore, NMR experiments reveal a calcium sensitive interaction between CM2 and calmodulin. Although unexpected because of sequence divergence, this suggests that centrosomin-mediated assemblies, like the mammalian pericentrin, may be calcium regulated. From these results, we suggest an expanded model where during interphase CM2 interacts with pericentrin-like-protein to form a layer of centrosomin around the centriole wall and that at the onset of mitosis this population acts as a nucleation site of intramolecular centrosomin interactions that support the expansion into the metaphase matrix

The centrosomin CM2 domain is a multi-functional binding domain with distinct cell cycle roles.

The centrosome serves as the main microtubule-organizing center in metazoan cells, yet despite its functional importance, little is known mechanistically about the structure and organizational principles that dictate protein organization in the centrosome. In particular, the protein-protein interactions that allow for the massive structural transition between the tightly organized interphase centrosome and the highly expanded matrix-like arrangement of the mitotic centrosome have been largely uncharacterized. Among the proteins that undergo a major transition is the Drosophila melanogaster protein centrosomin that contains a conserved carboxyl terminus motif, CM2. Recent crystal structures have shown this motif to be dimeric and capable of forming an intramolecular interaction with a central region of centrosomin. Here we use a combination of in-cell microscopy and in vitro oligomer assessment to show that dimerization is not necessary for CM2 recruitment to the centrosome and that CM2 alone undergoes significant cell cycle dependent rearrangement. We use NMR binding assays to confirm this intramolecular interaction and show that residues involved in solution are consistent with the published crystal structure and identify L1137 as critical for binding. Additionally, we show for the first time an in vitro interaction of CM2 with the Drosophila pericentrin-like-protein that exploits the same set of residues as the intramolecular interaction. Furthermore, NMR experiments reveal a calcium sensitive interaction between CM2 and calmodulin. Although unexpected because of sequence divergence, this suggests that centrosomin-mediated assemblies, like the mammalian pericentrin, may be calcium regulated. From these results, we suggest an expanded model where during interphase CM2 interacts with pericentrin-like-protein to form a layer of centrosomin around the centriole wall and that at the onset of mitosis this population acts as a nucleation site of intramolecular centrosomin interactions that support the expansion into the metaphase matrix

Conserved domain, CM2, of centrosomin characterized by SEC-MALS, SIM, and yeast-two-hybrid experiments.

<p>(A) Schematic of Centrosomin with coiled-coil prediction shown in blue. CM1 and CM2 are highlighted in red. Alignment of <i>D</i>. <i>Melanogaster’s</i> CM2 with the zebra fish, mouse and human (CDK5RAP2) homologues, with residue numbering according to <i>D</i>. <i>Melanogaster</i> residues. Higher conservation can be seen starting at residue E1090 in line with the start of the predicted coiled coil region. The predicted calmodulin binding site in human CDK5RAP2 is shown in red. (B) Molecular weights (left axis) measured with SEC-MALS are consistent with aa1064-1148 forming a dimer of CM2 (red) with a predicted monomer weight of 11kDa and a calculated weight of 22.4kDa. The construct aa1090-1148 exists as monomer (blue) with a predicted monomer weight of 7kDa and a calculated weight of 7.7kDa. (C) SEC-MALS of a more minimal dimer construct aa1074-1148 (blue) that has a calculated molecular weight of 19.4kDa and predicted monomer weight of 9kDa. A construct with the final 18 residues truncated (Δ1130–1148) maintains a dimeric oligomer state (red) with a calculated molecular weight of 21.6kDa and predicted monomer weight of 8.8kDA. (D) Z-projections of the aligned and averaged PLP and GFP distributions for both the monomer and dimer GFP-CM2 fusion constructs in interphase S2 cells. Scale bar represents 100nm. (E) Z-projections of the monomer and dimer GFP-CM2 fusion constructs in mitotic S2 cells. Scale bar represents 100nm (F) Radial averages of the data represented in figures D and E with vertical bars indicating the average and horizontal bars representing the standard deviation of the Gaussian fit. (G) Schematic summaries of yeast-two-hybrid experiments showing the CM2 construct (aa1087-1148) interacting with a minimal domain of PLP(aa583-740) and the middle domain of CNN (aa454-556). Yeast plates of the interaction test are shown with red boxes indicating plating on selection media.</p