The T-cell leukemia related rpl10-R98S mutant traps the 60S export adapter Nmd3 in the ribosomal P site in yeast

Mutations in the ribosomal protein Rpl10 (uL16) can be drivers of T-cell acute lymphoblastic leukemia (T-ALL). We previously showed that these T-ALL mutations disrupt late cytoplasmic maturation of the 60S ribosomal subunit, blocking the release of the trans-acting factors Nmd3 and Tif6 in S. cerevisiae. Consequently, these mutant ribosomes do not efficiently pass the cytoplasmic quality control checkpoint and are blocked from engaging in translation. Here, we characterize suppressing mutations of the T-ALL-related rpl10-R98S mutant that bypass this block and show that the molecular defect of rpl10-R98S is a failure to release Nmd3 from the P site. Suppressing mutations were identified in Nmd3 and Tif6 that disrupted interactions between Nmd3 and the ribosome, or between Nmd3 and Tif6. Using an in vitro system with purified components, we found that Nmd3 inhibited Sdo1-stimulated Efl1 activity on mutant rpl10-R98S but not wild-type 60S subunits. Importantly, this inhibition was overcome in vitro by mutations in Nmd3 that suppressed rpl10-R98S in vivo. These results strongly support a model that Nmd3 must be dislodged from the P site to allow Sdo1 activation of Efl1, and define a failure in the removal of Nmd3 as the molecular defect of the T-ALL-associated rpl10-R98S mutation

The structures of secretory and dimeric immunoglobulin A

Secretory (S) Immunoglobulin (Ig) A is the predominant mucosal antibody, which binds pathogens and commensal microbes. SIgA is a polymeric antibody, typically containing two copies of IgA that assemble with one joining-chain (JC) to form dimeric (d) IgA that is bound by the polymeric Ig-receptor ectodomain, called secretory component (SC). Here, we report the cryo-electron microscopy structures of murine SIgA and dIgA. Structures reveal two IgAs conjoined through four heavy-chain tailpieces and the JC that together form a β-sandwich-like fold. The two IgAs are bent and tilted with respect to each other, forming distinct concave and convex surfaces. In SIgA, SC is bound to one face, asymmetrically contacting both IgAs and JC. The bent and tilted arrangement of complex components limits the possible positions of both sets of antigen-binding fragments (Fabs) and preserves steric accessibility to receptor-binding sites, likely influencing antigen binding and effector functions

The structures of secretory and dimeric immunoglobulin A

Secretory (S) Immunoglobulin (Ig) A is the predominant mucosal antibody, which binds pathogens and commensal microbes. SIgA is a polymeric antibody, typically containing two copies of IgA that assemble with one joining-chain (JC) to form dimeric (d) IgA that is bound by the polymeric Ig-receptor ectodomain, called secretory component (SC). Here, we report the cryo-electron microscopy structures of murine SIgA and dIgA. Structures reveal two IgAs conjoined through four heavy-chain tailpieces and the JC that together form a β-sandwich-like fold. The two IgAs are bent and tilted with respect to each other, forming distinct concave and convex surfaces. In SIgA, SC is bound to one face, asymmetrically contacting both IgAs and JC. The bent and tilted arrangement of complex components limits the possible positions of both sets of antigen-binding fragments (Fabs) and preserves steric accessibility to receptor-binding sites, likely influencing antigen binding and effector functions

Characterization of Reactive Organometallic Species via MicroED

Here we apply microcrystal electron diffraction (MicroED) to the structural determination of transition-metal complexes. We find that the simultaneous use of 300 keV electrons, very low electron doses, and an ultrasensitive camera allows for the collection of data without cryogenic cooling of the stage. This technique reveals the first crystal structures of the classic zirconocene hydride, colloquially known as “Schwartz’s reagent”, a novel Pd(II) complex not amenable to solution-state NMR or X-ray crystallography, and five other paramagnetic and diamagnetic transition-metal complexes

Characterization of Reactive Organometallic Species via MicroED

Here we apply microcrystal electron diffraction (MicroED) to the structural determination of transition-metal complexes. We find that the simultaneous use of 300 keV electrons, very low electron doses, and an ultrasensitive camera allows for the collection of data without cryogenic cooling of the stage. This technique reveals the first crystal structures of the classic zirconocene hydride, colloquially known as “Schwartz’s reagent”, a novel Pd(II) complex not amenable to solution-state NMR or X-ray crystallography, and five other paramagnetic and diamagnetic transition-metal complexes

On a new genus and species of midaliid octocoral (Coelenterata: Octocorallia) from the South China Sea

Volume: 10Start Page: 41End Page: 4

The structure of the teleost Immunoglobulin M core provides insights on polymeric antibody evolution, assembly, and function

Abstract Polymeric (p) immunoglobulins (Igs) serve broad functions during vertebrate immune responses. Typically, pIgs contain between two and six Ig monomers, each with two antigen binding fragments and one fragment crystallization (Fc). In addition, many pIgs assemble with a joining-chain (JC); however, the number of monomers and potential to include JC vary with species and heavy chain class. Here, we report the cryo-electron microscopy structure of IgM from a teleost (t) species, which does not encode JC. The structure reveals four tIgM Fcs linked through eight C-terminal tailpieces (Tps), which adopt a single β-sandwich-like domain (Tp assembly) located between two Fcs. Specifically, two of eight heavy chains fold uniquely, resulting in a structure distinct from mammalian IgM, which typically contains five IgM monomers, one JC and a centrally-located Tp assembly. Together with mutational analysis, structural data indicate that pIgs have evolved a range of assembly mechanisms and structures, each likely to support unique antibody effector functions

Budding Pathway in the Templated Assembly of Viruslike Particles

A new pathway for the assembly of viral capsid protein around inorganic nanoparticle cores was observed by time-course light scattering and cryo-electron tomography. Gold nanoparticles with an average diameter of 11.3 nm have been used as a template for the assembly of Brome mosaic virus (BMV) capsid protein at different concentrations. At least at low protein concentrations the kinetic features of the scattering and extinction measurements are consistent with the initial rapid formation of large nanoparticle–protein clusters, which subsequently separate into individual viruslike particles (VLPs). The occurrence of multiparticle clusters at short times after mixing nanoparticles and proteins was confirmed by cryo-EM. Cryo-electron tomography of the multiparticle clusters yielded an average surface-to-surface interparticle distance of ∼7.5 nm, equivalent to ∼1.5 times the thickness of a protein shell. We propose a scenario in which VLP generation may take place through monomer exchange between aggregated particles with defect-ridden or incomplete shells, leading to the formation of stable icosahedral shells, which eventually bud off the aggregate. Together with results from previous works, the findings highlight the astonishing versatility of plant virus capsid protein assembly. This previously unknown mechanism for VLP formation has features that may have relevance for the crowded environment characterizing virus factories in the cell