Cryo-EM structure of the CBC-ALYREF complex.

In eukaryotes, RNAs transcribed by RNA Pol II are modified at the 5' end with a 7-methylguanosine (m7G) cap, which is recognized by the nuclear cap binding complex (CBC). The CBC plays multiple important roles in mRNA metabolism, including transcription, splicing, polyadenylation, and export. It promotes mRNA export through direct interaction with a key mRNA export factor, ALYREF, which in turn links the TRanscription and EXport (TREX) complex to the 5' end of mRNA. However, the molecular mechanism for CBC-mediated recruitment of the mRNA export machinery is not well understood. Here, we present the first structure of the CBC in complex with an mRNA export factor, ALYREF. The cryo-EM structure of CBC-ALYREF reveals that the RRM domain of ALYREF makes direct contact with both the NCBP1 and NCBP2 subunits of the CBC. Comparing CBC-ALYREF with other cellular complexes containing CBC and/or ALYREF components provides insights into the coordinated events during mRNA transcription, splicing, and export.

Cryo-EM structure of the CBC-ALYREF complex.

In eukaryotes, RNAs transcribed by RNA Pol II are modified at the 5' end with a 7-methylguanosine (m 7 G) cap, which is recognized by the nuclear cap binding complex (CBC). The CBC plays multiple important roles in mRNA metabolism including transcription, splicing, polyadenylation and export. It promotes mRNA export through direct interaction with ALYREF, which in turn links the TRanscription and EXport (TREX) complex to the 5' end of mRNA. However, the molecular mechanism for CBC mediated recruitment of the mRNA export machinery is not well understood. Here, we present the first structure of the CBC in complex with a mRNA export factor, ALYREF. The cryo-EM structure of CBC-ALYREF reveals that the RRM domain of ALYREF makes direct contacts with both the NCBP1 and NCBP2 subunits of the CBC. Comparison of CBC-ALYREF to other CBC and ALYREF containing cellular complexes provides insights into the coordinated events during mRNA transcription, splicing, and export.

Structural basis for high-order complex of SARNP and DDX39B to facilitate mRNP assembly.

mRNA in eukaryotic cells is packaged into highly compacted ribonucleoprotein particles (mRNPs) in the nucleus and exported to the cytoplasm for translation. mRNP packaging and export require the evolutionarily conserved transcription-export (TREX) complex. TREX facilitates loading of various RNA-binding proteins on mRNA through the action of its DDX39B subunit. SARNP (Tho1 [transcriptional defect of Hpr1 by overexpression 1] in yeast) is shown to interact with DDX39B and affect mRNA export. The molecular mechanism of how SARNP recognizes DDX39B and functions in mRNP assembly is unclear. Here, we determine the crystal structure of a Tho1/DDX39B/RNA complex, revealing a multivalent interaction mediated by tandem DDX39B interacting motifs in SARNP/Tho1. The high-order complex of SARNP and DDX39B is evolutionarily conserved, and human SARNP can engage with five DDX39B molecules. RNA sequencing (RNA-seq) from SARNP knockdown cells shows the most affected RNAs in export are GC rich. Our work suggests the role of the high-order SARNP/DDX39B/RNA complex in mRNP assembly and export.

Cryo-EM structure of the yeast TREX complex and coordination with the SR-like protein Gbp2.

The evolutionarily conserved TRanscript-EXport (TREX) complex plays central roles during mRNP (messenger ribonucleoprotein) maturation and export from the nucleus to the cytoplasm. In yeast, TREX is composed of the THO sub-complex (Tho2, Hpr1, Tex1, Mft1, and Thp2), the DEAD box ATPase Sub2, and Yra1. Here we present a 3.7 Šcryo-EM structure of the yeast THO•Sub2 complex. The structure reveals the intimate assembly of THO revolving around its largest subunit Tho2. THO stabilizes a semi-open conformation of the Sub2 ATPase via interactions with Tho2. We show that THO interacts with the serine-arginine (SR)-like protein Gbp2 through both the RS domain and RRM domains of Gbp2. Cross-linking mass spectrometry analysis supports the extensive interactions between THO and Gbp2, further revealing that RRM domains of Gbp2 are in close proximity to the C-terminal domain of Tho2. We propose that THO serves as a landing pad to configure Gbp2 to facilitate its loading onto mRNP.

Structure and activation mechanism of the yeast RNA Pol II CTD kinase CTDK-1 complex.

The C-terminal domain (CTD) kinase I (CTDK-1) complex is the primary RNA Polymerase II (Pol II) CTD Ser2 kinase in budding yeast. CTDK-1 consists of a cyclin-dependent kinase (CDK) Ctk1, a cyclin Ctk2, and a unique subunit Ctk3 required for CTDK-1 activity. Here, we present a crystal structure of CTDK-1 at 1.85-Å resolution. The structure reveals that, compared to the canonical two-component CDK-cyclin system, the third component Ctk3 of CTDK-1 plays a critical role in Ctk1 activation by stabilizing a key element of CDK regulation, the T-loop, in an active conformation. In addition, Ctk3 contributes to the assembly of CTDK-1 through extensive interactions with both Ctk1 and Ctk2. We also demonstrate that CTDK-1 physically and genetically interacts with the serine/arginine-like protein Gbp2. Together, the data in our work reveal a regulatory mechanism of CDK complexes.

Paratracheal abscess after traumatic tracheal intubation.

Major complications of laryngoscopy and tracheal intubation are rare. However, mucosal trauma during airway management can lead to the introduction of oropharyngeal bacterial flora into the deep neck spaces, with the potential for fatal complications. This report describes the development of a paratracheal abscess in a healthy 62-year-old man following an outpatient herniorrhaphy. The patient was treated with intravenous antibiotics and underwent ultrasound-guided needle aspiration of the abscess. He was later re-admitted to the hospital with re-accumulation of the abscess, which was successfully treated by open surgical drainage. Though deep neck space infection following laryngoscopy is more common in patients with significant comorbidities and when tracheal intubation has been difficult, this case highlights the need for careful airway management in all patients.

Identifying patterns in foraging-area origins in breeding aggregations of migratory species: Loggerhead turtles in the Northwest Atlantic.

Population assessments conducted at reproductive sites of migratory species necessitate understanding the foraging-area origins of breeding individuals. Without this information, efforts to contextualize changes in breeding populations and develop effective management strategies are compromised. We used stable isotope analysis of tissue samples collected from loggerhead sea turtles (Caretta caretta) nesting at seven sites in the Northern Recovery Unit (NRU) of the eastern United States (North Carolina, South Carolina and Georgia) to assign females to three separate foraging areas in the Northwest Atlantic Ocean (NWA). We found that the majority of the females at NRU nesting sites (84.4%) use more northern foraging areas in the Mid-Atlantic Bight, while fewer females use more proximate foraging areas in the South Atlantic Bight (13.4%) and more southerly foraging areas in the Subtropical Northwest Atlantic (2.2%). We did not find significant latitudinal or temporal trends in the proportions of NRU females originating from different foraging areas. Combining these findings with previous data from stable isotope and satellite tracking studies across NWA nesting sites showed that variation in the proportion of adult loggerheads originating from different foraging areas is primarily related differences between recovery units: individuals in the NRU primarily use the Mid-Atlantic Bight foraging area, while individuals from the three Florida recovery units primarily use the Subtropical Northwest Atlantic and Eastern Gulf of Mexico foraging areas. Because each foraging area is associated with its own distinct ecological characteristics, environmental fluctuations and anthropogenic threats that affect the abundance and productivity of individuals at nesting sites, this information is critical for accurately evaluating population trends and developing effective region-specific management strategies.

Cellular mRNA export factor UAP56 recognizes nucleic acid binding site of influenza virus NP protein.

Influenza A virus nucleoprotein (NP) is a structural component that encapsulates the viral genome into the form of ribonucleoprotein complexes (vRNPs). Efficient assembly of vRNPs is critical for the virus life cycle. The assembly route from RNA-free NP to the NP-RNA polymer in vRNPs has been suggested to require a cellular factor UAP56, but the mechanism is poorly understood. Here, we characterized the interaction between NP and UAP56 using recombinant proteins and showed that UAP56 features two NP binding sites. In addition to the UAP56 core comprised of two RecA domains, we identified the N-terminal extension (NTE) of UAP56 as a previously unknown NP binding site. In particular, UAP56-NTE recognizes the nucleic acid binding region of NP. This corroborates our observation that binding of UAP56-NTE and RNA to NP is mutually exclusive. Collectively, our results reveal the molecular basis for how UAP56 acts on RNA-free NP, and provide new insights into NP-mediated influenza genome packaging.

Measurement of the parity-violating asymmetry in inclusive electroproduction of π- near the Δ0 resonance.

The parity-violating (PV) asymmetry of inclusive π- production in electron scattering from a liquid deuterium target was measured at backward angles. The measurement was conducted as a part of the G0 experiment, at a beam energy of 360 MeV. The physics process dominating pion production for these kinematics is quasifree photoproduction off the neutron via the Δ0 resonance. In the context of heavy-baryon chiral perturbation theory, this asymmetry is related to a low-energy constant d(Δ)- that characterizes the parity-violating γNΔ coupling. Zhu et al. calculated d(Δ)- in a model benchmarked by the large asymmetries seen in hyperon weak radiative decays, and predicted potentially large asymmetries for this process, ranging from A(γ)-=-5.2 to +5.2 ppm. The measurement performed in this work leads to A(γ)-=-0.36±1.06±0.37±0.03 ppm (where sources of statistical, systematic and theoretical uncertainties are included), which would disfavor enchancements considered by Zhu et al. proportional to V(ud)/V(us). The measurement is part of a program of inelastic scattering measurements that were conducted by the G0 experiment, seeking to determine the N-Δ axial transition form factors using PV electron scattering.

Transverse beam spin asymmetries at backward angles in elastic electron-proton and quasielastic electron-deuteron scattering.

We have measured the beam-normal single-spin asymmetries in elastic scattering of transversely polarized electrons from the proton, and performed the first measurement in quasielastic scattering on the deuteron, at backward angles (lab scattering angle of 108°) for Q² = 0.22 GeV²/c² and 0.63 GeV²/c² at beam energies of 362 and 687 MeV, respectively. The asymmetry arises due to the imaginary part of the interference of the two-photon exchange amplitude with that of single-photon exchange. Results for the proton are consistent with a model calculation which includes inelastic intermediate hadronic (πN) states. An estimate of the beam-normal single-spin asymmetry for the scattering from the neutron is made using a quasistatic deuterium approximation, and is also in agreement with theory.

Strange quark contributions to parity-violating asymmetries in the backward angle G0 electron scattering experiment.

We have measured parity-violating asymmetries in elastic electron-proton and quasielastic electron-deuteron scattering at Q2=0.22 and 0.63 GeV2. They are sensitive to strange quark contributions to currents in the nucleon and the nucleon axial-vector current. The results indicate strange quark contributions of approximately < 10% of the charge and magnetic nucleon form factors at these four-momentum transfers. We also present the first measurement of anapole moment effects in the axial-vector current at these four-momentum transfers.

Economic costs of cataract surgery using a rigid and a foldable intraocular lens.

Optimal delivery of healthcare requires consideration of various costs. A foldable intraocular lens (IOL) is more expensive than an equivalent rigid IOL. However, surgical and post-operative costs may make a foldable IOL economically preferable. We compared the economic costs of cataract surgery plus implantation of a foldable IOL with implantation of a rigid IOL. Prospective audit of the clinical records of 82 pseudophakes; 39 implanted with a rigid IOL and 43 implanted with a foldable IOL by one surgeon. Average follow-up periods were 25 +/- 7 months and 23 +/- 5 months respectively. There was no difference between the two groups for the follow-up period (P = 0.55), number of post-operative complications (P = 0.25) or cost of post-operative visits (P = 0.83). The cost of single-use theatre equipment was greater for the rigid-IOL group (P= 0.0001). The total identified cost per patient was greater for the foldable-IOL group (P = 0.0001). Despite possible technical advantages, implantation of the foldable IOL did not provide an economic benefit, either in the initial cost or in the costs of post-operative care. Over the 2-year period, implanting with the rigid IOL cost, on average, Pound Sterling57 less per patient. Despite this economic difference, a cost-benefit analysis is required, since other factors may be more important.

The rapid diagnosis and clinical features of human herpesvirus 6.

Since human herpesvirus 6 (HHV6) was first linked with exanthem subitum in 1988 there has been increasing evidence that the morbidity associated with acute HHV6 infection may be more significant and variable. However, the clinical appreciation of HHV6 infection has been hampered by the lack of rapid and clinically useful diagnostic methods. In this prospective study of hospitalized febrile children under 3 years of age we compared three rapid viral diagnostic methods, (polymerase chain reaction assay (PCR), IgM serology and direct antigen detection), with conventional serology on paired serum samples. In addition, we documented the range of clinical features associated with acute HHV6 infection. Of 67 children recruited, 11 (16%) had evidence of acute HHV6 infection: six had detectable, specific, IgM; four were PCR positive; and one was PCR positive with IgM. Direct antigen testing on batched frozen samples detected no infections. Apart from high fever (median peak 38.5 degrees C), common features were non-specific. Two children had febrile convulsions and only one child had a non-specific rash. We conclude that rapid microbiological diagnosis at present requires two tests (IgM and PCR). HHV6 is a common cause of febrile illness in hospitalized infants with no rash and should be considered in their diagnosis.