ABSTRACT
Lung hernia is an uncommon diagnosis characterized by lung tissue protruding through a chest wall defect. It may occur spontaneously, as a congenital defect, as a result of trauma, or as a postsurgical complication. We describe the occurrence of lung hernia and subsequent successful herniorraphy in two patients following robotic-assisted mitral valve repair.
Subject(s)
Hernia/etiology , Lung Diseases/etiology , Mitral Valve Annuloplasty/methods , Mitral Valve Prolapse/surgery , Postoperative Complications , Robotics , Hernia/diagnosis , Herniorrhaphy/methods , Humans , Lung Diseases/diagnosis , Lung Diseases/surgery , Male , Middle Aged , Postoperative Complications/diagnosis , Postoperative Complications/surgeryABSTRACT
The 5'-untranslated regions of all gammaretroviruses contain a conserved "double-hairpin motif" (Ψ(CD)) that is required for genome packaging. Both hairpins (SL-C and SL-D) contain GACG tetraloops that, in isolated RNAs, are capable of forming "kissing" interactions stabilized by two intermolecular G-C base pairs. We have determined the three-dimensional structure of the double hairpin from the Moloney murine leukemia virus ([Ψ(CD)](2), 132 nt, 42.8 kDa) using a (2)H-edited NMR-spectroscopy-based approach. This approach enabled the detection of (1)H-(1)H dipolar interactions that were not observed in previous studies of isolated SL-C and SL-D hairpin RNAs using traditional (1)H-(1)H correlated and (1)H-(13)C-edited NMR methods. The hairpins participate in intermolecular cross-kissing interactions (SL-C to SL-D' and SLC' to SL-D) and stack in an end-to-end manner (SL-C to SL-D and SL-C' to SL-D') that gives rise to an elongated overall shape (ca 95 Å×45 Å×25 Å). The global structure was confirmed by cryo-electron tomography (cryo-ET), making [Ψ(CD)](2) simultaneously the smallest RNA to be structurally characterized to date by cryo-ET and among the largest to be determined by NMR. Our findings suggest that, in addition to promoting dimerization, [Ψ(CD)](2) functions as a scaffold that helps initiate virus assembly by exposing a cluster of conserved UCUG elements for binding to the cognate nucleocapsid domains of assembling viral Gag proteins.
Subject(s)
5' Untranslated Regions , Moloney murine leukemia virus/chemistry , RNA, Viral/chemistry , RNA, Viral/metabolism , Virus Assembly , Animals , Cryoelectron Microscopy , Dimerization , Electron Microscope Tomography , Magnetic Resonance Spectroscopy , Models, Biological , Models, Molecular , Moloney murine leukemia virus/physiology , Nucleic Acid ConformationABSTRACT
Retroviruses selectively package two copies of their RNA genomes via mechanisms that have yet to be fully deciphered. Recent studies with small fragments of the Moloney murine leukemia virus (MoMuLV) genome suggested that selection may be mediated by an RNA switch mechanism, in which conserved UCUG elements that are sequestered by base-pairing in the monomeric RNA become exposed upon dimerization to allow binding to the cognate nucleocapsid (NC) domains of the viral Gag proteins. Here we show that a large fragment of the MoMuLV 5' untranslated region that contains all residues necessary for efficient RNA packaging (Psi(WT); residues 147-623) also exhibits a dimerization-dependent affinity for NC, with the native dimer ([Psi(WT)](2)) binding 12+/-2 NC molecules with high affinity (K(d)=17+/-7 nM) and with the monomer, stabilized by substitution of dimer-promoting loop residues with hairpin-stabilizing sequences (Psi(M)), binding 1-2 NC molecules. Identical dimer-inhibiting mutations in MoMuLV-based vectors significantly inhibit genome packaging in vivo (approximately 100-fold decrease), whereas a large deletion of nearly 200 nucleotides just upstream of the gag start codon has minimal effects. Our findings support the proposed RNA switch mechanism and further suggest that virus assembly may be initiated by a complex comprising as few as 12 Gag molecules bound to a dimeric packaging signal.