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1. Reconstruction of PtERV1 p12 and CA and infectivity of PtERV1/MLV chimeric virus. (A) An alignment of the p12 and CA sequences of the consensus PtERV1 from chimpanzee (top line) and MLV (bottom line) is shown. A late domain (PPPY) characteristic for retroviruses is in p12 and the Major Homology Region (MHR) conserved in all retroviruses is present in CA are indicated by shaded boxes. ( An unrooted phylogenetic NJ tree was generated with nucleotide sequences of PtERV1 (MA through PRO) from chimpanzee, rhesus, and gorilla genomes. Branches are color coded: gorilla, orange; chimpanzee, blue; and rhesus, green. Fifteen amino acid changes were introduced into one extant copy of PtERV1 p12-CA from chimpanzees to generate the "ancestral" clone (fig. S1). The derived consensus sequence from chimpanzee is denoted by a black branch. The nucleotide consensus is slightly displaced from the node of the monophyletic clade containing chimp and gorilla sequences because synonymous changes were not introduced during the reconstruction process. (C) Schematic representation of the chimeric virus between MLV and PtERV1 is shown. The Gag-Pol expression vector used to make virus contains the MA, NC, and the entire POL coding sequence of MLV and the p12 and CA from PtERV1. Protease cleavage sites between MLV MA and PtERV1 p12, as well as between PtERV1 CA and MLV NC, were generated to match cleavage sites recognized by MLV protease. (D) Green fluorescent protein (GFP)–encoding virus was used to infect Crandell-Rees feline kidney (CRFK) cells with PtERV1 chimeric virions. Fluorescence-activated cell sorting (FACS) plots of virus-challenged cells are shown. The y axis indicates cell size, and the x axis denotes GFP positivity, which is an indicator of transduction. The initial PtERV1 clone used to reconstruct this consensus was tested in the chimeric context and was noninfectious (left). After 15 mutations were introduced to reconstruct the consensus p12 and CA sequence, infectious virus particles were generated (right).
原文出处：
Science 22 June 2007 Vol 316, Issue 5832
《生命科学研究》杂志网址：http://smky.chinajournal.net.cn
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作者:admin@医学,生命科学 2011-02-27 17:11
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