Simian retrovirus serogroup 2 constitutive transport element recognizes the ribosomal L10-like protein and translocon gamma subunit-like protein in a yeast three-hybrid assay

Biao Li, Xiaorong Li, Ying Bai, Jing Wen Hou, Mark Ma, Curtis Machida

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

The simian retrovirus (SRV) serogroup 2 genome contains a constitutive transport element (CTE) within its 3′ intergenic region (IR) that mediates the nuclear export of unspliced SRV RNA. In a previous report [Virology 264 (1999) 37], CTE RNA-protein complexes were detected using UV-crosslinking/sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). To identify these CTE-interacting cellular proteins, we utilized yeast three-hybrid interaction approaches using the complete IR as bait, modified to eliminate transcriptional termination signals recognized by RNA polymerase III, and identified several interactive clones from a Hela cell cDNA activation domain (AD) library. UV-crosslinking of RNA-protein complexes, using Hela cell extracts and the modified IR bait, were conducted prior to library screening, to verify appropriate interaction of CTE RNA-protein complexes. Over one million recombinants were screened, and our yeast hybrid results indicate that the CTE interacts with several molecules involved in cellular translational and translocation machinery, including the ribosomal L10-like protein and the tranlocon protein gamma subunit-like protein. UV-crosslinking/immunoblot assays have verified the interaction of the CTE region with molecules immunologically reactive to antibodies recognizing the ribosomal L10-like protein.

Original languageEnglish (US)
Pages (from-to)69-80
Number of pages12
JournalVirus Research
Volume99
Issue number1
DOIs
StatePublished - Jan 2004

Fingerprint

Mason-Pfizer monkey virus
Two-Hybrid System Techniques
Protein Subunits
Intergenic DNA
RNA Transport
HeLa Cells
Simian Retroviruses
Proteins
RNA
RNA Polymerase III
Virology
Fungal Proteins
Cell Nucleus Active Transport
Cell Extracts
Sodium Dodecyl Sulfate
Libraries
Polyacrylamide Gel Electrophoresis
Complementary DNA
Clone Cells
Yeasts

Keywords

  • Constitutive transport element
  • Ribosomal proteins
  • Secondary structural analyses
  • Simian retrovirus
  • Translation
  • UV-crosslinking/immunoblot experiments
  • Yeast three-hybrid interaction

ASJC Scopus subject areas

  • Cancer Research
  • Molecular Biology
  • Virology

Cite this

Simian retrovirus serogroup 2 constitutive transport element recognizes the ribosomal L10-like protein and translocon gamma subunit-like protein in a yeast three-hybrid assay. / Li, Biao; Li, Xiaorong; Bai, Ying; Hou, Jing Wen; Ma, Mark; Machida, Curtis.

In: Virus Research, Vol. 99, No. 1, 01.2004, p. 69-80.

Research output: Contribution to journalArticle

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