Identification of the minimal essential RNA sequences responsible for site-specific targeting of the Leishmania RNA virus 1-4 capsid endoribonuclease

Young Tae Ro, Jean L. Patterson

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

The Leishmania RNA virus 1-4 capsid protein possesses an endoribonuclease activity responsible for single-site-specific cleavage within the 450-nucleotide 5' untranslated region of its own viral RNA transcript. To characterize the minimal essential RNA determinants required for site-specific cleavage, mutated RNA transcripts were examined for susceptibility to cleavage by the virus capsid protein in an in vitro assay. Deletion analyses revealed that all determinants necessary for accurate cleavage are encoded in viral nucleotides 249 to 342. Nuclease mapping and site-specific mutagenesis of the minimal RNA sequence defined a stem-loop structure that is located 40 nucleotides upstream from the cleavage site (nucleotide 320) and that is essential for accurate RNA cleavage. Abrogation of cleavage by disruption of base pairing within the stem-loop was reversed through the introduction of complementary nucleotide substitutions that reestablished the structure. We also provide evidence that divalent cations, essential components of the cleavage reaction, stabilized the stem-loop structure in solution. That capsid-specific antiserum eliminated specific RNA cleavage provides further evidence that the virus capsid gene encodes the essential endoribonuclease activity.

Original languageEnglish (US)
Pages (from-to)130-138
Number of pages9
JournalJournal of Virology
Volume74
Issue number1
StatePublished - 2000
Externally publishedYes

Fingerprint

endoribonucleases
Endoribonucleases
capsid
Capsid
Leishmania
RNA Viruses
RNA Cleavage
Nucleotides
nucleotides
nucleotide sequences
Capsid Proteins
RNA
messenger RNA
coat proteins
stems
Viruses
viruses
nucleases
5' Untranslated Regions
Essential Genes

ASJC Scopus subject areas

  • Immunology

Cite this

Identification of the minimal essential RNA sequences responsible for site-specific targeting of the Leishmania RNA virus 1-4 capsid endoribonuclease. / Ro, Young Tae; Patterson, Jean L.

In: Journal of Virology, Vol. 74, No. 1, 2000, p. 130-138.

Research output: Contribution to journalArticle

@article{5bba323400bc447b99221b9e898548ee,
title = "Identification of the minimal essential RNA sequences responsible for site-specific targeting of the Leishmania RNA virus 1-4 capsid endoribonuclease",
abstract = "The Leishmania RNA virus 1-4 capsid protein possesses an endoribonuclease activity responsible for single-site-specific cleavage within the 450-nucleotide 5' untranslated region of its own viral RNA transcript. To characterize the minimal essential RNA determinants required for site-specific cleavage, mutated RNA transcripts were examined for susceptibility to cleavage by the virus capsid protein in an in vitro assay. Deletion analyses revealed that all determinants necessary for accurate cleavage are encoded in viral nucleotides 249 to 342. Nuclease mapping and site-specific mutagenesis of the minimal RNA sequence defined a stem-loop structure that is located 40 nucleotides upstream from the cleavage site (nucleotide 320) and that is essential for accurate RNA cleavage. Abrogation of cleavage by disruption of base pairing within the stem-loop was reversed through the introduction of complementary nucleotide substitutions that reestablished the structure. We also provide evidence that divalent cations, essential components of the cleavage reaction, stabilized the stem-loop structure in solution. That capsid-specific antiserum eliminated specific RNA cleavage provides further evidence that the virus capsid gene encodes the essential endoribonuclease activity.",
author = "Ro, {Young Tae} and Patterson, {Jean L.}",
year = "2000",
language = "English (US)",
volume = "74",
pages = "130--138",
journal = "Journal of Virology",
issn = "0022-538X",
publisher = "American Society for Microbiology",
number = "1",

}

TY - JOUR

T1 - Identification of the minimal essential RNA sequences responsible for site-specific targeting of the Leishmania RNA virus 1-4 capsid endoribonuclease

AU - Ro, Young Tae

AU - Patterson, Jean L.

PY - 2000

Y1 - 2000

N2 - The Leishmania RNA virus 1-4 capsid protein possesses an endoribonuclease activity responsible for single-site-specific cleavage within the 450-nucleotide 5' untranslated region of its own viral RNA transcript. To characterize the minimal essential RNA determinants required for site-specific cleavage, mutated RNA transcripts were examined for susceptibility to cleavage by the virus capsid protein in an in vitro assay. Deletion analyses revealed that all determinants necessary for accurate cleavage are encoded in viral nucleotides 249 to 342. Nuclease mapping and site-specific mutagenesis of the minimal RNA sequence defined a stem-loop structure that is located 40 nucleotides upstream from the cleavage site (nucleotide 320) and that is essential for accurate RNA cleavage. Abrogation of cleavage by disruption of base pairing within the stem-loop was reversed through the introduction of complementary nucleotide substitutions that reestablished the structure. We also provide evidence that divalent cations, essential components of the cleavage reaction, stabilized the stem-loop structure in solution. That capsid-specific antiserum eliminated specific RNA cleavage provides further evidence that the virus capsid gene encodes the essential endoribonuclease activity.

AB - The Leishmania RNA virus 1-4 capsid protein possesses an endoribonuclease activity responsible for single-site-specific cleavage within the 450-nucleotide 5' untranslated region of its own viral RNA transcript. To characterize the minimal essential RNA determinants required for site-specific cleavage, mutated RNA transcripts were examined for susceptibility to cleavage by the virus capsid protein in an in vitro assay. Deletion analyses revealed that all determinants necessary for accurate cleavage are encoded in viral nucleotides 249 to 342. Nuclease mapping and site-specific mutagenesis of the minimal RNA sequence defined a stem-loop structure that is located 40 nucleotides upstream from the cleavage site (nucleotide 320) and that is essential for accurate RNA cleavage. Abrogation of cleavage by disruption of base pairing within the stem-loop was reversed through the introduction of complementary nucleotide substitutions that reestablished the structure. We also provide evidence that divalent cations, essential components of the cleavage reaction, stabilized the stem-loop structure in solution. That capsid-specific antiserum eliminated specific RNA cleavage provides further evidence that the virus capsid gene encodes the essential endoribonuclease activity.

UR - http://www.scopus.com/inward/record.url?scp=0033988278&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033988278&partnerID=8YFLogxK

M3 - Article

C2 - 10590099

AN - SCOPUS:0033988278

VL - 74

SP - 130

EP - 138

JO - Journal of Virology

JF - Journal of Virology

SN - 0022-538X

IS - 1

ER -