TY - JOUR
T1 - The herpes simplex virus type 1 2.0-kilobase latency-associated transcript is a stable intron which branches at a guanosine
AU - Zabolotny, Janice M.
AU - Krummenacher, Claude
AU - Fraser, Nigel W.
PY - 1997/6
Y1 - 1997/6
N2 - We have used a minigene construct of the herpes simplex virus type I (HSV-1) latency-associated transcript (LAT) gene to analyze its transcripts in transient transfection assays. A 2.8-kb fragment of the approximately 8.5- kb LAT gene encompassing the 2.0-kb LAT was cloned into a eukaryotic expression vector downstream of the cytomegalovirus immediate-early gene promoter. Northern hybridization of RNA isolated from transfected COS-1 cells identified three LAT-specific transcripts, 3.4, 2.0, and 1.4 kb in size. Mapping of these transcripts by Northern hybridization indicated that the 1.4- and 2.0-kb RNAs are nonoverlapping, while the 3.4-kb RNA overlaps both smaller RNAs. Reverse transcription-PCR (RT-PCR) and partial sequencing of the 1.4-kb RNA revealed that this RNA is the spliced exons of the 3.4-kb primary transcript. The 2.0-kb LAT appears to be an intron accumulating after splicing of the minor LAT (mLAT) pre-mRNA. The splice donor and acceptor sites for the 2.0-kb LAT identified in transfected and HSV-1-infected cells are identical. Mapping of the branch point of this intron by RT-PCR in transfected and HSV-1-infected cells, as well as in latently infected murine trigemial ganglia, shows that it is a guanosine. This branch site does not bear homology to consensus mammalian branch site sequences. These data provide evidence that the 2.0-kb LAT is an intron of the mLAT pre-mRNA with a unique branch point.
AB - We have used a minigene construct of the herpes simplex virus type I (HSV-1) latency-associated transcript (LAT) gene to analyze its transcripts in transient transfection assays. A 2.8-kb fragment of the approximately 8.5- kb LAT gene encompassing the 2.0-kb LAT was cloned into a eukaryotic expression vector downstream of the cytomegalovirus immediate-early gene promoter. Northern hybridization of RNA isolated from transfected COS-1 cells identified three LAT-specific transcripts, 3.4, 2.0, and 1.4 kb in size. Mapping of these transcripts by Northern hybridization indicated that the 1.4- and 2.0-kb RNAs are nonoverlapping, while the 3.4-kb RNA overlaps both smaller RNAs. Reverse transcription-PCR (RT-PCR) and partial sequencing of the 1.4-kb RNA revealed that this RNA is the spliced exons of the 3.4-kb primary transcript. The 2.0-kb LAT appears to be an intron accumulating after splicing of the minor LAT (mLAT) pre-mRNA. The splice donor and acceptor sites for the 2.0-kb LAT identified in transfected and HSV-1-infected cells are identical. Mapping of the branch point of this intron by RT-PCR in transfected and HSV-1-infected cells, as well as in latently infected murine trigemial ganglia, shows that it is a guanosine. This branch site does not bear homology to consensus mammalian branch site sequences. These data provide evidence that the 2.0-kb LAT is an intron of the mLAT pre-mRNA with a unique branch point.
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U2 - 10.1128/jvi.71.6.4199-4208.1997
DO - 10.1128/jvi.71.6.4199-4208.1997
M3 - Article
C2 - 9151806
AN - SCOPUS:0030919832
SN - 0022-538X
VL - 71
SP - 4199
EP - 4208
JO - Journal of virology
JF - Journal of virology
IS - 6
ER -