Nucleic Acids Research, Vol 24, Issue 9 1653-1661, Copyright © 1996 by Oxford University Press
J Eul, M Graessmann and A Graessmann
The early SV40 BstXI-BamHI (Bst/Bam) DNA fragment encodes exclusively for
the second exon of the large T-antigen and contains the intact small
t-antigen intron. Rat cells transformed by the p14T, a construct that
carries the Bst/Bam DNA fragment as a tail-to-head tandem duplication,
synthesize a truncated T-antigen (T1-antigen) without having a direct
equivalent at the DNA level. Formation of the T1-mRNA occurs by means of
two distinct mechanisms: alternative-tandem-cis- splicing and
trans-splicing. To generate the T1-mRNA the cells utilize a cryptic 5'
splice site, located within the second exon of the large T- antigen and the
regular small t-antigen 3' splice site. Since these splice sites are in an
inverted order two Bst/Bam transcripts are required to generate one T1-mRNA
molecule. For alternative-tandem-cis- splicing the cells utilize a 4.4 kb
pre-mRNA that contains the sequence of the entire Bst/Bam tandem repeat.
The proximal Bst/Bam segment provides the 5' donor splice site and the
distal segment the 3' acceptor site. This requires that the pre-mRNA not be
cleaved after the RNA polymerase II has passed the polyadenylation signal
of the proximal Bst/Bam DNA segment. Synthesis of the 4.4 kb pre-mRNA was
demonstrable by RT-PCR but not by Northern blot analysis. For
trans-splicing, the cells utilize two separate pre-mRNA molecules. One
transcript provides the cryptic 5' splice donor site and the other the 3'
splice acceptor site. To demonstrate this a three base pair deletion was
introduced into the proximal Bst/Bam segment of the p14T DNA (p14Tdelta-3)
as a marker, destroying the recognition site for Pf/MI restriction enzyme.
This deletion allowed the differentiation between the proximal and distal
Bst/Bam segment. RT-PCR analysis and DNA sequencing confirmed that the
p14Tdelta-3 transformed cells generate the T1-mRNA by intra- and
inter-molecular RNA splicing.
ARTICLES
Trans-splicing and alternative-tandem-cis-splicing: two ways by which mammalian cells generate a truncated SV40 T-antigen
Institut fur Molekularbiologie und Biochemie, Freie Universitat, Berlin, Germany.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  R. Rigatti, J.-H. Jia, N. J. Samani, and I. C. Eperon Exon repetition: a major pathway for processing mRNA of some genes is allele-specific Nucleic Acids Res., January 22, 2004; 32(2): 441 - 446. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Labrador and V. G. Corces Extensive Exon Reshuffling Over Evolutionary Time Coupled to Trans-Splicing in Drosophila Genome Res., October 1, 2003; 13(10): 2220 - 2228. [Abstract] [Full Text] [PDF]
|
|