Hepatic Expression of SV40 Small-t Antigen Blocks the in Vivo CD95-Mediated Apoptosis

We have determined the effects of a number of mutations in the small-t antigen mRNA intron on the alternative splicing pattern of the simian virus 40 early transcript. Expansion of the distance separating the small-t pre-mRNA lariat branch point and the shared large T-small t 3' splice site from 18 to 29 nucleotides (nt) resulted in a relative enhancement of small-t splicing in vivo. This finding, coupled with the observation that large-T pre-RNA splicing in vitro was not affected by this expansion, suggests that small-t splicing is specifically constrained by a short branch point-3' splice site distance. Similarly, the distance separating the 5' splice site and branch point (48 nt) was found to be at or near a minimum for small-t splicing, because deletions in this region as small as 2 nt dramatically reduced the ratio of small-t to large-T mRNA that accumulated in transfected cells. Finally, a specific sequence within the small-t intron, encompassing the upstream branch sites used in large-T splicing, was found to be an important element in the cell-specific pattern of early alternative splicing. Substitutions within this region reduced the ratio of small-t to large-T mRNA produced in HeLa cells but had only minor effects in human 293 cells.

Download Full-text

Involvement of simian virus 40 (SV40) small t antigen in trans activation of SV40 early and late promoters.

Journal of Virology ◽

10.1128/jvi.66.3.1489-1494.1992 ◽

1992 ◽

Vol 66 (3) ◽

pp. 1489-1494 ◽

Cited By ~ 7

Author(s):

I Bikel ◽

M R Loeken

Keyword(s):

Simian Virus 40 ◽

Simian Virus ◽

T Antigen ◽

In Trans ◽

Small T Antigen ◽

Sv40 Small T

Download Full-text

Simian virus 40 small t antigen cooperates with mitogen-activated kinases to stimulate AP-1 activity

Molecular and Cellular Biology ◽

10.1128/mcb.14.9.6244-6252.1994 ◽

1994 ◽

Vol 14 (9) ◽

pp. 6244-6252

Author(s):

J A Frost ◽

A S Alberts ◽

E Sontag ◽

K Guan ◽

M C Mumby ◽

...

Keyword(s):

Simian Virus 40 ◽

Cell Types ◽

Simian Virus ◽

T Antigen ◽

Mitogen Activated Protein Kinase ◽

Early Gene ◽

Mitogen Activated Protein ◽

Small T Antigen

The simian virus 40 small tumor antigen (small t) specifically interacts with protein phosphatase type 2A (PP2A) in vivo and alters its catalytic activity in vitro. Among the substrates for PP2A in vitro are the activated forms of MEK and ERK kinases. Dephosphorylation of the activating phosphorylation sites on MEK and ERKs by PP2A in vitro results in a decrease in their respective kinase activities. Recently, it has been shown that overexpression of small t in CV-1 cells results in an inhibition of PP2A activity toward MEK and ERK2 and a constitutive upregulation of MEK and ERK2 activity. Previously, we have observed that overexpression of either ERK1, MEK1, or a constitutively active truncated form of c-Raf-1 (BXB) is insufficient to activate AP-1 in REF52 fibroblasts. We therefore examined whether overexpression of small t either alone or in conjunction with ERK1, MEK1, or BXB could activate AP-1. We found that coexpression of small t and either ERK1, MEK1, or BXB resulted in an increase in AP-1 activity, whereas expression of either small t or any of the kinases alone did not have any effect. Similarly, coexpression of small t and ERK1 activated serum response element-regulated promoters. Coexpression of kinase-deficient mutants of ERK1 and ERK2 inhibited the activation of AP-1 caused by expression of small t and either MEK1 or BXB. Coexpression of an interfering MEK, which inhibited AP-1 activation by small t and BXB, did not inhibit the activation of AP-1 caused by small t and ERK1. In contrast to REF52 cells, we observed that overexpression of either small or ERK1 alone in CV-1 cells was sufficient to stimulate AP-1 activity and that this stimulation was not enhanced by expression of small t and ERK1 together. These results show that the effects of small t on immediate-early gene expression depend on the cell type examined and suggest that the mitogen-activated protein kinase activation pathway is distinctly regulated in different cell types.

Download Full-text

Coordinated Activation of the Origin Licensing Factor CDC6 and CDK2 in Resting Human Fibroblasts Expressing SV40 Small T Antigen and Cyclin E

Journal of Biological Chemistry ◽

10.1074/jbc.m900687200 ◽

2009 ◽

Vol 284 (21) ◽

pp. 14126-14135 ◽

Cited By ~ 9

Author(s):

Elena Sotillo ◽

Judit Garriga ◽

Amol Padgaonkar ◽

Alison Kurimchak ◽

Jeanette Gowen Cook ◽

...

Keyword(s):

Cyclin E ◽

Human Fibroblasts ◽

T Antigen ◽

Origin Licensing ◽

Licensing Factor ◽

Small T Antigen ◽

Sv40 Small T

Download Full-text

Multiple cis-acting sequence elements are required for efficient splicing of simian virus 40 small-t antigen pre-mRNA

Molecular and Cellular Biology ◽

10.1128/mcb.8.9.3582-3590.1988 ◽

1988 ◽

Vol 8 (9) ◽

pp. 3582-3590

Author(s):

X Y Fu ◽

J D Colgan ◽

J L Manley

Keyword(s):

Alternative Splicing ◽

Branch Point ◽

Splice Site ◽

Simian Virus 40 ◽

Simian Virus ◽

T Antigen ◽

Specific Sequence ◽

Small T Antigen

We have determined the effects of a number of mutations in the small-t antigen mRNA intron on the alternative splicing pattern of the simian virus 40 early transcript. Expansion of the distance separating the small-t pre-mRNA lariat branch point and the shared large T-small t 3' splice site from 18 to 29 nucleotides (nt) resulted in a relative enhancement of small-t splicing in vivo. This finding, coupled with the observation that large-T pre-RNA splicing in vitro was not affected by this expansion, suggests that small-t splicing is specifically constrained by a short branch point-3' splice site distance. Similarly, the distance separating the 5' splice site and branch point (48 nt) was found to be at or near a minimum for small-t splicing, because deletions in this region as small as 2 nt dramatically reduced the ratio of small-t to large-T mRNA that accumulated in transfected cells. Finally, a specific sequence within the small-t intron, encompassing the upstream branch sites used in large-T splicing, was found to be an important element in the cell-specific pattern of early alternative splicing. Substitutions within this region reduced the ratio of small-t to large-T mRNA produced in HeLa cells but had only minor effects in human 293 cells.

Download Full-text

Regulation of Endosome Sorting by a Specific PP2A Isoform

The Journal of Cell Biology ◽

10.1083/jcb.142.6.1399 ◽

1998 ◽

Vol 142 (6) ◽

pp. 1399-1411 ◽

Cited By ~ 72

Author(s):

Sean S. Molloy ◽

Laurel Thomas ◽

Craig Kamibayashi ◽

Marc C. Mumby ◽

Gary Thomas

Keyword(s):

Protein Localization ◽

T Antigen ◽

Regulatory Subunits ◽

State Dependent ◽

Early Endosomes ◽

Wide Range ◽

Small T Antigen ◽

Endosomal System

The regulated sorting of proteins within the trans-Golgi network (TGN)/endosomal system is a key determinant of their biological activity in vivo. For example, the endoprotease furin activates of a wide range of proproteins in multiple compartments within the TGN/endosomal system. Phosphorylation of its cytosolic domain by casein kinase II (CKII) promotes the localization of furin to the TGN and early endosomes whereas dephosphorylation is required for efficient transport between these compartments (Jones, B.G., L. Thomas, S.S. Molloy, C.D. Thulin, M.D. Fry, K.A. Walsh, and G. Thomas. 1995. EMBO [Eur. Mol. Biol. Organ.] J. 14:5869–5883). Here we show that phosphorylated furin molecules internalized from the cell surface are retained in a local cycling loop between early endosomes and the plasma membrane. This cycling loop requires the phosphorylation state-dependent furin-sorting protein PACS-1, and mirrors the trafficking pathway described recently for the TGN localization of furin (Wan, L., S.S. Molloy, L. Thomas, G. Liu, Y. Xiang, S.L. Ryback, and G. Thomas. 1998. Cell. 94:205–216). We also demonstrate a novel role for protein phosphatase 2A (PP2A) in regulating protein localization in the TGN/endosomal system. Using baculovirus recombinants expressing individual PP2A subunits, we show that the dephosphorylation of furin in vitro requires heterotrimeric phosphatase containing B family regulatory subunits. The importance of this PP2A isoform in directing the routing of furin from early endosomes to the TGN was established using SV-40 small t antigen as a diagnostic tool in vivo. The role of both CKII and PP2A in controlling multiple sorting steps in the TGN/endosomal system indicates that the distribution of itinerant membrane proteins may be acutely regulated via signal transduction pathways.

Download Full-text