Gene Regulation in Animal Cells 
Joseph R. Nevins, Ph.D. — Investigator 
Dr. Nevins is also Professor of Genetics and of Microbiology and Immunology at Duke University Medical 
Center. He received his Ph.D. degree in virology at Duke University, where he studied with Wolfgang Joklik. 
His postdoctoral studies as a Jane Cofftn Childs fellow focused on the mechanisms of mRNA biogenesis 
and were conducted with James Darnell at the Rockefeller University, where he later became a faculty 
member. 
REGULATION of gene expression is central in 
the determination of cellular phenotype and 
in the complex transformations that take place 
during such events as oncogenesis. The goal of 
our laboratory is to elucidate the molecular mech- 
anisms of gene control pathways. 
Adenovirus Transcriptional Regulation — 
A Role in Oncogenesis 
Complex cellular events are often best studied 
through the use of simple model systems. The 
control of transcription (transfer of DNA-coded 
information to RNA to guide protein synthesis) 
mediated by viral regulatory proteins is instruc- 
tive as regards the v^^orkings and control of tran- 
scription factor activity in eukaryotic cells. 
Work in our laboratory has focused on the elu- 
cidation of transcription control by viral pro- 
teins, using in vitro assays. We have found that a 
cellular transcription factor termed E2F is regu- 
lated by other cellular proteins and that these in- 
teractions prevent E2F from being co-opted by 
the viral genome. The adenovirus regulatory pro- 
tein El A, however, can dissociate these com- 
plexes, releasing E2F, which can then be utilized 
by another viral product, the E4 protein. This re- 
directs the cellular transcription factor for viral 
purposes. 
These findings are significant for several rea- 
sons. First, they provide insight into the evolu- 
tion of viral regulatory events that manipulate the 
host cell for the virus's benefit. In particular, they 
highlight the interplay between two viral pro- 
teins (El A and E4) that together redirect the E2F 
factor to a viral-specific role. 
Second, this activity of ElA correlates with the 
oncogenic activity of the protein. The sequences 
in ElA that are responsible for this activity are 
shared with other viral oncogene products, such 
as the SV40 T antigen and the human papilloma- 
virus E7 protein. Each of these viral proteins has 
been shown to activate transcription, dependent 
on E2F. 
Third, our recent experiments have shown that 
the viral T antigen and E7 protein can also dissoci- 
ate complexes containing the E2F transcription 
factor. In short, this latest discovery has defined a 
common biochemical activity of these viral regu- 
latory proteins that is likely part of their onco- 
genic activity. 
The identification of the cellular proteins that 
are complexed to the E2F factor has provided im- 
portant insight into the role of ElA as an onco- 
gene. We have found that there are multiple such 
E2F complexes and that their formation is regu- 
lated during the cell cycle. One complex con- 
tains the cell cycle-regulated protein cyclin A, 
and another the retinoblastoma gene product. 
The product of the retinoblastoma susceptibil- 
ity gene {RBI) is a 1 10-kDa nuclear phosphopro- 
tein that is expressed at equivalent levels in all 
cell types examined except certain tumor cells in 
which the RBI gene has been inactivated by mu- 
tation or deletion. Given that it is the loss of RBI 
function that is correlated with the development 
of certain human tumors, it is widely believed 
that the Rb protein functions to limit or constrain 
cell proliferation. 
It now appears that the ability of E 1 A, as well as 
T antigen and E7, to dissociate the E2F-Rb com- 
plex, releasing free E2F, is at least one of the 
events of Rb inactivation. The E2F-Rb complex, 
like the E2F-cycIin complex, is also regulated by 
cell proliferation. Moreover, the E2F-Rb complex 
is absent in cells that express a nonfunctional 
form of the Rb protein. 
This latter result strongly argues that E2F is a 
functional target for Rb action. Thus the conse- 
quence of either ElA action, the deletion or mu- 
tation of Rb, or the phosphorylation of Rb is the 
loss of the E2F-Rb interaction and the generation 
of free E2F molecules. In short, "inactivation" of 
Rb can be viewed as the loss of the E2F-Rb inter- 
action. Moreover, it would appear that the conse- 
quence of disruption of either the E2F-cyclin A 
complex or the E2F-Rb complex is the release of 
transcriptionally active E2F and that this event 
may be important for stimulating the transcrip- 
tion of genes whose products are critical for en- 
hancing cell proliferation. 
Molecular Mechanisms for Polyadenylation 
The generation of the 3' terminus of the mature 
mRNA, commonly termed poly(A) site formation. 
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