Biology Reference
In-Depth Information
Chapter 8
Determination of the Transforming Activities
of Adenovirus Oncogenes
Thomas Speiseder , Michael Nevels , and Thomas Dobner
Abstract
The last 50 years of molecular biological investigations into human adenoviruses (Ads) have contributed
enormously to our understanding of the basic principles of normal and malignant cell growth. Much of
this knowledge stems from analyses of the Ad productive infection cycle in permissive host cells. Also,
initial observations concerning the transforming potential of human Ads subsequently revealed decisive
insights into the molecular mechanisms of the origins of cancer and established Ads as a model system for
explaining virus-mediated transformation processes. Today it is well established that cell transformation by
human Ads is a multistep process involving several gene products encoded in early transcription units 1A
(E1A) and 1B (E1B). Moreover, a large body of evidence now indicates that alternative or additional
mechanisms are engaged in Ad-mediated oncogenic transformation involving gene products encoded in
early region 4 (E4) as well as epigenetic changes resulting from viral DNA integration. In particular, stud-
ies on the transforming potential of several E4 gene products have now revealed new pathways that point
to novel general mechanisms of virus-mediated oncogenesis. In this chapter we describe in vitro and in
vivo assays to determine the transforming and oncogenic activities of the E1A, E1B, and E4 oncoproteins
in primary baby rat kidney cells, human amniotic fl uid cells and athymic nude mice.
Key words
Adenovirus, Transformation, Oncogenes, E1A, E1B, E4, Tumorigenicity, Baby rat kidney
cells, Primary human cells, Human amniotic fl uid cells, Nude mice
1
Introduction
Oncogenic transformation of primary cells by human adenoviruses
(Ads) is a multistep process that is initiated by the viral E1A gene.
Autonomous ectopic expression of E1A gene products is suffi cient
to induce unscheduled cellular DNA synthesis and proliferation by
virtue of their ability to interact with and modulate the function of
several growth-regulatory cellular proteins that control transcrip-
tion and cell cycle progression, including several Rb family mem-
bers and p300/CBP (reviewed in ref.
1
). Consequently, upon E1A
expression, primary cells with a limited life span can become
immortal. The Ad type 5 (Ad5) E1A transcription unit encodes
two major proteins of 289 and 243 amino acids derived from
