download report - Istituto Pasteur

P a r t i c i p a n t s :
Alessandra Vacca, Maria Pia Felli, professors; Diana
Bellavia, Antonio F. Campese, researchers; Claudio Talora,
Saula Checquolo, post-doc fellows; Samantha Cialfi, Rocco
Palermo, Giuseppina Di Mario, PhD students; Massimo
Zani, technician.
C o l l a b o r a t i o n s :
Department of Cell and Molecular Biology, Medical Nobel
Institute, Karolinska Institute, Stockholm, Sweden (Prof. Urban
Lendahl); Harvard Medical School, Dana-Farber Cancer Institute,
Boston, MA, USA (Prof. Harald von Boehmer).
Report of activity
We previously observed that thymocytes from
Notch3-IC (N3-IC) transgenic (tg) mice display
overexpression of pTα/pre-TCR chain and
increased NF-κB activity (Bellavia et al., EMBO J.
2000, 19:3337 ). Furthermore, we demonstrated that
combined overexpression of pTα and Notch3 in tg
mice, triggers important molecular events, some of
which are NF-κB-mediated and cooperate in sustaining
Notch3-induced lymphomagenesis (Felli et al.,
Oncogene 2005, 24:992; Talora et al., EMBO Rep.
2003, 4:1067).
The ability that we demonstrated of activated Notch3
to lead to constitutive activation of NF-kB both, in
vivo and in vitro, together with previous reports that
demonstrated on one hand that Notch1 is able to bind
to the p50 NF-kB subunit and on the other hand that
NF-kB2, the precursor of p52, is a putative target
gene of activated Notch1 suggest strict relationships
between activated Notch signaling and different NFkB
activation pathways. In accord with this hypothesis,
we recently demonstrated that Notch3 is indeed
able to activate canonical and alternative NF-kB pathways
by regulating the assembling and activation of
different IKK complexes, depending on the presence
of pre-TCR (Vacca et al., EMBO J. 2006, 25:1000).
Moreover, IKKalpha forms a complex with Notch3
Principal investigator: Isabella Screpanti
Professor of General Pathology
Dipartimento di Medicina Sperimentale
Tel: (+39) 06 44700816; Fax: (+39) 06 4464129
isabella.screpanti@uniroma1.it
103
Cellular and molecular immunology - AREA 5
Analysis of the role of preTCR-triggered NF-kB in T cell
leukemogenesis: relationship with activated Notch signaling
even independently on the presence of pre-TCR , but
only the presence of pre-TCR allows NIK to participate
at the Notch3-IKKalpha complex. This last
observation suggests that Notch3 may activate the
alternative pathway through an additional direct
mechanism.
In order to genetically approach this issue, we
planned to generate two lines of double transgenic
mice, one overexpressing Notch3-IC and mutant for
p50, a component of the canonical heterodimer, and
the other one overexpressing Notch3 and mutant
forp52, a component of the alternative heterodimer,
to the purpose to obtain mice in which only one NFkB
activation pathway is impaired.
The following results have been obtained in the last
two years:
Generation of Notch3-IC/ p50-/- and Notch3-
IC/p52-/- double transgenic mice
We obtained p50-/- and p52-/- mice from Dr. G.
Franzoso (University of Chicago, ILL, USA). These
mice were generated by homologous recombination
and are totally lacking of functional p50 or p52 protein.
We have now established Notch3-IC/p50-/and,
more recently, Notch3-IC/p52-/- double mutant
mice and started their phenotypic characterization.
Phenotypic and functional characterization
of cell population in different lymphoid
organs with respect to the kinetic of
development of multicentric T cell lymphoma
The T cell oncogenic potential of Rel/NFkB is well
established. If the oncogenic role of Notch3 is mediated
by the activation of NF-kB, we would predict
that in double transgenic mice the lymphoma development
would be affected. For this reason, the mice were
clinically observed and mortality curves were drawn.
a) Notch3-IC/p52-/- double mutant mice. We previously
showed that the canonical pathway of NF-kB
activation, mediated by the nuclear translocation of
p50/p65 heterodimer, is mostly responsible of the