MOLECULAR GENETICS OF LYMPHOCYTE DEVELOPMENT 
StanieyJ. Korsmeyer, M.D., Associate Investigator 
The Bcl-2 Proto-oncogene 
The t(l4;18) chromosomal breakpoim found in 
follicular B cell lymphoma juxtaposes the new 
proto-oncogene Bcl-2 with the immunoglobulin 
(Ig) heavy-chain gene, resulting in marked over- 
production of Bcl-2. Bcl-2 is novel among proto- 
oncogenes in that it localizes to mitochondria and 
demonstrates the unique role of blocking pro- 
grammed cell death without promoting cell divi- 
sion. 
Transgenic mice bearing a Bcl-2-lg minigene, 
which recapitulates the t(l4;18), initially display a 
polyclonal expansion of resting B cells with a pro- 
longed life span. Over time these mice develop 
monoclonal, high-grade lymphomas. Extended cell 
survival can be a primary oncogenic event that ap- 
pears to favor the acquisition of secondary genetic 
abnormalities. 
Bcl-2 protein is widespread during embryonic 
development but topographically restricted in 
adult tissues. Within adult tissues that demonstrate 
apoptotic cell turnover, Bcl-2 is often geographi- 
cally restricted to long-lived or proliferating cell 
zones. Many developing organs express Bcl-2 at high 
levels and in a more widespread pattern. For exam- 
ple, the relatively undifferentiated neuroepithe- 
lium in the mouse retina at embryonic day 14.5 uni- 
formly expresses Bcl-2. Following differentiation 
and migration, the inner nuclear layer of inter- 
neuron is much more intense than the outer layer of 
photoreceptors. At day 11.5, Bcl-2 is present 
throughout the embryonic limb bud, but by day 
14.5, when digits have separated, expression is lost 
in the interdigital regions of death. The distribution 
pattern of Bcl-2 suggests that it is selectively down- 
regulated to enable cell death to occur. 
Maintenance ofB cell memory represents a nor- 
mal physiologic role for Bcl-2. Bcl-2-lg mice that 
overproduce Bcl-2 provide an in vivo model to as- 
sess the role of Bcl-2 upon immune responsiveness. 
Secondary immune responses by these transgenics 
were markedly protracted. This resulted from long- 
term persistence of antibody-secreting plasma cells 
as well as an extended lifetime for resting memory B 
cells. Bcl-2 spared the need for antigen in maintain- 
ing immune responsiveness. 
Bcl-2 inhibits multiple forms of apoptosis but 
not negative selection within thymocytes. In the 
thymus, Bcl-2 is present in the mature T cells of the 
medulla but only in rare cells of the cortex. To as- 
sess the role of Bcl-2 in the programmed death of 
thymocytes, Ictf ''-Bcl-2 transgenic mice were gen- 
erated that redirected Bcl-2 expression to cortical 
thymocytes. Bcl-2 protected immature CD4^CD8"^ 
thymocytes from glucocorticoid, radiation, and 
anti-CD3-induced apoptosis. Moreover, Bcl-2 alters 
T cell maturation, increasing €03"""* thymocytes 
felt to represent an intermediate stage after posi- 
tive selection. Despite this, clonal deletion of T 
cells that recognize endogenous superantigen still 
occurred. 
Moreover, matings with T cell receptor transgenic 
mice indicate that the negative selection of thymo- 
cytes reactive with class 1- or class Il-restricted an- 
tigen also occurred. Such double-transgenic mice, 
when placed on a neutral MHC background, suggest 
that Bcl-2 may participate in positive selection. It 
appears that multiple death pathways operate 
within the thymus and can be distinguished by their 
dependence on Bcl-2. 
A novel negative regulatory element inhibits ex- 
pression from an upstream promoter. Nuclear 
run-on assays demonstrate that Bcl-2 transcription 
decreases in parallel with RNA levels during B cell 
differentiation. Promoter-reporter constructs iden- 
tified a negative regulatory element (NRE) in the 
Bcl-2 5'-untranslated region. The NRE dramatically 
decreases expression from the Bcl-2 PI promoter or 
heterologous promoters in a position-dependent 
fashion. 
Multiple functionally redundant sequence ele- 
ments mediate NRE activity. Chromatin structure of 
the endogenous NRE differs from pre-B to mature B 
lines. These results indicate that negative control of 
transcription is an important determinant of the dif- 
ferential expression of Bcl-2. 
Transgenic Mice That Redirect 
Transcription Factors to the Thymus 
Develop T Cell Acute Lymphoblastic 
Leukemia / Lymphoma 
Interchromosomal translocations within T cell 
neoplasms often involve their antigen receptor 
genes and another chromosome that provides a new 
transcription factor gene. Ttg-1, from chromosome 
segment 11 pi 5, encodes a nuclear protein — pre- 
dominantly expressed in neuronal cells — and be- 
longs to a novel family of transcription factors with 
LIM (Jin- 11, Isl-1, mec-3) domains. 
Transgenic mice that overexpress this candidate 
oncogene in early thymocytes develop immature, 
aggressive T cell leukemia/lymphoma. A minority 
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