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Blood, 1 September 2003, Vol. 102, No. 5, pp. 1566-1567
More is not always better
The t(15;17) is present in more than 95% of patients with acute
promyelocytic leukemia (APL). This translocation generates 2 fusion proteins:
promyelocytic leukemia–retinoic acid receptor alpha (PML-RAR ) and
RAR-PML. To understand the development of APL and to search for more
effective treatments of this disease, several PML-RAR APL transgenic
mouse models have been generated using myeloid specific regulatory elements,
such as cathepsin G and myeloid-related protein 8 (MRP8), to direct
PML-RAR expression into early myeloid cells. All of these
PML-RAR mice develop a myeloproliferative syndrome early in life, and
15% to 20% of these mice develop an APL-like disease after 6 to 14 months (see
Grisolano et al, Blood. 1997; 89:376-387; Brown et al, PNAS. 1997;94:
2551-2556; and He et al, PNAS. 1997;94: 5302-5307). Additional expression of
the t(15;17) reciprocal fusion protein RAR-PML can increase the
percentage of APL in transgenic mice by 4-fold. However, the long latency
remains. These results indicate that PML-RAR is necessary but not
sufficient for APL development. Additional mutations are probably required for
leukemia to occur.
The current explanation of how PML-RAR is involved in leukemogenesis
centers on the dominant-negative effect of PML-RAR. PML-RAR
contains the DNA and retinoic acid (RA) ligand-binding domains of wild-type
RAR. In the absence of a ligand, both RAR and PML-RAR
repress transcription due to the interaction with nuclear receptor
corepressor/silencing mediator for retinoid and thyroid receptor (NCoR/SMRT)
and form complexes with histone deacetylase (HDAC). Removing acetyl groups by
HDACs increases the positive charge on proteins and enhances protein
interactions with negatively charged DNA to keep chromatin in a more compacted
confirmation, which does not favor the initiation of transcription.
Furthermore, removing acetyl groups may also enhance the binding of repressor
proteins. In the presence of physiologic concentrations of RA, RA binds to RAR
and changes the conformation of RAR, replacing the HDAC complex with
transcription coactivators, including histone acetylase (HAT) complex.
Acetylation of histones changes chromatin structure to favor gene expression.
The PML portion of PML-RAR contains the oligomerization domain of PML.
The oligomerized PML-RAR forms a more stable complex with NCoR/SMRT and
HDAC. Dissociation of this complex requires a much higher concentration of RA.
This theory explains why a high dosage of RA can be effectively used to treat
APL. It also suggests that relatively higher levels of PML-RAR
expression may be more effective at initiating APL development.
It has been difficult to detect PML-RAR expression in the above
mentioned PML-RAR transgenic mice. Therefore, Westervelt and colleagues
(page 1857)
hypothesized that increasing the expression of PML-RAR may enhance the
penetration of APL development in transgenic mice because the upstream
fragment of the human cathepsin G used in their previously reported
PML-RAR transgenic mice may lack critical regulatory elements required
for high-level transgene expression. In order to fully capture cathepsin
regulatory elements to express PML-RAR in early myeloid cells,
Westervelt et al generated another PML-RAR mouse model by knocking the
PML-RAR cDNA into the cathepsin G 5' untranslated region. In
contrast to the 15% to 20% penetration of APL in previously reported
PML-RAR transgenic mouse models, more than 90% of PML-RAR
knock-in mice developed APL, although the latency was similar to other
transgenic models. These results suggested that the new knock-in model
provides a level of PML-RAR expression that is more optimal for APL
development, and most of us would have predicted that this level would be
higher. However, when real-time reverse transcriptase–polymerase chain
reaction (RT-PCR) analysis was used to compare the expression of
PML-RAR in bone marrow cells and APL cells from this knock-in model (vs
their human cathepsin G–PML-RAR transgenic mouse model) it was
surprising to discover that PML-RAR was expressed at an extremely low
level in the knock-in mice—less than 3% of the expression in the
transgenic mice. This result goes directly against the original hypothesis
(based on the dominant-negative effect of the interaction of PML-RAR
with the HDAC complex) that more PML-RAR expression will enhance the
development of APL. Instead, an optimal low level of PML-RAR expression
appears to be favorable for APL development. Furthermore, the knock-in model
may target PML-RAR expression to a more proper population of myeloid
cells for APL development. It has been reported previously that PML-RAR
triggers cell death in most cell lines. On one hand, PML-RAR expression
may favor leukemia development; on the other hand, it may mitigate against
leukemia. The disruption of such a balance with additional mutations may lead
to leukemogenesis. Most importantly, this report urges the search for
additional mechanisms besides the dominant-negative effect of PML-RAR
via HDAC to fully explain how t(15;17) is involved in the development of
APL.
Dong-Er Zhang
The Scripps Research Institute
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