Structural Determinants of Human a-Glohin 
Gene Expression 
Stephen A. Liebhaber, M.D. — Investigator 
Dr. Liebhaber is also Professor of Genetics and Medicine (Hematology) at the University of Pennsylvania 
School of Medicine. He received his B.A. degree in chemistry from Brandeis University and his M.D. degree 
from Yale University. He took clinical training in internal medicine and hematology at Case Western 
Reserve, the University of Colorado, Washington University, and the University of California, San 
Francisco. As a postdoctoral fellow with David Schlessinger at Washington University, Dr. Liebhaber 
examined ribosomal RNA processing, and with Yuet Wai Kan at the University of California, San 
Francisco, he studied human globin gene expression and genetic defects in a-thalassemia. Before moving 
to Philadelphia, he was a faculty member of the Department of Medicine at UCSF. 
OUR laboratory has largely concentrated on 
studying the expression of the human glo- 
bin genes. These genes encode hemoglobin, the 
major red cell protein responsible for transport of 
oxygen from the lungs to peripheral tissues. 
Since the hemoglobin molecule, a2i^2> is com- 
posed of an equal number of a- and ;8-globin 
chains, normal synthesis demands balanced ex- 
pression of both sets of genes, which are located 
on different chromosomes. Defects in either set 
result in an imbalance of expression and conse- 
quent hereditary anemia: a- or (S-thalassemia. 
Thalassemias result from more than 150 different 
mutations in the globin genes, affecting the 
health of millions worldwide. 
Certain characteristics of globin gene expres- 
sion make it particularly interesting for study. 
The extremely high level of globin mRNA in the 
differentiating red cell (over 95 percent of total 
cellular mRNA) has no equal in any other cell 
type. This abundance reflects both high levels of 
globin gene transcription and an unusual stability 
of the mature globin mRNA. 
An additional interesting aspect of globin ex- 
pression is that the genes in the a- and |S-globin 
gene clusters follow an orderly sequence of ex- 
pression during embryologic development. This 
results in a well-defined switch from embryonic 
to adult globin gene expression during develop- 
ment of the fetus. The switching results in the 
synthesis of successive hemoglobins with oxygen 
affinities that match changes in the uterine envi- 
ronment. The active transcription, the clearly de- 
fined pattern of developmental switching, and 
the unusual mRNA stability are areas of special 
focus in our laboratory. 
The loss of a-globin expression observed in 
a-thalassemia usually results from deletion or ab- 
normal structure of one or more of the a-globin 
genes. We have recently studied three unrelated 
patients from Germany, Portugal, and Italy who 
have an unusual form of a-thalassemia. In each 
case the loss of a-globin synthesis reflects loss of 
expression of a structurally normal a-globin clus- 
ter. In other words, one of the a-globin gene clus- 
ters in each of these patients is not functioning, 
even though the genes in these clusters are struc- 
turally normal and synthesize normal levels of 
a-globin when isolated and expressed in tissue 
culture cells. Although initially puzzling, the 
mechanism of this remarkable expression defect 
was eventually defined by extensive DNA map- 
ping. In each of the three independent cases we 
demonstrated a large deletion 5' to the inacti- 
vated cluster itself. In one case this deletion be- 
gan as much as 50,000 bases 5' to the silenced 
a-globin genes. By comparing the maps of each of 
these deletions, a region of common overlap was 
noted. These studies define a new category of 
a-thalassemia and demonstrate a critical determi- 
nant of a-globin gene expression located entirely 
external to the a-globin gene cluster. A similar set 
of transcriptional signals has been localized adja- 
cent to the j8-globin gene cluster. One can there- 
fore speculate that such signals serve coordi- 
nately to activate and balance the expression of 
the a- and ;8-globin clusters in the red cell. 
The human a-globin gene cluster contains a 
globin gene expressed specifically in the embryo 
and two a-globin genes, al and a2, expressed in 
the fetus and the adult. The switch from embry- 
onic f-globin to adult a-globin expression occurs 
at the end of embryonic development (7-8 
weeks of gestation) . This critical developmental 
switch, which occurs widely in mammals, pres- 
ents a well-defined model system for studying de- 
velopmental control of gene expression. 
To establish a system in which to study switch- 
ing within the human a-globin gene cluster, we 
have injected the human embryonic and adult 
a-globin genes into fertilized mouse eggs to gen- 
erate transgenic mice. The red cells of these 
transgenic mice appropriately express the human 
transgenes during development. In the embry- 
onic period, there is a parallel expression of the 
human and mouse f-globin genes and, by day 1 2 
of development, parallel expression of the 
a-globin genes. These data suggest that 1 ) the hu- 
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