Molecular Genetics of the Major 
Histocompatibility Complex 
Jan Geliebter, Ph.D. — Assistant Investigator 
Dr. Geliebter is also Assistant Professor and Head of the Mammalian Molecular Genetics Laboratory at the 
Rockefeller University. He received his Ph.D. degree in microbiology and immunology from the State 
University of New York, Downstate Medical Center. He was a postdoctoral fellow and research associate 
in the laboratory of Stanley Nathenson at the Albert Einstein College of Medicine, Bronx, New York. 
THE immune system functions to rid the body 
of foreign objects such as bacteria, viruses, 
tumors, and transplants. The portion of such mat- 
ter that is recognized as foreign by the immune 
system is called an antigen. Antigens that are 
found on cells are "presented" to the immune 
system by cell surface molecules called histocom- 
patibility molecules (also called HLA molecules 
in humans and H-2 molecules in the mouse). His- 
tocompatibility molecules are able to bind anti- 
genic fragments of, for example, viruses, and 
stimulate white blood cells (lymphocytes) to 
attack the virus-infected cell, thereby limiting 
the spread of infection. Without these antigen- 
presenting molecules the host would be unable 
to mount an immune response against pathogens 
and would not survive. 
Different H-2 molecules can bind and present 
different types of antigens. Because inbred mice 
have about three different types of H-2 molecules 
on their cells, they can bind and present a large, 
but limited, number of antigens to the immune 
system. 
To ensure the survival of the species, it is bene- 
ficial that many varieties of H-2 molecules be 
present in the population. In this way there will 
always be some portion of the population that 
will mount an immune response to a given anti- 
gen. Indeed, an extraordinary number of differ- 
ent histocompatibility molecules have been 
found in almost all species investigated. In hu- 
mans, the large variety of HLA molecules ensures 
our survival but is the major obstacle confound- 
ing tissue transplantation. Our interest lies in the 
genetic mechanism that generates the different 
histocompatibility genes in mice and other 
species. 
The H-2 genes of the mouse are part of the 
larger major histocompatibility complex class I 
multigene family. This gene family also contains 
genes that are structurally similar to H-2 genes 
and have unknown functions. The genetic mecha- 
nism that generates variety in H 2 genes is the 
microrecombination process, which reassorts 
DNA among H-2 genes and other related class I 
genes. By substituting small segments of class I 
gene sequences into //-2 genes, the microrecom- 
bination process can create new H-2 molecules 
that have different antigen-presenting capabili- 
ties, thereby expanding the immune responsive- 
ness of the population. 
Our interest is to understand better the mecha- 
nism underlying the microrecombination pro- 
cess. This process has previously been studied by 
identifying microrecombinant mice that differed 
from their otherwise identical siblings by altered 
H-2 genes. Since microrecombinant H-2 mole- 
cules elicit skin graft rejection, these studies 
were accomplished by skin graft testing thou- 
sands of mice. The rejection of a skin graft by a 
sibling mouse signaled an alteration in H-2 mole- 
cules. These labor-intensive studies found that, 
on the average, one microrecombinant mouse 
was detected for every 5,000 skin grafts 
performed. 
To gain further insight into the microrecom- 
bination process, we are using in vitro-engi- 
neered constructs to detect microrecombinant 
H-2 genes. We have constructed a fusion gene in 
which /3-galactosidase sequences replace two cy- 
toplasmic exons for the gene. The fusion pro- 
tein can be detected by staining for |8-galactosi- 
dase activity, which is manifested as blue-colored 
cells. We have also site-directed two in-frame ter- 
mination codons in the gene at positions that 
undergo frequent microrecombinations. This 
prevents the expression of |S-galactosidase. 
/3-Galactosidase expression can be rescued by a 
microrecombination with a linked class I gene, 
Q4, that recombines away the termination co- 
dons. Thus microrecombinations can be scored as 
blue cells. 
This microrecombination construct, once in- 
troduced into a variety of cell types, will help 
determine the microrecombination frequencies 
of different cells. Its introduction into transgenic 
mice will help determine frequencies in germ 
cells. Data from previous studies indicate that mi- 
crorecombinations occur in female germ cells. 
We also hope to determine if they occur in sperm 
cells as well and at what frequency. Some strains 
of mice may undergo microrecombinations more 
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