II. Characterization of the Moloney Murine Leuke- 
mia Virus i|i Region. 
Dr. Belmont's laboratory has investigated some 
of the molecular mechanisms involved in vector as- 
sembly. Initial studies have used chemical protec- 
tion mapping of the RNA in virus particles and phy- 
logenetic analysis to develop a model for the 
secondary structure of the RNA in the v|; packaging 
region. Computer analysis of the chemical modifica- 
tion data and primary sequence was conducted 
using PCFOLD 4.0, which contains algorithms for 
minimization of free energy. A model of the RNA 
secondary structure based on all these data pro- 
poses that the two strands of RNA are hybridized in 
a unique homodimer within the 5' region of On- 
going studies are now aimed at using site-directed 
mutagenesis to allow genetic tests of the secondary 
structure model. It is suspected that the NC pep- 
tide encoded in the gag gene is involved in the spe- 
cific recognition of the vector RNA. NC is a proto- 
typic finger protein and is known to have nucleic 
acid-binding properties. Several vectors for over- 
expression of NC have been constructed and are 
being tested. The ability of the free NC peptide to 
bind viral RNA and thus inhibit the normal packag- 
ing process is also being investigated. 
III. Use of TCR7 to Analyze T Cell Development. 
A major concern for gene therapy research cen- 
ters on the normal biology of the target cells for 
gene transfer. Retrovirus vector infection has been 
used to mark the progeny of hematopoietic stem 
cells genetically and thus investigate their develop- 
PUBLICATIONS 
mental potential and dynamics. A different ap- 
proach is being taken to the problem of T cell pop- 
ulation kinetics and lineage relationships. In these 
experiments the rearrangement of the TCR7 loci 
provides natural markers for families of developing 
T cells. It is known that Cyl begins rearranging 
early in fetal thymocyte development. Current 
models suggest that TCR7 loci rearrange in lym- 
phoid progenitors before TCRP or TCRa. Thus the 
unique rearrangement, and in particular the 
specific sequence at the V-J junction in single 
TCR7S, should mark families of T cells before they 
clonally diverge with rearrangement of TCRp. With 
the help of Dr. Richard Gibbs (Baylor College of 
Medicine), specific PCR and direct sequencing tech- 
niques for V72, V73, and V74 have been developed. 
Initial experiments have focused on the production 
of cells bearing rearrangements of V73. Previous 
data had suggested that cells expressing this gene 
were produced only during early prenatal thymic 
ontogeny, that the diversity was very restricted, and 
that they were anatomically sequestered in the den- 
dritic epidermal cell population. Analysis of adult 
and developing thymic cells indicates that this re- 
ceptor can be highly diverse and that the cells 
expressing this gene are present throughout life. 
The laboratory is currently using this method to in- 
vestigate the possible clonal relationships within 
thymic microanatomic compartments, i.e., thymic 
nurse cell-enclosed lymphocytes and macrophage 
rosettes. 
Dr. Belmont is also Assistant Professor of Molecu- 
lar Genetics, Pediatrics, Microbiology, and Immu- 
nology at Baylor College of Medicine. 
Article 
Belmont, J.W , MacGregor, G.R., Wager-Smith, K. , Fletcher, F.A., Moore, K.A., Hawkins, D. , Villalon, D. , Chang, 
S.M.-W, and Caskey C.T. 1988. Expression of human adenosine deaminase in murine hematopoietic cells. 
Mol Cell Biol 8:5116-512 5 . 
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