BRAIN AND TESTIS SELECTIVE EXPRESSION OF THE GLUTATHIONE-S-TRANSFERASE YB-3 SUBUNIT IS GOVERNED BY TANDEM DIRECT REPEAT OCTAMER MOTIFS IN THE 5'-FLANKING REGION OF ITS GENE
M. Abramovitz et al., BRAIN AND TESTIS SELECTIVE EXPRESSION OF THE GLUTATHIONE-S-TRANSFERASE YB-3 SUBUNIT IS GOVERNED BY TANDEM DIRECT REPEAT OCTAMER MOTIFS IN THE 5'-FLANKING REGION OF ITS GENE, Molecular brain research, 28(1), 1995, pp. 37-46
To gain insight into mechanisms of cell type-specific transcription of
class mu-glutathione S-transferase genes, the gene encoding the Yb-3
subunit was cloned. Yb-3 subunits are selectively expressed at high le
vels in rat brain and testis but not in liver or kidney. The Yb-3 subu
nit gene spans over 6 kb and consists of 8 exons and 7 introns and a s
equence consisting of tandem direct repeat consensus octamer DNA bindi
ng motifs separated by a 6 base pair (bp) spacer was identified in its
5'-flanking region. Gel shift assays with a 40 bp segment of DNA cont
aining the two consensus octamer sequences, revealed the presence of s
pecific binding proteins in nuclear extracts of rat brain, testis and
C6 glioma cells. DNA binding activity was greatly reduced in liver, ki
dney and HTC cells. Reporter vectors carrying segments of the 5'-flank
ing region of the Yb-3 subunit gene fused to a luciferase gene were in
troduced into C6 glioma cells which express high levels of Yb-3 subuni
ts, and into HTC cells which do not. The plasmids consisting of the Yb
-3 gene promoter up to, but not including, the octamer motifs did not
support luciferase transcription in the C6 glioma cells, but larger fr
agments that included the octamer repeat sequences, effectively direct
ed transcription in the C6 glioma cells. With mutated octameric sequen
ces transcriptional activity was greatly reduced, and none of the same
Yb-3 constructs directed substantial luciferase transcription in the
HTC cells. The results show that octamer motifs in the 5'-flanking reg
ion of the Yb-3 subunit gene are functional and are the principal cis-
acting elements that account for its discrete cell type-selective expr
ession. This gene is one of the few known targets for octamer DNA bind
ing transcription factors in brain.