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J. Biol. Chem., Vol. 276, Issue 40, 37011-37019, October 5, 2001
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EF1 Binds to a Far Upstream Sequence of the Mouse Pro-
1(I)
Collagen Gene and Represses Its Expression in Osteoblasts*
§,
,
, and
From the The transcription of type I collagen genes is
tightly regulated, but few cis-acting elements have been identified
that can modulate the levels of expression of these genes. Generation
of transgenic mice harboring various segments of the mouse pro-
INSERM U489 and Université Paris VI,
Paris, France and ¶ Cohesion Technologies,
Palo Alto, California 94303
1(I) collagen promoter led us to suspect that a repressor element was located between
10.5 and
17 kilobase pairs. Stable and
transient transfection experiments in ROS17/2.8 osteoblastic cells
confirmed the existence of such a repressor element at about
14
kilobase pairs and showed that it consisted in an almost perfect
three-time repeat of a 41-base pair sequence. This element, which we
named COIN-1, contains three E2-boxes, and a point mutation in at least two of them completely abolished its repressor effect. In gel shift
assays, COIN-1 bound a DNA-binding protein named
EF1/ZEB-1, and
mutations that abolished the repressor effect of COIN-1 also suppressed
the binding of
EF1. We also showed that the repressor effect of
COIN-1 was not mediated by chromatin compaction. Furthermore, overexpression of
EF1 in ROS17/2.8 osteoblastic cells enhanced the
inhibitory effect of COIN-1 in a dose-dependent manner and repressed the expression of the pro-
1(I) collagen gene. Thus,
EF1
appears to repress the expression of the mouse pro-
1(I) collagen
gene, through its binding to COIN-1.
To whom correspondence should be addressed: INSERM U489,
Hôpital TENON, 4 rue de la Chine, 75020 Paris, France. Tel.:
33-1-56-01-69-93; Fax: 33-1-56-01-69-99; E-mail:
jerome.rossert@tnn.ap-hop-paris.fr.
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