Nucleic Acids Research, Vol 25, Issue 15 3175-3182, Copyright © 1997 by Oxford University Press
CA Seid, RK Ramachandran, JM George, V Govindarajan, MF Gonzalez-Rimbau, CN Flytzanis and CR Tomlinson
The extracellular matrix (ECM) has been shown to play an important role in
development and tissue-specific gene expression, yet the mechanism by which
genes receive signals from the ECM is poorly understood. The aboral
ectoderm-specific LpS1-alpha and -beta genes of Lytechinus pictus , members
of the Spec gene family, provide an excellent model system to study ECM-
mediated gene regulation. Disruption of the ECM by preventing collagen
deposition using the lathrytic agent beta- aminopropionitrile (BAPN)
inhibits LpS1 gene transcription. LpS1 transcription resumes after removal
of BAPN and subsequent collagen reformation. Using a chloramphenicol
acetyltransferase (CAT) reporter gene assay, we show that a 125 bp region
of the LpS1-beta promoter from -108 to +17 contains an ECM response element
(ECM RE). Insertion of the 125 bp region into the promoter of the
metallothionein gene of L. pictus, a gene unaffected by ECM disruption,
caused the fused promoter to become ECM dependent. As with the endogenous
LpS1 genes, CAT activity directed by the fused LpS1-beta promoter resumed
in embryos recovered from ECM disruption. A mutation in a cis -acting
element called the proximal G-string, which lies in the 125 bp region,
caused CAT activity levels in ECM-disrupted embryos to equal that of the
wild- type LpS1-bet apromoter in ECM-intact embryos. These results suggest
that the intact ECM normally transmits signals to inhibit repressor
activity at the proximal G-string in aboral ectoderm cells. Consistent with
these results were our findings which showed that in addition to expression
in the aboral ectoderm, the proximal G-string mutation caused expression of
the CAT gene in oral ectoderm cells. These studies suggested that the
proximal G-string serves as a binding site for negative regulation of the
LpS1 genes in oral ectoderm during development. We also examined trans
-acting factors binding the proximal G-string following ECM disruption.
Band shift gels revealed a predominant set of slower migrating nuclear
proteins from ECM-disrupted embryos which bound the proximal G-string. This
work suggested that ECM disruption initiates signaling that induces a
repressor to bind the ECM RE and/or modifies ECM RE binding proteins, which
in turn represses LpS1 gene activity.
ARTICLES
An extracellular matrix response element in the promoter of the LpS1 genes of the sea urchin Lytechinus pictus
Department of Biology, University of Houston, Houston, TX 77204-5513, USA.
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