Determinants of the brain-specific expression of the rat aldolase C gene: ex vivo and in vivo analysis

Authors


Correspondence to M. Thomas, CHU Cochin, ICGM U.129 INSERM, 24 rue du Fg St Jacques, F-75014 Paris, France
Fax:+33 1 44 41 24 21

Abstract

A 115-bp promoter fragment of the aldolase C gene is sufficient for conferring neural cell specificity on a reporter gene, in cultured PC12 cells and in transgenic mice. In vitro DNase I protection experiments detected two footprints on the promoter, termed boxes A/A', and B. The 5′ A/A' box contains overlapping Sp1 and Krox20/Krox24 binding sites; it binds Sp1 in fibroblasts (box A') and a different complex in brain (box A). Any deletion or mutation of this box that impairs protein recognition also suppresses promoter activity. The replacement of box A/A' by a Sp1 consensus binding site results in the loss of the brain specificity of expression in transgenic mice. Further 3′, box B is composed of a 5′ direct repeat and a 3′ GC box consisting of overlapping Sp1 and Krox20/Krox24 binding sites. Mutation of the direct repeat subregion appears to be more deleterious for the promoter activity than mutation of the G+C-rich subregion.

Abbreviations
CAT

chloramphenicol acetyltransferase

UMS

upstream mouse sequence

SV40

simian virus 40

PCR

polymerase chain reaction

DOTAP

N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium methylsulfate

Enzyme
 

Aldolase C, fructose-1,6-bisphosphate aldolase (EC 4.1.2.13)

Ancillary