Screening of human gene promoter activities using transfected-cell arrays.
Promoters are the best characterized transcriptional regulatory sequences in complex genomes because of their predictable location immediately upstream of transcription start sites. Despite a substantial body of literature describing transcriptional promoters, the identification of true start sites for all human transcripts is far from complete. The same is true of the key structural and functional elements responsible for promoter action in different cell types. In order to identify elements responsible for promoter activity, we applied transfected-cell array technology to functionally evaluate promoters for genes involved in inflammatory bowel disease. Seventy-four promoters were examined by reverse transfection of a promoter-fluorescent reporter constructs into a human embryonic kidney cell line (HEK293T). Sixteen (21.6%) promoters were found to be active in HEK293 T cells. Correlations between promoter activity and endogenous transcript level were calculated, and 75% of active promoters were found to be associated with transcriptional activity of their gene counterparts. These results provide experimental evidence of promoter activity, which may aid in understanding the regulation of gene expression. Moreover, this is the first large-scale functional study of regulatory sequences to use a high-throughput transfected-cell array technique.
Department of Vertebrate Genomics, Max Planck Institute for Molecular Genetics, 14195 Berlin, Germany.
Xi
X
Cheng
Anna
A
Guerasimova
Thomas
T
Manke
Philip
P
Rosenstiel
Stefan
S
Haas
Hans-Joerg
HJ
Warnatz
Robert
R
Querfurth
Wilfried
W
Nietfeld
Dominique
D
Vanhecke
Hans
H
Lehrach
Marie-Laure
ML
Yaspo
Michal
M
Janitz
D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation; D006801:Humans; D011401:Promoter Regions, Genetic / Q000235:genetics*; D046888:Tissue Array Analysis / Q000379:methods*; D014158:Transcription, Genetic; D014162:Transfection