A zebrafish ftz-F1 (Fushi tarazu factor 1) homologue requires multiple subdomains in the D and E regions for its transcriptional activity.
A zebrafish Ftz-F1 homologue, zFF1A (zebrafish Ff1a or Nr5a2, a member of nuclear receptor superfamily) and its C-terminally truncated variant (zFF1B) were previously identified. Due to lack of the identity box (I-box) and activation function 2 (AF-2) domain, zFF1B lacks transactivation function and fails to synergize with estrogen receptor (ER) in regulating promoters. It was speculated that the I-box might be involved in the zFF1A/ER interaction. In the present study, the function of the I-box was examined. In the absence of the I-box or with an altered heptad 9, the AF-2 of zFF1A was not functional, either in the presence or absence of ER. The GST pull-down assay showed that zFF1A and its mutants exerted similar physical contacts with ER-LBD, suggesting that the "dimerization" domain (I-box) is essential for the transcriptional activity of zFF1A. Moreover, nuclear receptor coactivator selectively activated zFF1 with the I-box but exerted no effect on zFF1B, indicating that the I-box is able to interact with the coactivators. By deletion study and analysis of the identified domains in GAL4-DNA binding domain, other regions of zFF1A critical for its AF were also delineated. Consistent with the mutation analysis, AF-2 was active only in the presence of the I-box. We also identified a novel AF domain (AF-3) located in the hinge region (amino acids 155-267), although the activity of AF-3 was inhibited by its flanking region. We suggest that the D and E regions of zFF1A possess both positive and negative transactivation functions, and interdomain "cross-talk" may confer the full transcriptional activity of the protein.
Liu
D
D
Division of Structural Biology and Biochemistry, Hospital for Sick Children, Toronto, Ontario M5G 1L5, Canada.
Chandy
M
M
Lee
S K
SK
Le Dréan
Y
Y
Ando
H
H
Xiong
F
F
Woon Lee
J
J
Hew
C L
CL
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017931:DNA Primers; D019281:Dimerization; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D015027:Zebrafish
D017931:DNA Primers; D051717:Fushi Tarazu Transcription Factors; D018398:Homeodomain Proteins