The Xenopus B2 factor involved in TFIIIA gene regulation is closely related to Sp1 and interacts in a complex with USF.
In the Xenopus laevis oocyte there is a million fold more transcription factor IIIA (TFIIIA) and its corresponding mRNA than in a somatic cell. These high levels of TFIIIA gene expression are achieved primarily by transcriptional regulation. The TATA box along with three positive cis-elements in the control region of the TFIIIA gene located at positions -269 to -264 (E1), -235 to -220 (E2), and -669 to -636 (E3) are required for this high level of expression in oocytes. The proteins that bind E1 and E3 of the TFIIIA gene have been identified as Xenopus USF (Xl-USF) and B3 (homolog of Vg1 RBP/VERA). In this study the B2 protein was found to bind E2 in a zinc-dependent fashion and anti-human Sp1 (but not Sp2, Sp3, nor Sp4) supershifted the B2:element 2 complex. The E2 binding protein was purified by DNA affinity chromatography. Based on supershift analysis, molecular weight estimation experiments, and purified human Sp1 DNA binding affinity tests the data strongly support the idea that the B2 protein is the Xenopus ortholog of Sp1, but not Sp2, Sp3, nor Sp4. Xl-USF binds to element 1 of the TFIIIA gene which is immediately adjacent to element 2. Coimmunoprecipitation experiments using crude whole oocyte extracts revealed that Xenopus Sp1 and USF or closely related factors are present together in a high-affinity complex. This structure contributes positively to the initiation of TFIIIA gene transcription in Xenopus oocytes.
Penberthy
William T
WT
Molecular, Cellular, and Developmental Biology, UCLA, Los Angeles, CA 90095-1606, USA. wtpenber@ucla.edu
Griffin
David
D
Hall
Robert K
RK
Taylor
William L
WL
D000818:Animals; D000906:Antibodies / Q000276:immunology / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D004268:DNA-Binding Proteins; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D015151:Immunoblotting; D008970:Molecular Weight; D009841:Oligonucleotides / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D020218:Response Elements / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000276:immunology / Q000378:metabolism*; D035561:Transcription Factor TFIIA / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051720:Upstream Stimulatory Factors; D014982:Xenopus laevis / Q000235:genetics / Q000378:metabolism*; D016335:Zinc Fingers / Q000235:genetics
D000906:Antibodies; D004268:DNA-Binding Proteins; D009841:Oligonucleotides; D016329:Sp1 Transcription Factor; D035561:Transcription Factor TFIIA; D014157:Transcription Factors; C497181:USF1 protein, human; D051720:Upstream Stimulatory Factors