Identification of DNA-binding proteins that recognize a Conserved Type I repeat sequence in the replication origin region of Tetrahymena rDNA

Date
1994
Authors
Umthun, Angela
Hou, Zhen
Sibenaller, Zita
Shaiu, Wen-Ling
Dobbs, Drena
Dobbs, Drena
Major Professor
Advisor
Committee Member
Journal Title
Journal ISSN
Volume Title
Publisher
Altmetrics
Authors
Research Projects
Organizational Units
Zoology and Genetics
Organizational Unit
Journal Issue
Series
Department
Zoology and GeneticsMolecular, Cellular and Developmental BiologyGenetics
Abstract

An origin of DNA replication has been mapped within the 5' non-transcribed spacer region of the amplified macronuclear rRNA genes (rDNA) of Tetrahymena thermophila. Mutations in 33 nt conserved AT-rich Type I repeat sequences located in the origin region cause defects in the replication and/or maintenance of amplified rDNA in vivo. Fe(ll)EDTA cleavage footprinting of restriction fragments containing the Type I repeat showed that most of the conserved nucleotides were protected by proteins in extracts of Tetrahymena cells. Two classes of proteins that bound the Type I repeat were identified and characterized using synthetic oligonucleotides in electrophoretic mobility shift assays. One of these, ds-TIBF, bound preferentially to duplex DNA and exhibited only moderate specificity for Type I repeat sequences. In contrast, a single-stranded DNA-binding protein, ssATIBF, specifically recognized the A-rich strand of the Type I repeat sequence. Deletion of the 5′ or 3′ borders of the conserved sequence significantly reduced binding of ssA-TIBF. The binding properties of ssATIBF, coupled with genetic evidence that Type I sequences function as c/s-acting rDNA replication control elements in vivo, suggest a possible role for ssA-TIBF in rDNA replication in Tetrahymena.

Comments

This article is from Nucleic Acids Research 22 (1994): 4432, doi: 10.1093/nar/22.21.4432. Posted with permission.

Description
Keywords
Citation
DOI
Collections