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PMID: 3352603 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

Effects of single-base substitutions within the Acanthamoeba castellanii rRNA promoter on transcription and on binding of transcription initiation factor and RNA polymerase I.

Molecular and cellular biology ·Vol. 8 ·No. 2 ·1988-02-00 ·Pages 747-53

Kownin P, Bateman E, Paule MR

Abstract

Single-point mutations were introduced into the promoter region of the Acanthamoeba castellanii rRNA gene by chemical mutagen treatment of a single-stranded clone in vitro, followed by reverse transcription and cloning of the altered fragment. The promoter mutants were tested for transcription initiation factor (TIF) binding by a template commitment assay plus DNase I footprinting and for transcription by an in vitro runoff assay. Point mutations within the previously identified TIF interaction region (between -20 and -47, motifs A and B) indicated that TIF interacts most strongly with a sequence centered at -29 and less tightly with sequences upstream and downstream. Some alterations of the base sequence closer to the transcription start site (and outside the TIF-protected site) also significantly decreased specific RNA synthesis in vitro. These were within the region which is protected from DNase I digestion by polymerase I, but these mutations did not detectably affect the binding of polymerase to the promoter.

MeSH Terms
Acanthamoeba/genetics Animals Deoxyribonuclease I Genes Mutation Promoter Regions, Genetic Protein Binding RNA Polymerase I/metabolism RNA, Ribosomal/genetics Templates, Genetic Transcription Factors/metabolism Transcription, Genetic
Chemicals
RNA, Ribosomal Transcription Factors RNA Polymerase I Deoxyribonuclease I
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Kownin P
Department of Biochemistry, Colorado State University, Fort Collins 80523.
Bateman E
Paule M R
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Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.8.2.747-753.1988
Published
1988-02-00
Pages
747-53
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC363200
Subset
IM
Grants
NIGMS NIH HHS · GM22580 · United States
NIGMS NIH HHS · GM26059 · United States
External Links
PubMed source DOI: 10.1128/mcb.8.2.747-753.1988 PMC Full Text View journal details
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