title: Regulation of BLM Nucleolar Localization.
abstract: Defects in coordinated ribosomal RNA (rRNA) transcription in the nucleolus cause cellular and organismal growth deficiencies. Bloom's syndrome, an autosomal recessive human disorder caused by mutated recQ-like helicase BLM, presents with growth defects suggestive of underlying defects in rRNA transcription. Our previous studies showed that BLM facilitates rRNA transcription and interacts with RNA polymerase I and topoisomerase I (TOP1) in the nucleolus. The mechanisms regulating localization of BLM to the nucleolus are unknown. In this study, we identify the TOP1-interaction region of BLM by co-immunoprecipitation of in vitro transcribed and translated BLM segments and show that this region includes the highly conserved nuclear localization sequence (NLS) of BLM. Biochemical and nucleolar co-localization studies using site-specific mutants show that two serines within the NLS (S1342 and S1345) are critical for nucleolar localization of BLM but do not affect the functional interaction of BLM with TOP1. Mutagenesis of both serines to aspartic acid (phospho-mimetic), but not alanine (phospho-dead), results in approximately 80% reduction in nucleolar localization of BLM while retaining the biochemical functions and nuclear localization of BLM. Our studies suggest a role for this region in regulating nucleolar localization of BLM via modification of the two serines within the NLS.
authors.affiliation: Department of Cancer Biology and Genetics, College of Medicine, The Ohio State University, Columbus, OH 43210, USA. larissa.tangeman@osumc.edu.
authors.forename: Larissa
authors.initials: L
authors.lastname: Tangeman
authors.affiliation: Department of Cancer Biology and Genetics, College of Medicine, The Ohio State University, Columbus, OH 43210, USA. michael.mcilhatton@osumc.edu.
authors.forename: Michael A
authors.initials: MA
authors.lastname: McIlhatton
authors.affiliation: Divisions of Hematology and Medical Oncology, Department of Internal Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA. grierson@wustl.edu.
authors.forename: Patrick
authors.initials: P
authors.lastname: Grierson
authors.affiliation: Department of Cancer Biology and Genetics, College of Medicine, The Ohio State University, Columbus, OH 43210, USA. joanna.groden@osumc.edu.
authors.forename: Joanna
authors.initials: J
authors.lastname: Groden
authors.affiliation: Department of Cancer Biology and Genetics, College of Medicine, The Ohio State University, Columbus, OH 43210, USA. samir.acharya@osumc.edu.
authors.forename: Samir
authors.initials: S
authors.lastname: Acharya
keywords: BLM; Bloom’s syndrome; growth defects; nucleolar localization; rRNA transcription; topoisomerase I