Thesis
The nuclear actions of IGFBP-3
Washington State University
Master of Science (MS), Washington State University
2006
Handle:
https://hdl.handle.net/2376/554
Abstract
IGF-binding protein (IGFBP)-3 has intrinsic antiproliferative and proapoptotic functions that are independent of IGF binding and may involve nuclear localization. We determined that exogenous IGFBP-3 rapidly translocates to myoblast nuclei and that a 22-residue peptide containing the metal binding domain (MBD) and nuclear localization sequence (NLS) can similarly direct chimeric GFP into myoblast nuclei. Furthermore, a non-IGF-binding IGFBP-3 mutant inhibited myoblast proliferation without stimulating apoptosis. These results suggest that IGFBP-3 inhibits muscle cell growth in an IGFindependent manner that may be influenced by its rapid nuclear localization. We therefore identified IGFBP-3 interacting proteins by screening a rat L6 myoblast cDNA library using the yeast two-hybrid assay and two N-terminal deletion mutants as bait: BP3/231 (231 residues, L61 to K291) and BP3/ 111 (K181 to K291). Proteins previously known to interact with IGFBP-3 as well as several novel proteins were identified, including RNA polymerase II binding subunit 3 (Rpb3). The domain necessary for Rpb3 binding was subsequently identified using different IGFBP-3 deletion mutants and was localized to the MBD/NLS epitope. Rpb3/IGFBP-3 binding was confirmed by coimmunoprecipitation assays with specific antisera, whereas a NLS mutant IGFBP-3 did not associate with Rpb3, suggesting that a functional NLS is required. Rpb3 facilitates recruitment of the polymerase complex to specific transcription factors and is necessary for the transactivation of many genes. Its association of Rpb3 with IGFBP-3 provides a functional role for IGFBP-3 in the direct modulation of gene transcription.
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Details
- Title
- The nuclear actions of IGFBP-3
- Creators
- Wan-Jung Lin
- Contributors
- Buel Dan Rodgers (Degree Supervisor)
- Awarding Institution
- Washington State University
- Academic Unit
- Animal Sciences, Department of
- Theses and Dissertations
- Master of Science (MS), Washington State University
- Publisher
- Washington State University; Pullman, Wash. :
- Identifiers
- 99900525031001842
- Language
- English
- Resource Type
- Thesis