Two distinct domains of protein 4.1 critical for assembly offunctional nuclei in Vitro
Protein 4.1R, a multifunctional structural protein, acts asan adaptor in mature red cell membrane skeletons linking spectrin-actincomplexes to plasma membrane-associated proteins. In nucleated cellsprotein 4.1 is not associated exclusively with plasma membrane but isalso detected at several important subcellular locations crucial for celldivision. To identify 4.1 domains having critical functions in nuclearassembly, 4.1 domain peptides were added to Xenopus egg extract nuclearreconstitution reactions. Morphologically disorganized, replicationdeficient nuclei assembled when spectrin-actin binding domain orNuMA-binding C-terminal domain peptides were present. However, controlvariant spectrin-actin binding domain peptides incapable of bindingactin, or mutant C-terminal domain peptides with reduced NuMA binding,had no deleterious effects on nuclear reconstitution. To test if 4.1 isrequired for proper nuclear assembly, 4.1 isoforms were depleted withspectrin-actin binding or C-terminal domain-specific antibodies. Nucleiassembled in depleted extracts ha d deranged phenotypes. However, nuclearassembly could be rescued by addition of recombinant 4.1R. Our dataestablishes that protein 4.1 is essential for nuclear assembly andidentifies two distinct 4.1 domains, initially characterized incytoskeletal interactions, that have crucial and versatile functions innuclear assembly.
- Research Organization:
- Ernest Orlando Lawrence Berkeley NationalLaboratory, Berkeley, CA (US)
- Sponsoring Organization:
- National Institutes of Health
- DOE Contract Number:
- AC02-05CH11231
- OSTI ID:
- 860210
- Report Number(s):
- LBNL--48155; BnR: 400412000
- Journal Information:
- Journal Biological Chemistry, Journal Name: Journal Biological Chemistry Journal Issue: 46 Vol. 277
- Country of Publication:
- United States
- Language:
- English
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