Phosphorylation and microtubule association of the Opitz syndrome protein mid-1 is regulated by protein phosphatase 2A via binding to the regulatory subunit α4

Jun Liu, Todd D. Prickett, Elizabeth Elliott, G. Meroni, David L. Brautigan

Research output: Contribution to journalArticle

85 Scopus citations

Abstract

Opitz syndrome (OS) is a human genetic disease characterized by deformities such as cleft palate that are attributable to defects in embryonic development at the midline. Gene mapping has identified OS mutations within a protein called Mid1. Wild-type Mid1 predominantly colocalizes with microtubules, in contrast to mutant versions of Mid1 that appear clustered in the cytosol. Using yeast two-hybrid screening, we found that the α4-subunit of protein phosphatases 2A/4/6 binds Mid1. Epitope-tagged α4 coimmunoprecipitated endogenous or coexpressed Mid1 from COS7 cells, and this required only the conserved C-terminal region of α4. Localization of Mid1 and α4 was influenced by one another in transiently transfected cells. Mid1 could recruit α4 onto microtubules, and high levels of α4 could displace Mid1 into the cytosol. Metabolic 32P labeling of cells showed that Mid1 is a phosphoprotein, and coexpression of full-length α4 decreased Mid1 phosphorylation, indicative of a functional interaction. Association of green fluorescent protein-Mid1 with microtubules in living cells was perturbed by inhibitors of MAP kinase activation. The conclusion is that Mid1 association with microtubules, which seems important for normal midline development, is regulated by dynamic phosphorylation involving MAP kinase and protein phosphatase that is targeted specifically to Mid1 by α4. Human birth defects may result from environmental or genetic disruption of this regulatory cycle.

Original languageEnglish (US)
Pages (from-to)6650-6655
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume98
Issue number12
DOIs
StatePublished - Jun 5 2001

ASJC Scopus subject areas

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