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BMC Cell Biol. 2009 Jan 28;10:7. doi: 10.1186/1471-2121-10-7.

Mucin 1 (MUC1) is a novel partner for MAL2 in breast carcinoma cells.

Author information

  • 1Molecular Oncology Laboratory, Oncology Research Unit, The Children's Hospital at Westmead, Locked Bag 4001, Westmead, 2145 NSW, Australia. sfanayan@cbms.mq.edu.au

Abstract

BACKGROUND:

The MAL2 gene, encoding a four-transmembrane protein of the MAL family, is amplified and overexpressed in breast and other cancers, yet the significance of this is unknown. MAL-like proteins have trafficking functions, but their molecular roles are largely obscure, partly due to a lack of known binding partners.

METHODS:

Yeast two-hybrid screening of a breast carcinoma cDNA expression library was performed using a full-length MAL2 bait, and subsequent deletion mapping experiments were performed. MAL2 interactions were confirmed by co-immunoprecipitation analyses and confocal microscopy was employed to compare protein sub-cellular distributions. Sucrose density gradient centrifugation of membranes extracted in cold Triton X-100 was employed to compare protein distributions between Triton X-100-soluble and -insoluble fractions.

RESULTS:

The tumor-associated protein mucin 1 (MUC1) was identified as a potential MAL2 partner, with MAL2/MUC1 interactions being confirmed in myc-tagged MAL2-expressing MCF-10A cells using co-immunoprecipitation assays. Deletion mapping experiments demonstrated a requirement for the first MAL2 transmembrane domain for MUC1 binding, whereas the MAL2 N-terminal domain was required to bind D52-like proteins. Confocal microscopy identified cytoplasmic co-localisation of MUC1 and MAL2 in breast cell lines, and centrifugation of cell lysates to equilibrium in sucrose density gradients demonstrated that MAL2 and MUC1 proteins were co-distributed between Triton X-100-soluble and -insoluble fractions. However co-immunoprecipitation analyses detected MAL2/MUC1 interactions in Triton X-100-soluble fractions only. Myc-MAL2 expression in MCF-10A cells was associated with both increased MUC1 detection within Triton X-100-soluble and -insoluble fractions, and increased MUC1 detection at the cell surface.

CONCLUSION:

These results identify MUC1 as a novel MAL2 partner, and suggest a role for MAL2 in regulating MUC1 expression and/or localisation.

PMID:
19175940
[PubMed - indexed for MEDLINE]
PMCID:
PMC2644682
Free PMC Article
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