| Autor: |
Roberto J; Department of Chemistry and Biochemistry, University of Windsor, 401 Sunset Avenue, Windsor, Ontario N9B 3P4, Canada., Sykes CE; Department of Chemistry and Biochemistry, University of Windsor, 401 Sunset Avenue, Windsor, Ontario N9B 3P4, Canada., Vacratsis PO; Department of Chemistry and Biochemistry, University of Windsor, 401 Sunset Avenue, Windsor, Ontario N9B 3P4, Canada. |
| Abstrakt: |
The transcriptional co-activator with the PDZ binding motif (TAZ) is a critical regulator of numerous cellular processes such as cell differentiation, development, proliferation, and cell growth. Aberrant expression and activity of TAZ are also featured in many human malignancies. A hallmark of TAZ biology is its cytoplasmic retention mediated by 14-3-3 isoforms in response to phosphorylation of Ser 89 by members of the LATS family of kinases. Following the observation that TAZ is a highly phosphorylated protein even when Ser 89 is mutated, high-resolution mass spectrometry employing data-independent acquisition and ion mobility separation was conducted to elucidate additional TAZ phosphorylation sites that may play a role in regulating this critical transcriptional rheostat. Numerous phosphorylation sites on TAZ were identified, including several novel modifications. Of notable interest was the identification of positional phosphoisomers on a phosphopeptide containing Ser 89 . Optimized use of a so-called wideband enhancement acquisition technique yielded higher-quality fragmentation data that confirmed the detection of Ser 93 as the positional phosphoisomer partner of Ser 89 and identified diagnostic fragment ions for the phosphorylation events. Functional analysis indicated that Ser 93 phosphorylation reduces the level of 14-3-3 association and increases the level of nuclear translocation, indicating this phosphorylation event attenuates the 14-3-3-mediated TAZ cytoplasmic retention mechanism. These findings suggest that the biological activities of TAZ are likely dynamically regulated by multisite phosphorylation. |
