Therapeutic aspect of 14-3-3 related trypanosome-specific modulation with phosphorylation and dephosphorylation.

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We have previously shown that Trypanosoma brucei 14-3-3 I and II play pivotal roles in motility, cytokinesis and cell cycle (J.Biol.Chem.280, 14085-14096). We here report the unique dimerization pattern of 14-3-3 isoforms ; I forms both heterodimer and homodimer, whereas II forms mainly heterodimer. The α-helices 1-4 are responsible for dimeraization as in the cases of mammalian 14-3-3. In addition, we found unique N-terminal sequences of both isoforms existed in Trypoanosomatidae but not in mammals are indispensable for dimeraization. We further characterized that amino acid ^4Phe, ^6IIe, ^7Pro and ^<13>Leu in the N-terminal sequence of 14-3-3 II are crucial for dimeraization. 14-3-3 proteins act as phoshoserine-dependent molecular chaperones which recognize well-conserved motifs among eukaryotes. However, the precise mechanism by which T.brucei 14-3-3 controls cellular events have yet to be proven. We report that the protein recognition of T.brucei 14-3-3 is different from other eukaryotes ; 1)the surface plasmon resonance analysis reveals that 14-3-3 I but not II possesses weak affinity to the conserved phosphopeptides. 2)14-3-3II interacts with many proteins, not truly phosphorylation-dependent. We further searched how the conserved motifs are utilized, extensively in cultured T.brucei, 3)over expression of the human 14-3-3 isoform but not the mutated form, induces a drastic reduction in a growth rates and morphological changes that are observed in 14-3-3I and/or II knockdown cells, 4)the number of phosphorylated proteins that carry the conserved recognition motifs are small in T.brucei. The new pathway was suggested that phosphorylation would play a roll as so called dephosphorylation in a mirroring fashion.