tailieunhanh - Báo cáo khoa học: Analysis of the regulatory motifs in eukaryotic initiation factor 4E-binding protein 1

Mammalian target of rapamycin complex 1 (mTORC1) phosphorylates proteins such as eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) and the S6 kinases. These substrates contain short sequences, termed TOR signalling (TOS) motifs, which interact with the mTORC1 component rap-tor. | ỊFEBS Journal Analysis of the regulatory motifs in eukaryotic initiation factor 4E-binding protein 1 Vivian H. Y. Lee1 Timothy Healy1 Bruno D. Fonseca1 Amanda Hayashi2 and Christopher G. Proud1 1 Department of Biochemistry and Molecular Biology University of British Columbia Vancouver Canada 2 Institute of Food Nutrition and Human Health Massey University and Food Metabolism and Microbiology AgResearch Limited Palmerston North New Zealand Keywords 4E-BP1 mTOR mTORC1 RAIP motif TOS motif Correspondence C. G. Proud Department of Biochemistry and Molecular Biology University of British Columbia Life Sciences Centre 2350 Health Sciences Mall Vancouver V6T 1Z3 BC Canada Fax 1 604 822 5227 Tel 1 604 827 3923 E-mail cgpr@ Website http fac_research faculty Received 10 December 2007 revised 22 February 2008 accepted 3 March 2008 doi Mammalian target of rapamycin complex 1 mTORCl phosphorylates proteins such as eukaryotic initiation factor 4E-binding protein 1 4E-BP1 and the S6 kinases. These substrates contain short sequences termed TOR signalling TOS motifs which interact with the mTORC1 component raptor. Phosphorylation of 4E-BP1 requires an additional feature termed the RAIP motif Arg-Ala-Ile-Pro . We have analysed the interaction of 4E-BP1 with raptor and the amino acid residues required for functional RAIP and TOS motifs as assessed by raptor binding and the phosphorylation of 4E-BP1 in human cells. Binding of 4E-BP1 to raptor strongly depends on an intact TOS motif but the RAIP motif and additional C-terminal features of 4E-BP1 also contribute to this interaction. Mutational analysis of 4E-BP1 reveals that isoleucine is a key feature of the RAIP motif that proline is also very important and that there is greater tolerance for substitution of the first two residues. Within the TOS motif the first position phenylalanine in the known motifs is most critical whereas a wider range of residues