Identification of functional domains involved in BTG1 cell localization.
We have previously shown that BTG1 stimulates myoblast differentiation. In addition, this protein displays a major nuclear localization in confluent myoblasts, decreasing during the early steps of differentiation, and is essentially detected in the cytoplasm of mature myotubes. To identify the domains involved in the cellular trafficking of BTG1, we observed the localization of several BTG1 sequences fused to betaGalactosidase. The highly conserved B box among all members of the BTG family induces a significant nuclear localization of the betaGal moiety, enhanced by presence of the BTG1 carboxy-terminal sequence. In addition, a functional Nuclear Export Signal (NES) overlaps the B box. Moreover, presence of the first 43 NH(2)-terminal amino acids reduced the nuclear localization of each chimeric protein tested. Last, the BTG1 amino-terminal domain bears an LxxLL motif favouring nuclear accumulation, and another region encompassing the A box inhibiting nuclear localization. In contrast to a BTG1 mutant exclusively localized in the cytoplasm, transient expression of a mutant displaying a nuclear localization enhanced myoblasts withdrawal from the cell cycle and terminal differentiation, thus mimicking the myogenic influence of BTG1. In conclusion, several regions of BTG1 are implicated in its cellular localization, and BTG1 myogenic activity is induced at the nuclear level.
Rodier
A
A
UMR Différenciation Cellulaire et Croissance (INRA, Université Montpellier II, ENSAM), Unité d'Endocrinologie Cellulaire, INRA, 2 place Viala, 34060 Montpellier Cedex 1, France.
Rochard
P
P
Berthet
C
C
Rouault
J P
JP
Casas
F
F
Daury
L
L
Busson
M
M
Magaud
J P
JP
Wrutniak-Cabello
C
C
Cabello
G
G
D021581:Active Transport, Cell Nucleus / Q000502:physiology; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D017124:Conserved Sequence; D006801:Humans; D008969:Molecular Sequence Data; D009132:Muscles / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D017434:Protein Structure, Tertiary; D011784:Quail; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D009363:Neoplasm Proteins; D011993:Recombinant Fusion Proteins; C074086:BTG1 protein, human; D001616:beta-Galactosidase