Identification and characterization of a novel activated RhoB binding protein containing a PDZ domain whose expression is specifically modulated in thyroid cells by cAMP.
In a search for genes regulated in response to cAMP we have identified a new protein, p76RBE, whose mRNA and protein expression is enhanced in thyrocytes following thyrotropin stimulation of the cAMP transduction cascade. This protein presents important similarities with Rhophilin and contains different protein-protein interaction motifs. The presence of HR1 and PDZ motifs as well as a potential PDZ binding domain motif suggests that p76RBE could be implicated in targeting or scaffolding processes. By yeast two-hybrid screenings and coimmunoprecipitation, we show here that p76RBE is a specific binding protein of RhoB and binds selectively to the GTP-bound form of this small GTPase. p76RBE also binds in vitro to components of the cytoskeleton, including cytokeratin 18. p76RBE is essentially cytoplasmic in transfected COS-7 mammalian cells and seems to be recruited to an endosomal compartment when coexpressed with the activated form of RhoB. p76RBE was shown to be mainly expressed in tissues with high secretion activity. Our data suggest that p76RBE could play a key role between RhoB and potential downstream elements needed under stimulation of the thyrotropin/cAMP pathway in thyrocytes and responsible for intracellular motile phenomena such as the endocytosis involved in the thyroid secretory process.
Mircescu
Hortensia
H
Institute of Interdisciplinary Research, School of Medicine, Free University of Brussels, Belgium.
Steuve
Séverine
S
Savonet
Valérie
V
Degraef
Chantal
C
Mellor
Harry
H
Dumont
Jacques E
JE
Maenhaut
Carine
C
Pirson
Isabelle
I
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D019556:COS Cells; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D004285:Dogs; D004705:Endocytosis; D006160:Guanosine Triphosphate / Q000378:metabolism; D006801:Humans; D007633:Keratins / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D013961:Thyroid Gland / Q000166:cytology*; D013972:Thyrotropin / Q000378:metabolism; D014018:Tissue Distribution; D020798:Two-Hybrid System Techniques; D020743:rhoB GTP-Binding Protein / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; C471419:RHPN2 protein, human; D012333:RNA, Messenger; D007633:Keratins; D006160:Guanosine Triphosphate; D013972:Thyrotropin; D000242:Cyclic AMP; D020743:rhoB GTP-Binding Protein