J Biol Chem, Vol. 274, Issue 17, 11773-11781, April 23, 1999

A Pivotal Role for the Transmembrane Domain in Transforming Growth Factor- Receptor Activation

From the Ludwig Institute for Cancer Research, Post Office, Royal Melbourne Hospital, Victoria 3050, Australia

Transforming growth factor- (TGF-) delivers diverse growth and differentiation signals by binding two distantly related transmembrane serine/threonine kinase receptors: the type I receptor (TRI) and the type II receptor (TRII). In an attempt to establish the role of the transmembrane domain in receptor signaling, two chimeric TGF- receptors, TRI-II-I and TRII-I-II, containing the opposite transmembrane domain were generated. When transfected into a mutant mink lung epithelial cell line R1B, which lacks functional TRI, TRI-II-I restored TGF-1-induced transcriptional activation of a TGF- reporter p3TP-Lux to ~25% of the levels restored by wild-type TRI. In the mutant mink lung epithelial cell line DR26, which contains a truncated, nonfunctional TRII, wild-type receptor TRII restored the TGF- responsiveness, while the TRII-I-II cDNA was inactive. When both TRI and TRII were transfected into R1B, DR26, or Mv1Lu cells, a low level of constitutive p3TP-Lux activity was observed. However, cotransfection of both transmembrane chimeric receptors, TRI-II-I and TRII-I-II, or the wild-type TRI with the transmembrane chimeric TRII-I-II resulted in high levels of ligand-independent receptor activation. These results suggest that the transmembrane domains of both TGF- receptors are essential and play a pivotal role in receptor activation. To investigate the role of the transmembrane domain further, four type II transmembrane mutants were generated: TRII-1, TRII-2, TRII-3, and TRII-4, which have one, two, three, or four amino acids deleted at the N terminus of the transmembrane domain, respectively. Interestingly, co-expression of TRII-1 with the wild-type TRI in DR26 cells resulted in high levels of constitutive activation, while only low levels of the activation were observed when TRII-2, TRII-3, or TRII-4 were co-expressed with the wild-type TRI. However, TRII-1 restored very little the TGF- responsiveness in DR26cells. Expression of TRII-2, TRII-3, and TRII-4 resulted in a progressive increase in TGF- responsiveness, with TRII-4 reaching the level of activity of the wild-type TRII. Furthermore, like TRII-I-II, co-expression of TRII-1 with TRI-II-I also resulted in high levels of constitutive activation. These results are consistent with an important role for the transmembrane region of the receptors. We further propose a model of receptor activation in which receptor activation occurs via relative orientational rotation.