The putative tumor suppressor RASSF1A homodimerizes and heterodimerizes with the Ras-GTP binding protein Nore1

Abstract

Nore and RASSF1A are noncatalytic proteins that share 50% identity over their carboxyterminal 300 AA, a segment that encompasses a putative Ras-Rap association (RA) domain. RASSF1 is expressed as several splice variants, each of which contain an RA domain, however the 340 AA RASSF1A, but not the shorter RASSF1C variant, is a putative tumor suppressor. Nore binds to Ras and several Ras-like GTPases in a GTP dependent fashion however neither RASSF1 (A or C) or the C. elegans Nore/RASSF1 homolog, T24F1.3 exhibit any interaction with Ras or six other Ras-like GTPases in a yeast two-hybrid expression assay. A low recovery of RASSF1A (but not RASSF1C) in association with RasG12V is observed however on transient expression in COS cells. Nore and RASSF1A can each e ciently homodimerize and heterodimerize with each other through their nonhomologous aminoterminal segments. Recombinant RASSF1C exhibits a much weaker ability to homodimerize or heterodimerize; thus the binding of RASSF1C to Nore is very much less than the binding of RASSF1A to Nore. The association of RASSF1A with RasG12V in COS cells appears to re¯ect the heterodimerization of RASSF1A with Nore, inasmuch the recovery of RASSF1A with RasG12V is increased by concurrent expression of full length Nore, and abolished by expression of Nore deleted of its RA domain. The preferential ability of RASSF1A to heterodimerize with Nore and thereby associate with Ras-like GTPases may be relevant to its putative tumor suppressor function.

Evaluation of the 3p21.3 tumour-suppressor gene cluster

“…For several RASSF members growth suppression is enhanced by co-transfection of constitutively active Ras proteins (Vos et al, 2000(Vos et al, , 2003a or by artificially forcing membrane localization thereby facilitating interaction with Ras (Eckfeld et al, 2004). Co-immunoprecipitation experiments show that RASSF proteins bind several Ras superfamily members as Ras-guanosine triphosphate (GTP) but not Ras-GDP and that this interaction occurs between the RA domain and the effector domain of Ras proteins (Vavvas et al, 1998;Vos et al, 2000Vos et al, , 2003aKhokhlatchev et al, 2002;Ortiz-Vega et al, 2002;Eckfeld et al, 2004). In the case of RASSF1A association with Ras may occur indirectly through heterodimerization with NORE1A or the scaffold protein CNK1 (Ortiz-Vega et al, 2002;Rabizadeh et al, 2004).…”

“…Co-immunoprecipitation experiments show that RASSF proteins bind several Ras superfamily members as Ras-guanosine triphosphate (GTP) but not Ras-GDP and that this interaction occurs between the RA domain and the effector domain of Ras proteins (Vavvas et al, 1998;Vos et al, 2000Vos et al, , 2003aKhokhlatchev et al, 2002;Ortiz-Vega et al, 2002;Eckfeld et al, 2004). In the case of RASSF1A association with Ras may occur indirectly through heterodimerization with NORE1A or the scaffold protein CNK1 (Ortiz-Vega et al, 2002;Rabizadeh et al, 2004). Taken together these data indicate the RASSF family of proteins are genuine Ras effectors that mediate some of the growth inhibitory functions of Ras proteins.…”

RASSF1A interacts with and activates the mitotic kinase Aurora-A

“…The homology of RASSF1A with the mammalian Ras effector novel Ras effector (NORE)1 suggests that the RASSF1A gene product may function in signal transduction pathways involving Ras-like proteins. However, recent data indicate that RASSF1A itself binds to RAS only weakly and that binding to RAS may require heterodimerization of RASSF1A and NORE1 (Ortiz-Vega et al, 2002). In addition, there is evidence for an association of both NORE1 and RASSF1A with the pro-apoptotic kinase MST1 and that this interaction is involved in apoptosis induced by activated Ras .…”

Frequent hypermethylation of the RASSF1A gene in prostate cancer

“…It was shown that RASSF1 binds Ras in a GTP-dependent manner and that its overexpression induces apoptosis (Vos et al, 2000). Our recent data indicate that the pro-apoptotic effect of RASSF1 may require heterodimerization with NORE1 and that RASSF1 itself binds to Ras only very weakly (Ortiz-Vega et al, 2002). In addition, there is evidence for an association of both NORE1 and RASSF1 with the pro-apoptotic kinase MST1, and that this interaction is involved in apoptosis induced by activated Ras .…”