Destabilization of tetraplex structures of the fragile X repeat sequence (CGG)n is mediated by homolog-conserved domains in three members of the hnRNP family

Hairpin ortetrahelical structures formed by a d(CGG)(n) sequence in the FMR1 gene are thought to promote expansion of the repeat tract. Subsequent to this expansion FMR1 is silenced and fragile X syndrome ensues. The injurious effects of d(CGG), secondary structures may potentially be countered by agents that act to decrease their stability. We showed previously that the hnRNP-related protein CBF-A destabilized G'2 bimolecular tetraplex structures of d(CGG)(n). Analysis of mutant proteins revealed that the CBF-A-conserved domains RNP1(1) and ATP/GTP binding box were sufficient and necessary for G'2 d(CGG), disruption while the RNP2(1) motif inhibited the destabilization activity. Here, we report that a C-terminal fragment of CBF-A whose only remaining conserved domain was the ATP/GTP binding motif, disrupted G'2 d(CGG), more selectively than wildtype CBF-A. Further, two additional members of the hnRNP family, hnRNP A2 and mutant hnRNPA1 effectively destabilized G'2d(CGG)(n). Examination of mutant hnRNP A2 proteins revealed that, similar to CBF-A, their RNP1(1) element and ATP/GTP binding motif mediated G'2 d(CGG)(n) disruption, while the RNP2(1) element blocked their action. Similarly, the RNP1(1) and RNP2(1) domains of hnRNP All were, respectively, positive and negative mediators of G'2 d(CGG), destabilization. Last, employing the same conserved motifs that mediated disruption of the DNA tetraplex G'2 d(CGG)(n), hnRNP A2 destabilized r(CGG)(n) RNA tetraplex.