The role of human PinX1 in TRF1-mediated telomere protection

Abstract

Telomere maintenance is essential for chromosome stability. TRF1, a telomere binding protein, has a critical role in telomere protection. PinX1, identified as a TRF1 interacting protein, inhibits telomerase activity and affects tumorigenicity. However, little is known about the roles of the PinX1 on TRF1 function. Here we report that PinX1 regulates nucleolar accumulation, telomeric association of TRF1 and TRF1 protein stability. In HeLa, HA-PinX1 was co-localized with fibrillarin, a nucleolar protein, in 51% of the transfected cells and also resided in the nucleoplasm of the remaining 48%. Mutant analysis showed that the C-terminal region was important for nucleolar localization, while the N-terminus exhibited an inhibitory effect on nucleolar localization. Unlike HA- and Myc-PinX1, GFP-PinX1 resided predominantly in the nucleoli. Nuclear PinX1 bound to telomeres and other repeat sequences as well, but nucleolar PinX1 did not bind to telomeres, despite its interaction with TRF1. Nucleolar PinX1 forced endogenous TRF1 accumulation in the nucleoli. Furthermore, TRF1 binding to telomeres was upregulated in cells overexpressing PinX1. Depletion of PinX1 by siRNA resulted in rapid TRF1 degradation, enhanced TRF1 ubiquitination, less TRF1 binding to telomeres and increased telomere-dysfunction-induced foci (TIF). Overexpression of TRF1 compromised the reduction of TIF in PinX1-depleted cells, indicating that PinX1 plays a critical role in telomere protection by regulating TRF1 protein homeostasis. PinX1 also stabilized a mutant form of TRF1, a telomere-unbound form and exhibited the same extent of ability to the TRF1 stabilization in GCN5-depleted cells in which deubiquitination of TRF1 is inhibited, suggesting that PinX1 involves in TRF1 stability prior to ubquitin degradation pathway. In summary, PinX1 plays crucial roles in nucleolar accumulation of TRF1, TRF1 binding to telomere, TRF1 stability and telomere protection.