Akt-mediated phosphorylation increases the binding affinity of hTERT for importin α to promote nuclear translocation

Abstract

There was an error published in J. Cell Sci. 128, 2287-2301.In Fig. 2, the HA-importin-α3 immunoprecipitation and corresponding Flag-NLS blots were inadvertently duplicated as the HA-importin-α5 immunoprecipitation and corresponding Flag-NLS blots. The two HA-importin-α5 blots have been replaced with the correct images in panel D in the figure shown below. There are no changes to the figure legend, which is accurate. This error does not affect the conclusions of the study.The authors apologise to the readers for any confusion that this error might have caused. Fig. 2. The hTERT NLS interacts physically with importin α. (A) GST-importin-α1, GST-importin-α3, GST-importin-α5 and GST-importin-β1 were affinity purified and incubated with lysates that had been prepared from cells expressing Flag-NLS or Flag-S227A/7A, followed by immunoblotting with anti-Flag antibody. The purified GST fusion proteins were visualized by Coomassie staining and are indicated with arrowheads. Molecular mass makers are shown in kDa. (B) GST-NLS and GST-S227A/7A were purified and incubated with lysates prepared from cells expressing HA-importin-α1, HA-importin-α3, HA-importin-α5 and HA-importin-β1; the HA-tagged proteins were then detected. HA-tagged importin proteins are indicated with arrowheads. The purified GST fusion proteins were visualized by Coomassie staining. (C) MCF7 cells that had been transfected with Flag-NLS and either HA-importin-α proteins or HA-importin-β1 were subjected to immunoprecipitation (IP) with anti-Flag antibody, followed by immunoblotting with anti-HA antibody. HA-tagged importin proteins are indicated with arrowheads. (D) MCF7 cells transfected with Flag-NLS, HA-importin-α proteins and HA-importin-β1 were subjected to immunoprecipitation with anti-Flag antibody, followed by immunoblotting with anti-HA antibody. (E) MCF7 cells were subjected to immunoprecipitation with an anti-hTERT antibody; the immunoprecipitates were then probed for endogenous importin-α proteins and importin-β1. IgG antibody was used as a negative control. In B,C,D, asterisks mark the position of nonspecific immunoglobulin chains. ABSTRACTTelomeres are essential for chromosome integrity and protection, and their maintenance requires the ribonucleoprotein enzyme telomerase. Previously, we have shown that human telomerase reverse transcriptase (hTERT) contains a bipartite nuclear localization signal (NLS; residues 222-240) that is responsible for nuclear import, and that Akt-mediated phosphorylation of residue S227 is important for efficient nuclear import of hTERT. Here, we show that hTERT binds to importin-α proteins through the bipartite NLS and that this heterodimer then forms a complex with importin-β proteins to interact with the nuclear pore complex. Depletion of individual importin-α proteins results in a failure of hTERT nuclear import, and the resulting cytoplasmic hTERT is degraded by ubiquitin-dependent proteolysis. Crystallographic analysis reveals that the bipartite NLS interacts with both the major and minor sites of importin-α...

Telomere length and reactive oxygen species levels are positively associated with a high risk of mortality and recurrence in hepatocellular carcinoma

“…This study suggested that AKT pathway inhibitors such as perifosine could function as potential telomerase inhibitors . Several lines of evidence also suggested that blockade of AKT action inhibits telomerase activity, which could be translated into a promising approach for the treatment of human cancers . Here, we report that H 2 O 2 is a positive regulator of AKT‐mediated telomere elongation in liver HCC tumors.…”

“…Considerable evidence suggests that AKT increases telomerase activity by phosphorylating the TERT protein . Despite this evidence, there is a lack of clinical data and the existence of controversial data, such as a report showing that telomerase activity was not influenced by mutations in all of the putative AKT phosphorylation sites within TERT, which warrants further studies on the casual relationship of AKT and TERT. Our study also revealed that ROS induced telomere lengthening through AKT activation and pAKT inhibitors decreased telomere length and cancer cell proliferation.…”

Fucoidan Induces ROS‐Dependent Apoptosis in 5637 Human Bladder Cancer Cells by Downregulating Telomerase Activity via Inactivation of the PI3K/Akt Signaling Pathway

“…Telomerase activity can be tightly regulated by direct phosphorylation of hTERT linked to its nuclear localization by the PI3K/Akt signaling pathway [Kang et al, ; Chung et al, ; Jeong et al, ]. The PI3K/Akt pathway is also a well‐characterized cell survival signaling pathway, and the aberrant activation of this pathway is implicated in the initiation and progression of several types of human malignancies, including bladder cancer [Ching and Hansel, ; Bartholomeusz and Gonzalez‐Angulo, ; Houédé and Pourquier, ].…”

Towards an understanding of regulating Cajal body activity by protein modification

“…155 Furthermore, phosphorylation of TERT, especially by Akt at S227, has been shown to be important for the nuclear localization of TERT. 156,157 Despite these efforts, no studies have investigated how TERT PTMs regulate its association with the CB, or the delivery of telomerase by CBs to telomeres.…”