Cytoplasmic Retention of Protein Phosphatase 2A Inhibitor 2 (I2PP2A) Induces Alzheimer-like Abnormal Hyperphosphorylation of Tau

Mohammad Arif,Jianshe Wei,Qi Zhang,Fei Liu,Gustavo Basurto-Islas,Inge Grundke-Iqbal,Khalid Iqbal

doi:10.1074/jbc.m114.565358

Abstract

Abnormal hyperphosphorylation of Tau leads to the formation of neurofibrillary tangles, a hallmark of Alzheimer disease (AD), and related tauopathies. The phosphorylation of Tau is regulated by protein phosphatase 2A (PP2A), which in turn is modulated by endogenous inhibitor 2 (I2 (PP2A)). In AD brain, I2 (PP2A) is translocated from neuronal nucleus to cytoplasm, where it inhibits PP2A activity and promotes abnormal phosphorylation of Tau. Here we describe the identification of a potential nuclear localization signal (NLS) in the C-terminal region of I2 (PP2A) containing a conserved basic motif, (179)RKR(181), which is sufficient for directing its nuclear localization. The current study further presents an inducible cell model (Tet-Off system) of AD-type abnormal hyperphosphorylation of Tau by expressing I2 (PP2A) in which the NLS was inactivated by (179)RKR(181) → AAA along with (168)KR(169) → AA mutations. In this model, the mutant NLS (mNLS)-I2 (PP2A) (I2 (PP2A)AA-AAA) was retained in the cell cytoplasm, where it physically interacted with PP2A and inhibited its activity. Inhibition of PP2A was associated with the abnormal hyperphosphorylation of Tau, which resulted in microtubule network instability and neurite outgrowth impairment. Expression of mNLS-I2 (PP2A) activated CAMKII and GSK-3β, which are Tau kinases regulated by PP2A. The immunoprecipitation experiments showed the direct interaction of I2 (PP2A) with PP2A and GSK-3β but not with CAMKII. Thus, the cell model provides insights into the nature of the potential NLS and the mechanistic relationship between I2 (PP2A)-induced inhibition of PP2A and hyperphosphorylation of Tau that can be utilized to develop drugs preventing Tau pathology.

Highlights

In Alzheimer brain, I2PP2A is translocated from the neuronal nucleus to the cytoplasm and promotes abnormal hyperphosphorylation of Tau
Mutant nuclear localization signal (NLS)-I2PP2A Translocates to the Cell Cytoplasm from the Nucleus and Inhibits phosphatase 2A (PP2A) Activity—We previously reported that I2PP2A is translocated from neuronal nucleus to the cytoplasm, co-localizes with PP2A, and promotes abnormal hyperphosphorylation of Tau by inhibiting PP2A in Alzheimer disease (AD) brain [27, 28], suggesting the critical role of I2PP2A translocation and its association with PP2A in the pathogenesis of AD
Tauopathies are age-associated neurodegenerative diseases characterized by neurofibrillary pathology made up of abnormally hyperphosphorylated Tau, and the severity of these lesions directly correlates with dementia

Summary

Background

In Alzheimer brain, I2PP2A is translocated from the neuronal nucleus to the cytoplasm and promotes abnormal hyperphosphorylation of Tau. In AD brain, I2PP2A is translocated from neuronal nucleus to cytoplasm, where it inhibits PP2A activity and promotes abnormal phosphorylation of Tau. Here we describe the identification of a potential nuclear localization signal (NLS) in the C-terminal region of I2PP2A containing a conserved basic motif, 179RKR181, which is sufficient for directing its nuclear localization. We report a PC12 stably expressing human Tau441 cell line that allows the inducible expression of mNLS-I2PP2A (168KR169 3 AA/179RKR181 3 AAA) based on the Tet-Off system Employing this cell line, we found that cytoplasmic accumulation of I2PP2A is associated with inhibition of PP2A activity and activation of Tau kinases regulated by PP2A, hyperphosphorylation of Tau, and promotion of microtubule instability. The mNLS-I2PP2A cell model can be useful in screening of therapeutic drugs for AD and other tauopathies

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION