Neuronal thread protein regulation and interaction with microtubule-associated proteins in SH-Sy5y neuronal cells

Abstract

In Alzheimer's disease (AD), neuronal thread protein (NTP) accumulates in cortical neurons and colocalizes with phospho- tau-immunoreactive cytoskeletal lesions that correlate with dementia. To generate additional information about the potential role of NTP in AD, we characterized its expression and regulation in human SH-Sy5y neuronal cells. Quantitative real-time reverse transcription-polymerase chain reactin and Western blot analysis demonstrated prominent insulin, moderate insulin-like growth factor, type 1 (IGF-1) and minimal nerve growth factor stimulation of NTP expression. In addition, NTP protein was more stable and it progressively accumulated in cells that were stimulated with insulin for 24 or 48 h. Metabolic labeling and phospho-amino acid analysis demonstrated phosphorylation of NTP on Serine residues, 30-60 min after insulin or IGF-1 stimulation, when glycogen synthase kinase 3beta (GSK-3beta) activity would no longer have been suppressed. Kinase inhibitor and in vitro phosphorylation studies demonstrated a role for GSK-3beta in the positive regulation of NTP expression and phosphorylation. Coimmunoprecipitation studies demonstrated physical interactions between NTP and tau or microtubule-associated protein 1b (MAP-1b), and ubiquitin immunoreactivity in NTP immunoprecipitates. In summary, these studies showed that (i) NTP expression is regulated at the level of transcription by insulin and IGF-1 stimulation; (ii) NTP is phosphorylated by GSK-3beta; (iii) NTP can physically interact with tau and MAP-1b and (iv) NTP-MAP complexes are ubiquitinated. The results suggest a functional role for NTP in relation to the turnover or processing of neuronal cytoskeletal proteins, attributes that may be modulated by insulin/IGF-1-mediated signaling.

Brain metabolic dysfunction at the core of Alzheimer's disease

“…GSK-3β [36,41] and reductions in tau gene expression [8,32,46]. Attendant failure to generate sufficient normal tau protein, vis-a-vis accumulation of hyper-phosphorylated insoluble fibrillar tau likely promotes cytoskeletal collapse, neurite retraction, and synaptic disconnection.…”

Insulin Resistance, Cognitive Impairment and Neurodegeneration: Roles of Nitrosamine Exposure, Diet and Lifestyles

“…In addition, tau hyper-phosphorylation in AD is mediated by inhibition of protein phosphatases 1 and 2A (Hanger, Seereeram et al 2009;Iqbal, Liu et al 2009;Morales, Farias et al 2010), which are regulated by insulin/IGF. Besides hyper-phosphorylation, tau pathology in AD is mediated by impaired tau gene expression due to reduced insulin and IGF signaling (de la Monte, Chen et al 2003). Consequences include failure to generate sufficient amounts of normal soluble tau protein, vis-a-vis accumulation of hyper-phosphorylated insoluble fibillar tau, and attendant exacerbation of cytoskeletal collapse, neurite retraction, and synaptic disconnection.…”

Pin1 Modulation in Physiological Status and Neurodegeneration. Any Contribution to the Pathogenesis of Type 3 Diabetes?

“…Insulin and IGFs levels were reduced in brain tissues of AD patients and this was associated with the reduced synthesis of tau [82,83]. Growth factors stimulated tau protein expression in neuroblastoma cells [83].…”

“…Growth factors stimulated tau protein expression in neuroblastoma cells [83]. The condition of brain insulin resistance was characterized by reduced levels of insulin and IGFs, no compensatory hyper-expression of their receptors and therefore a reduced transduction pathway [16,17,82].…”