PuF, an antimetastatic and developmental signaling protein, interacts with the Alzheimer’s amyloid-β precursor protein via a tissue-specific proximal regulatory element (PRE)

Abstract

BackgroundAlzheimer’s disease (AD) is intimately tied to amyloid-β (Aβ) peptide. Extraneuronal brain plaques consisting primarily of Aβ aggregates are a hallmark of AD. Intraneuronal Aβ subunits are strongly implicated in disease progression. Protein sequence mutations of the Aβ precursor protein (APP) account for a small proportion of AD cases, suggesting that regulation of the associated gene (APP) may play a more important role in AD etiology. The APP promoter possesses a novel 30 nucleotide sequence, or “proximal regulatory element” (PRE), at −76/−47, from the +1 transcription start site that confers cell type specificity. This PRE contains sequences that make it vulnerable to epigenetic modification and may present a viable target for drug studies. We examined PRE-nuclear protein interaction by gel electrophoretic mobility shift assay (EMSA) and PRE mutant EMSA. This was followed by functional studies of PRE mutant/reporter gene fusion clones.ResultsEMSA probed with the PRE showed DNA-protein interaction in multiple nuclear extracts and in human brain tissue nuclear extract in a tissue-type specific manner. We identified transcription factors that are likely to bind the PRE, using competition gel shift and gel supershift: Activator protein 2 (AP2), nm23 nucleoside diphosphate kinase/metastatic inhibitory protein (PuF), and specificity protein 1 (SP1). These sites crossed a known single nucleotide polymorphism (SNP). EMSA with PRE mutants and promoter/reporter clone transfection analysis further implicated PuF in cells and extracts. Functional assays of mutant/reporter clone transfections were evaluated by ELISA of reporter protein levels. EMSA and ELISA results correlated by meta-analysis.ConclusionsWe propose that PuF may regulate the APP gene promoter and that AD risk may be increased by interference with PuF regulation at the PRE. PuF is targeted by calcium/calmodulin-dependent protein kinase II inhibitor 1, which also interacts with the integrins. These proteins are connected to vital cellular and neurological functions. In addition, the transcription factor PuF is a known inhibitor of metastasis and regulates cell growth during development. Given that APP is a known cell adhesion protein and ferroxidase, this suggests biochemical links among cell signaling, the cell cycle, iron metabolism in cancer, and AD in the context of overall aging.

Alzheimer's disease therapeutics targeted to the control of amyloid precursor protein translation: Maintenance of brain iron homeostasis

“…Additional evidence to connect APP over-expression with amyloidosis and dementia stems from the finding that known transcription mutations in the APP gene promoter may be critical to increase risk of AD pathology [68] Transcription factors like PUF-9 may contribute to increasing APP levels by generating sufficient template to alter the balance toward increased Aβ levels and amyloidosis [69], observations that are supported by the fact that most cases of familial AD result from APP processing by presenilins and BACE [70,71]. …”

Association between NME8 Locus Polymorphism and Cognitive Decline, Cerebrospinal Fluid and Neuroimaging Biomarkers in Alzheimer's Disease

“…NME8 (NM23 family member 8), a member of NM23 family, is known to be responsible for primary ciliary dyskinesia type 6 [14] . Although the relationship between NME8 and neurodegenerative diseases had not been well reported, proteins encoded by its family members had been reported to be associated with neurodegenerative disease in two papers [15] , [16] . In brain, nucleoside diphosphate kinase (NDPK), encoded by NM23 family, have been implicated to modulate neuronal cell proliferation, differentiation, and neurite outgrowth [15] , [17] , [18] .…”

“…Based on the fact that the specific activity of NDPK in brain was higher than in other tissues [19] , and its expression was accumulating preferentially in the nervous system during early embryonic development [20] , NDPK were predicted to play a pivotal role in the brain functions. Besides, nm23 nucleoside diphosphate kinase/metastatic inhibitory protein (PuF), encoded by nm34-H2 gene, may regulate the promoter of Aβ precursor protein gene (a gene that account for a part of familial AD) and that AD risk may be increased by interference with PuF regulation at the proximal regulatory element (PRE) [16] . All these potential connection between the NM23 family and AD may help to understand the underlying role of NME8 in AD.…”

The fallacy of placing confidence in confidence intervals

“…This is not a mere theoretical issue; moderately-sized p values often occur. In a cursory review of papers citing Steiger ( 2004 ), we found many that obtained and reported, without note, suspect confidence intervals bounded at 0 (e.g., Cumming, Sherar, Gammon, Standage, & Malina, 2012 ; Gilroy & Pearce 2014 ; Hamerman & Morewedge, 2015 ; Lahiri, Maloney, Rogers, & Ge, 2013 ; Hamerman & Morewedge, 2015 ; Todd, Vurbic, & Bouton, 2014 ; Winter et al, 2014 ). The others did not use confidence intervals, instead relying on point estimates of effect size and p values (e.g., Hollingdale & Greitemeyer, 2014 ); but from the p values it could be inferred that if they had followed “good practice” and computed such confidence intervals, they would have obtained intervals that according to Steiger could not be interpreted as anything but an inverted F test.…”