Activity-dependent Neuroprotective Protein Constitutes a Novel Element in the SWI/SNF Chromatin Remodeling Complex

Abstract

Complete deficiency in activity-dependent neuroprotective protein (ADNP), a heterochromatin 1-binding protein, results in dramatic changes in gene expression, neural tube closure defects, and death at gestation day 9 in mice. To further understand the cellular roles played by ADNP, the HEK293 human embryonic kidney cell line that allows efficient transfection with recombinant DNA was used as a model for the identification of ADNP-interacting proteins. Recombinant green fluorescent protein (GFP)-ADNP was localized to cell nuclei. When nuclear extracts were subjected to immunoprecipitation with specific GFP antibodies followed by polyacrylamide gel electrophoresis, several minor protein bands were observed in addition to GFP-ADNP. In-gel protein digests followed by mass spectrometry identified BRG1, BAF250a, and BAF170, all components of the SWI/SNF (mating type switching/sucrose nonfermenting) chromatin remodeling complex, as proteins that co-immunoprecipitate with ADNP. These results were verified utilizing BRG1 antibodies. ADNP short hairpin RNA down-regulation resulted in microtubule reorganization and changes in cell morphology including reduction in cell process formation and cell number. These morphological changes are closely associated with the SWI/SNF complex multifunctionality. Taken together, the current study uncovers a molecular basis for the essential function of the ADNP gene and protein.Activity-dependent neuroprotective protein (ADNP 3 (1)) was originally discovered as a gene product associated with neuroprotection/neuroglia interactions (2). ADNP immunoreactivity was shown to localize to the astrocyte nucleus as well as to the cytoplasm and to the extracellular space, where the protein enhances neuronal survival (3, 4). Further studies suggested very high gene conservation in mammals including humans, increased expression in malignant cells, and close association with cellular survival (5). In the mature brain, ADNP expression was modulated by injury, suggesting a potential protective effect for the protein in vivo (6, 7). Additionally, ADNP expression shows sexual dichotomy and is modulated during the estrous cycle in the arcuate nucleus of the hypothalamus (8) and in the vagina (9), suggesting a potential neurotrophic/plasticity-associated effect for the protein.Importantly, complete deficiency in ADNP results in neural tube closure defects and death at gestation day 9 in mice (10). ADNP-deficient embryos exhibit dramatic increases in gene transcripts associated with lipid metabolism coupled to a reduction in organogenesis/neurogenesis related transcripts (11). In pluripotent P19 cells, ADNP was shown to interact with specific chromatin regions in the neuro-differentiated state, which was associated with binding to heterochromatin protein 1 (11). To further understand ADNP function, the HEK293 human embryonic kidney cell line that allows efficient transfection with recombinant DNA was used as a model for the identification of ADNP-interacting proteins and cellular function, suggesting that ADN...

Addressing Alzheimers Disease Tangles: From NAP to AL-108

“…Consistent with these findings, recent work has shown that ADNP differentially interacts with chromatin to regulate essential genes [25][26][27][28] and ADNP was found to play a role in neurite outgrowth [18], potentially as part of a chromatin re-modeling complex, interacting with essential proteins like BRG1 and heterochromatin protein 1 alpha, as well as by direct activity in the neuronal cytoplasm [18,25]. The essential role of ADNP in neuro-differentiation and potentially in neuronal injury is exemplified in that, complete ADNP deficiency is embryonic lethal at the time of neural tube closure [29].…”

Tau Pathology and Future Therapeutics

“…Our results placed ADNP at a crucial point of gene regulation, repressing potential endoderm genes and enhancing genes associated with organogenesis/neurogenesis. Immunoprecipitation experiments showed interactions with heterochromatin protein1 (HP1 ), [100] and with BRG1, BAF250a, and BAF170, all components of the SWI/SNF (mating type switching/sucrose nonfermenting) chromatin remodeling complex [101]. Together, our results place ADNP in a chromatin remodeling epigenetic role in neurodifferentiation and neuroplasticity.…”

Protection Against Tauopathy by the Drug Candidates NAP (Davunetide) and D-SAL: Biochemical, Cellular and Behavioral Aspects

“…Immunoprecipitation with specific antibodies identified ADNP interacting proteins including BRG1, BAF250a, and BAF170, all components of the SWI/SNF (mating type switching/sucrose nonfermenting) chromatin remodeling complex. ADNP short hairpin RNA down-regulation resulted in microtubule reorganization and changes in cell morphology including reduction in cell process formation and cell number [39], [40].…”