Neurodegenerative Diseases Associated Protein Alpha-synuclein And Major Depressive Disorder Associated Protein Dcnp1 Function Study

Parkinson disease (PD) is the most common neurodegenerative movement disorder (Dawson and Dawson 2003). Pathologically, it is characterized by loss of dopamine (DA) neurons and the presence of cytoplasmic inclusions (Lewy bodies, LBs) in surviving neurons in the substantia nigra pars compacta (SNc), accompanied by the presence of dystrophic neurites (Lewy neurites, LNs). Although many studies have focused on the LBs, the mechanism that underlies LB biogenesis is poorly understood.α-SYN was identified as the first "PD-gene". Three PD related mutations inα-SYN gene contribute to the rare familial forms of PD. The deposition ofα-syn has been found in PD, dementia with LBs (DLB), multiple system atrophy (MSA), and juvenile onset neuroaxonal dystrophy. These diseases are so called synucleinopathies, suggesting thatα-syn plays a common role in certain neurodegenerative diseases. In synucleinopathic brains and transgenic animal models,α-syn is selectively and extensively phosphorylated at serine 129 (S129). Phosphorylation ofα-syn at S129 is critical forα-syn neurotoxicity and furthermore, targetsα-syn for ubiquitination. Alpha-synuclein (α-syn) and ubiquitin (Ub) are major protein components deposited in Lewy bodies and Lewy neurites, which are pathologic hallmarks of idiopathic Parkinson's disease (PD). Almost 90% ofα-syn in LBs is phosphorylated at serine 129 (S129). However, the role of S129 phosphorylatedα-syn in the biogenesis of LBs remains unclear. Here, we show that compared to coexpression of wild type (WT)α-syn and Ub, coexpression of phospho-mimic mutantα-syn (S129D) and Ub in neuro2a cells results in an increase of Ub-conjugates and the formation of ubiquitinated inclusions. Furthermore, S129Dα-syn fails to increase the Ub-conjugates and form ubiquitinated inclusions in the presence of a K63R mutant Ub. In addition, as compared to WTα-syn, S129Dα-syn increased cytoplasmic and neuritic aggregates of itself in neuro2a cells treated with H₂O₂ and serum deprivation. These results suggest that the contribution of S129 phosphorylatedα-syn to the K63-linked Ub-conjugates and aggregation of itself may be involved in the biogenesis of LBs in PD and other related synucleinopathies. Major depression (MD), is a common, costly pathology that is often characterized by recurrent episodes. The morbidity of the disorder is high and has been projected to become the second leading cause of disability worldwide by 2020 (second to ischemic heart disease). The enhanced secretion of the proinflammatory cytokines, such as interleukin (IL)-1, tumor necrosis factor (TNF)-αand interferon (IFN)-γin MD, indicates the activation of immune system. The activation of immune system plays an important role in the etiology and pathophysiology of major depression. Dendritic cells (DC) are the most potent antigen presenting cells (APC) that can specifically activate the naive T cells within the immune system and are involved in the induction of both immunity and tolerance. DCs play a key role in the immune response system and have been used for immunological tolerance for transplantation and the immunotherapies for cancers. DCNP1 (dendritic cell nuclear protein 1), a protein composed of 244 amino acids, localizes to the perinucleus of mature, and to a lesser extent immature dendritic cells. A recent study has identified DCNP1 as a candidate gene for MD and the mutant gene of DCNP1 encode a protein terminated at the 117th amino acid. However, the role of immunological function of DCs and the relevance of DCNP1 to the pathogenesis of MD is still poorly understood. Here we cloned the DCNP1 gene from human HEK293 cells and constructed reconstitute plasmids that express His-tag fused DCNP1 (His-DCNP1) and EGFP fused DCNP1 (DCNP1-EGFP). The His-DCNP1 expressed in E coli was used for polyclonal antibody preparation in mice. The DCNP1-EGFP expressed in mammalian cells localizes mostly in nuclei. We identified a nuclear localization signal (NLS) that locates between the 117^th amino acid and the 119^th amino acid in DCNP1. Our results suggest that the role of MD related DCNP1 mutant in the pathogenesis of MD may be associated with its wrong location in cell.