The mouse nicotinamide mononucleotide adenylyltransferase chaperones diverse pathological amyloid client proteins.

Molecular chaperones safeguard cellular protein homeostasis and obviate proteotoxicity. In the process of aging, as chaperone networks decline, aberrant protein amyloid aggregation accumulates in a mechanism that underpins neurodegeneration, leading to pathologies such as Alzheimer's disease and Parkinson's disease. Thus, it is important to identify and characterize chaperones for preventing such protein aggregation. In this work, we identified that the NAD+ synthase-nicotinamide mononucleotide adenylyltransferase (NMNAT) 3 from mouse (mN3) exhibits potent chaperone activity to antagonize aggregation of a wide spectrum of pathological amyloid client proteins including α-synuclein, Tau (K19), amyloid ß, and islet amyloid polypeptide. By combining NMR spectroscopy, cross-linking mass spectrometry, and computational modeling, we further reveal that mN3 uses different region of its amphiphilic surface near the active site to directly bind different amyloid client proteins. Our work demonstrates a client recognition mechanism of NMNAT via which it chaperones different amyloid client proteins against pathological aggregation and implies a potential protective role for NMNAT in different amyloid-associated diseases.

Structural basis of the interplay between α-synuclein and Tau in regulating pathological amyloid aggregation.

Amyloid aggregation of pathological proteins is closely associated with a variety of neurodegenerative diseases, and α-synuclein (α-syn) deposition and Tau tangles are considered hallmarks of Parkinson's disease and Alzheimer's disease, respectively. Intriguingly, α-syn and Tau have been found to co-deposit in the brains of individuals with dementia and parkinsonism, suggesting a potential role of cross-talk between these two proteins in neurodegenerative pathologies. Here we show that monomeric α-syn and the two variants of Tau, Tau23 and K19, synergistically promote amyloid fibrillation, leading to their co-aggregation in vitro NMR spectroscopy experiments revealed that α-syn uses its highly negatively charged C terminus to directly interact with Tau23 and K19. Deletion of the C terminus effectively abolished its binding to Tau23 and K19 as well as its synergistic effect on promoting their fibrillation. Moreover, an S129D substitution of α-syn, mimicking C-terminal phosphorylation of Ser129 in α-syn, which is commonly observed in the brains of Parkinson's disease patients with elevated α-syn phosphorylation levels, significantly enhanced the activity of α-syn in facilitating Tau23 and K19 aggregation. These results reveal the molecular basis underlying the direct interaction between α-syn and Tau. We proposed that this interplay might contribute to pathological aggregation of α-syn and Tau in neurodegenerative diseases.

Underlying mechanisms for LTF inactivation and its functional analysis in nasopharyngeal carcinoma cell lines.

The lactoferrin (LTF) gene, located at 3p21.3, behaves like a tumor suppressor gene in diverse tumors. To elucidate the exact role of LTF in NPC, we first detected its expression level in seven NPC cell lines by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). The results showed the mRNA level of LTF was nearly undetectable in all the seven NPC cell lines, while it could be detected in chronic nasopharyngitis tissues. Subsequently, we used methylation-specific PCR (MSP), microsatellite assay, PCR-single-strand conformation polymorphism (PCR-SSCP) and sequencing methods to examine the promoter methylation, loss of heterozygosity (LOH) and gene mutation of LTF in NPC cell lines respectively. Consequently, we found that 100% (7 of 7) of NPC cell lines were methylated in LTF promoter, only one cell line (14%, 1 of 7) had LOH and gene mutation of LTF, respectively, while LTF exhibited re-expression in all cell lines after 5-aza-dC treatment, indicating promoter methylation should be the key mechanism causing LTF downregulation in NPC cell lines. Furthermore, patched methylation assay confirmed that promoter methylation could down-regulate LTF gene expression in NPC cells. Finally, we investigated the function of LTF in NPC cell lines by gene transfection. Restoration of LTF expression in NPC cells resulted in blockage of cell cycle progression, significant inhibition of cell growth and a reduced colony-formation capacity in vitro and obviously weaker tumor formation potential in vivo. In conclusion, our data indicate LTF may participate in NPC carcinogenesis as a negative effector, that is, a tumor suppressor gene.