A novel EZH2/NXPH4/CDKN2A axis is involved in regulating the proliferation and migration of non-small cell lung cancer cells.

NXPH4 is discovered to be a neuropeptide-like glycoprotein, belonging to the Neurexophilins (Nxphs) family. NXPH4 shares a similar domain structure with NXPH1, which, however, is poorly understood in terms of its function. Bioinformatics analysis and experimental verification in this study confirmed the abnormal high expression of NXPH4 in non-small cell lung cancer (NSCLC) tissues and cells. Knockdown of NXPH4 by siRNA can inhibit the proliferation and migration of cells, resulting in significant cell cycle arrest in S1 phase. Furthermore, in NSCLC cells, NXPH4 was regulated by transcriptional activation of enhancer of zeste homolog 2 (EZH2) in its upstream. While downstream, NXPH4 could interact with CDKN2A and downregulate its protein stability, thus participating in the cell cycle regulation through interacting with cyclinD-CDK4/6-pRB-E2F signaling pathway. To sum up, the present study reveals a regulatory pathway of EZH2/NXPH4/CDKN2A in NSCLC, providing possible reference for understanding the function of NXPH4 in tumors.

A multiplexed electrochemical quantitative polymerase chain reaction platform for single-base mutation analysis.

Detection of single-based mutation (SbM), which is of ultra-low abundance against wild-type alleles, are typically constrained by the level of multiplexing, sensitivity for single-base resolution and quantification accuracy. In this work, an electrochemical quantitative polymerase chain reaction (E-PCR) platform was developed for multiplexed and quantitative SbM analysis in limited and precious samples with single-nucleotide discrimination. A locked nucleic acid (LNA)-mediated multiplexed PCR system in a single, closed tube setup was firstly constructed to selectively amplify the SbM genes while suppressing the wild-type alleles. The amplicons were detected simultaneously through hybridization with the sequence-specific hairpin probes anchored on a reduced graphene oxide-gold nanoparticles functionalized electrode surface. With the inclusion of an LNA-mediated PCR step upstream of the electrochemical detection, we improved the limit of detection (LOD) by 2 orders of magnitude, down to an ultralow-level of 5 copies µL-1. The platform achieved an ultra-sensitive and specific detection with 0.05% against a background of 10, 000 copies of wild-type alleles. It is highly adaptive and has the potential to enable expanded multiplexed detection in parallel, thus providing a universal tool for multiplexed SbM identification.