Association of Gpx1 fluctuation in cell cycle progression

Abstract

This research demonstrates fluctuation of glutathione peroxidase1 (Gpx1) throughout cell cycle progression with significant decreased expression at mitosis of HeLa cell. This was achieved with western blot (WB) analysis of target proteins from each phase of synchronized cells. The synchronizations were performed with double thymidine (T/T) for G1/S arrest and thymidine followed by nocodazol (T/N) for G2/M arrest. The G1/S arrested cells were released in fresh medium for 3, 6, 9, 10 and 15h to obtain cell at each phase such as Gap1 (G1), synthesis (S), gap2 (G2), mitosis (M) and gap1 (G1) phase, respectively for investigating Gpx1 expression throughout a complete cycle. The synchronizations were confirmed using fluorescence activated cell sorting (FACS) and WB analysis of phase specific markers. The fluctuations of Gpx1 expression were verified with universal protein actin and peroxiredoxin 1 (Prx1) which are stable throughout the cell cycle. Intriguingly, immunoblots showed the level of Gpx1 decreases at mitosis phase and increased during mitotic exit to G1 phase in HeLa cells, while Prx1 protein level remained constant. The fractionation experiments reveal that only the cytosolic Gpx1 was decreased while their levels at mitochondria remain constant. The heighest levels of mitochondrial ROS were measured in mitosis phase with FACS analysis using Mito sox indicating that antioxidant activity of Gpx1 for detoxifying excessive induced endogenous reactive oxygen species (ROS) in the mitosis phase could be the reason for such decreasing level.For unfolding the molecular mechanism of such decreased expression, the Gpx1 was investigated at transcriptional, translational and proteosomal level. The results revealed that translational mechanism is involve in the decreased expression rather than transcriptional or proteosomal degradation at mitosis phase. This finding supports that Gpx1 involved in the cell cycle progression through regulation of endogenous ROS. Based on this observation further research could uncover their possible association with the infinitive division of a cancer cell.

Using Information from Public Databases to Critically Evaluate Studies Linking the Antioxidant Enzyme Selenium-Dependent Glutathione Peroxidase 2 (GPX2) to Cancer

“…In agreement with DepMap, we identified two lines (NCIH841 and NCIH1229) as having neither GPX1 nor GPX2 expression (Table in Figure 13) [83]. The DepMap values for the cell line, HeLa, also seem to agree with reports in the literature, having almost no GPX2 expression and mid-level GPX1 expression [60,88,89]. Another issue found in a recent paper was the representation of a cell line called LNCaP as having high GPX2 levels via Western blotting [90], while three other papers counter this by showing LNCaP to have very low levels of GPX2 (Western and Northerns); consulting DepMap and THPA also quickly settles this issue [4,66,79].…”

The Encystment-Related MicroRNAs and Its Regulation Molecular Mechanism in Pseudourostyla cristata Revealed by High Throughput Small RNA Sequencing

“…It mainly scavenges organic peroxides, especially the large amount of lipid peroxides produced when free radicals cause lipid peroxidation. Moreover, GPX is also involved in a series of important biological processes such as the cell cycle, cell signal transduction, chromatin remodeling, DNA, and histone modification [17][18][19][20]. Therefore, the upregulation of GPX enhances the antioxidant and anti-stress capabilities of the cysts.…”