PGC-1a coordinates with Bcl-2 to control cell cycle in U251 cells through reducing ROS

Kun Yao; Xufeng Fu; Du Xing; Yan Li; Bing Han; Chen Zexi; Shanshan Yang; Ran Wei; Jiaqi Zhou; Qinghua Cui

Introduction 39 B-cell lymphoma 2 (Bcl-2) has both pro-apoptotic and anti-apoptotic potentials. It 40 is well known that Bcl-2 protein is anchored to the mitochondrial outer membrane, 41 and antagonizes with the pro-apoptotic protein BAX by forming Bcl-2/BAX 42 heterodimers that control mitochondrial membrane permeability and promote formation are not fully understood. 52 We have recently observed that quiescent Bcl-2 overexpressing cells maintain The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/111567 doi: bioRxiv preprint 3 modulate mitochondrial biogenesis and bioenergetics is peroxisome 58 proliferator-activated receptor-γ co-activator 1α (PGC-1α), which is predominantly 59 expressed in tissues with high energy demands, such as the heart, brain, and muscle The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/111567 doi: bioRxiv preprint 4 3T3PB cells. Our cell cycle profiles showed that both 3T3PB and 3T3Bcl-2 were 87 arrested successfully in G 0 /G 1 phase, the proportion of cells in S phase dropped from 88 ~20% (normal growing, NG) to less than 3% (serum starved, SS) for both cells (Fig. 89 1A and 1B). We also observed a significant elevation in p27 levels in SS3T3Bcl-2 90 cells, confirming that Bcl-2 function as tumor-repressive gene through upregulating 91 p27 ( Fig. 1C and 1D ). At this stage, PGC-1α expression was clearly increased in 92 SS3T3Bcl-2 comparing with NG3T3Bcl-2 cells, while there was no significant 93 difference between control cells SS3T3PB and NG3T3PB ( Fig. 1C and 1E ). This 94 result suggests that PGC-1α expression associates with Bcl-2 after serum depletion. Fig. 1 and 2) , suggesting a relationship between PGC-1α and Bcl-2. We hypothesized 187 that Bcl-2 may recruit and rely on PGC-1α to reduce ROS, which mediated cell cycle 188 as a critical signal. To test this possibility, Bcl-2 knockdown by siRNA was performed, 189 and as we expected, PGC-1α expression was correspondingly decreased (Fig.3) . 190 Together with the facts that PGC-1α regulates mitochondrial function to eliminate 191 ROS, and arrested Bcl-2 cells contain less ROS, these results suggest that Bcl-2 might 192 recruit PGC-1α to reduce ROS levels and block cell cycle progression. 193 To further clarify the relationship between Bcl-2 and PGC-1α, we investigated the 194 influence of PGC-1α knockdown on Bcl-2 expression. Endogenous expression of 195 PGC-1α is low in normal growing 3T3Bcl-2 and control cells (Fig. 1C, Fig. 2C The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/111567 doi: bioRxiv preprint . CC-BY 4.0 International license is made available under a