Knockdown of Atg7 Induces Nuclear-LC3 Dependent Apoptosis and Augments Chemotherapy in Colorectal Cancer Cells

Abstract

Autophagy is a catabolic process that enables cells to degrade obsolete content and refuel energy depots. In colorectal cancer (CRC) autophagy has been shown to promote tumorigenesis through energy delivery in the condition of uncontrolled proliferation. With this study, we aimed at evaluating whether autophagy sustains CRC cell viability and if it impacts therapy resistance. Initially, a colorectal cancer tissue micro array, containing mucosa (n = 10), adenoma (n = 18) and adenocarcinoma (n = 49) spots, was stained for expression of essential autophagy proteins LC3b, Atg7, p62 and Beclin-1. Subsequently, central autophagy proteins were downregulated in CRC cells using siRNA technology. Viability assays, flow cytometry and immunoblotting were performed and three-dimensional cell culture was utilized to study autophagy in a tissue mimicking environment. In our study we found an upregulation of Atg7 in CRC. Furthermore, we identified Atg7 as crucial factor within the autophagy network for CRC cell viability. Its disruption induced cell death via triggering apoptosis and in combination with conventional chemotherapy it exerted synergistic effects in inducing CRC cell death. Cell death was strictly dependent on nuclear LC3b, since simultaneous knockdown of Atg7 and LC3b completely restored viability. This study unravels a novel cell death preventing function of Atg7 in interaction with LC3b, thereby unmasking a promising therapeutic target in CRC.

Keap1 inhibition sensitizes head and neck squamous cell carcinoma cells to ionizing radiation via impaired non-homologous end joining and induced autophagy

“…Further, the nuclear pool of LC3 has a specific function in priming starved cells for autophagy induction underpinning our finding that autophagy induction might be part of the enhanced cell killing upon Keap1 inhibition. In line, recent studies showed nuclear LC3 accumulation to be associated with cell death in colorectal cancer cells and human oocytes 41,42 . Regarding altered levels of Rad50, cleavage PARP1, pATM S1981, Ku80 and MRE11, we believe that the detected changes occur through a cellular attempt to compensate between NHEJ and HR repair processes.…”

Nucleolar Stress Functions Upstream to Stimulate Expression of Autophagy Regulators

“…Given the fact that nucleolar impairment by multiple triggers similarly induced expression of autophagy related factors, our findings suggest a novel nucleolar stress response that might globally be activated as compensatory mechanism to inhibit or delay apoptosis in the cells. Note that enhanced levels of ATG7, for instance, have been reported in diverse cancer types and were shown to promote cancer survival and resistance to chemotherapy, whereas a downregulation could counteract cancer development, progression and resistance [ 53 , 54 ]. Whether ATG7 upregulation indeed represents a cause for diverse diseases associated with nucleolar stress needs to be further investigated.…”

Autophagy-targeted therapy to modulate age-related diseases: Success, pitfalls, and new directions

“…Since blocking the autophagy process by pharmacological or genetic (e.g., knockdown of Beclin 1 , ATG, or ULK1/2 genes) modulation improves tumor sensitivity chemotherapy ( Xiao et al., 2021 ) or photodynamic therapy ( Martins et al., 2021 ), there has been considerable interest in developing new clinically relevant autophagy inhibitors. For instance, the disruption of up-regulated ATG7 induces tumor cell death by triggering apoptosis, which is strictly dependent on nuclear LC3B ( Scherr et al., 2020 ). On the other hand, with inhibition of VPS34 signal transduction (siRNA therapy), tumor cell proliferation was significantly suppressed after combination chemotherapy ( Zhu et al., 2015 ).…”