Abstract LB014: Translational activation of ATF4 through mitochondrial anaplerotic metabolic pathways is required for DLBCL growth and survival
Authors: Li Meng , Teater Matthew R. , Hong Jun Young , Duy Cihangir , Shen Hao , Wang Ling , Chen Zhengming , Cerchietti Leandro , Lin Hening , Melnick Ari 
Published: July 2021

Diffuse large B-cell lymphomas (DLBCLs) tolerate various forms of cellular stress associated with their rapid proliferation and depletion of nutrients in their microenvironment. Our previous finding discovered that DLBCLs are broadly dependent mitochondrial SIRT3 as a critical source of non-oncogene addiction in DLBCL. SIRT3 promotes tumorigenesis in DLBCL, where it is required for glutamine fueled anaplerosis, and its loss of function leads to tumor suppressive autophagy. However, it is not known why SIRT3 deficient DLBCL cells are so vulnerable to such metabolic changes and autophagy, which points to potentially novel and critical nutrient regulatory circuits occurring in this disease.

We set out to identify downstream signals induced by SIRT3 deficiency in DLBCL cells in order to gain insight into how SIRT3 could interface with nutrient flux stress response pathways to support tumorigenesis of DLBCLs. First, we carried out RNA sequencing study in three DLBCL cell lines representing different subtypes of DLBCLs. The data showed that all SIRT3 deficient DLBCL cells experienced transcriptional inhibitions of ATF4 target genes. We further observed that ATF4 protein was decreased due to the translational inhibition in SIRT3 deficient DLBCL cells, which indicates that ATF4 may play a role downstream of SIRT3 contributing to DLBCL cell proliferation. Secondly, we proved that ATF4 is indeed required to DLBCL cells' proliferation. ATF4 target genes are more expressed in DLBCL tumor cells than normal germinal center B cells. Loss of ATF4 induced proliferation arrest as SIRT3 depletion and over-expression of ATF4 can partially rescue the proliferation and viability inhibited by SIRT3 deficiency. Consistently, we observed that loss of SIRT3 led to ATF4 reduction in the vavP-Bcl2 mouse lymphoma model. Mechanistically, we identified that ATF4 functions downstream of SIRT3 through the metabolic cascade of GDH-TCA cycle-autophagy. GDH and DMKG can rescue the ATF4 protein decreased by SIRT3 depletion. Blockage of autophagy with chloroquine or knocking down ATG5 can also neutralize SIRT3's inhibition on ATF4. Furthermore, we discovered that

ATF4's translation is highly sensitive to nutrient/amino acid levels in DLBCL cells as a stress signal for cell survival. However, ATF4 translation failed to respond to glutamine starvation in SIRT3 depleted DLBCL cells. SIRT3 deficiency underwent drastic changes of amino acid levels because of the TCA cycle inhibition and autophagy activation, which interfered the ATF4's translation and can lead to deleterious metabolic stresses. Collectively, we identify a novel SIRT3-ATF4 axis required to maintain survival of DLBCL cells by enabling them to optimize amino acid utilization. Lack of this coordination is lethal to DLBCL cells, exposing a critical and exploitable metabolic vulnerability.

Citation Format: Meng Li, Matthew R. Teater, Jun Young Hong, Cihangir Duy, Hao Shen, Ling Wang, Zhengming Chen, Leandro Cerchietti, Hening Lin, Ari Melnick. Translational activation of ATF4 through mitochondrial anaplerotic metabolic pathways is required for DLBCL growth and survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB014.