Understanding how PI3K/AKt/mTOR and AMPK signaling pathways affect glutaminase activity.

• Grant number: 13/23510-4
• Support Opportunities: Scholarships in Brazil - Doctorate
• Start date: May 01, 2014
• End date: February 28, 2018
• Field of knowledge: Biological Sciences - Biochemistry - Chemistry of Macromolecules
• Principal Investigator: Sandra Martha Gomes Dias
• Grantee: Carolline Fernanda Rodrigues Ascenção
• Host Institution: Centro Nacional de Pesquisa em Energia e Materiais (CNPEM). Ministério da Ciência, Tecnologia e Inovação (Brasil). Campinas , SP, Brazil
• Associated research grant: 09/10875-9 - Cellular and biochemical studies of the glutaminase enzyme and its relation with cancer, AP.JP

Abstract

Cell proliferation is crucial for embryogenesis and organism growth, being also essential for the proper function of several adult tissues. Although important for the homeostasis of the organism, its deregulation composes the driving force of tumor development. In the past ten years the relationship between the processes of signal translation stimulated by growth factors and reorganization of metabolic activity has become more evident. Growing cells need to prioritize the biosynthesis and biomass increase, processes essential for cell division. In tumor cells, the glutamine consumption is increased concurrently with the increasing in glutaminase activity on those cells. Three glutaminase isoenzymes are expressed in most tissues (liver- type glutaminase, kidney -type glutaminase and glutaminase C), but little is known about the necessity of each isoform for the tumor metabolism. Several recent papers have defined the role of glutaminolysis or glutamine metabolism in mTOR activation. So it is was valid hypothesis to speculate that mTOR can counter-regulate glutaminase. Results obtained during the master program of the candidate showed that MDA - MB 231 cells stably knocked down for PTEN did not present altered KGA and GAC protein levels, as well as there was no change on their subcellular location. Enzyme kinetics of the MDA-MB 231 mitochondrial fraction revealed that PTEN knockdown led to a decrease in the KM of the enzyme without changing Vmax. Accordingly, the treatment with rapamycin (mTOR inhibitor), led to a increase in KM back to the level detected in control cells. The glutaminase activity of total cellular lysate of MDA - MB 231, NIH 3T3, IMR90 and BJ5TA was also affected by rapamycin treatment in a dose- and time-response fashion. Moreover, glucose and glutamine cell deprivation prompted mTOR inhibition and concomitant reduction on glutaminase activity. Accordling, stable TSC2 knockdown in MDA - MB 231 and BJ5TA promoted overstimulation of mTOR and increasing on glutaminase activity. The activation of AMPK, a known energy stress sensor, led to a decreased glutaminase activity in the prostate tumor cell line DU145. The first goal of this project is to define the mTOR mechanism of action on the glutaminase. In a related subject, the comparison between PC3 and DU145 revealed that DU145 has higher GAC expression and was more responsive to metformin, a known AMPK activator. Stable AMPK knockdown in DU145, MDA- MB231 and NIH-3T3 cells led to increased protein level of KGA, while GAC was reduced. Curiously, no change was seen in PC-3. The second goal of this project is to understand the mechanism of glutaminase regulation at a protein level governed by AMPK .

Scientific publications

(4)

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

COSTA, RENNA K. E.; RODRIGUES, CAMILA T.; CAMPOS, JEAN C. H.; PARADELA, LUCIANA S.; DIAS, MARILIA M.; DA SILVA, BIANCA NOVAES; DE VALEGA NEGRAO, CYRO VON ZUBEN; GONCALVES, KALIANDRA DE ALMEIDA; ASCENCAO, CAROLLINE F. R.; ADAMOSKI, DOUGLAS; et al. High-Throughput Screening Reveals New Glutaminase Inhibitor Molecules. ACS PHARMACOLOGY & TRANSLATIONAL SCIENCE, v. 4, n. 6, p. 1849-1866, DEC 10 2021. (13/23510-4, 15/25832-4, 16/09077-4, 13/07600-3, 19/16351-3, 14/17820-3, 14/15968-3)

DOS REIS, LARISSA MENEZES; ADAMOSKI, DOUGLAS; OLIVEIRA SOUZA, RODOLPHO ORNITZ; RODRIGUES ASCENCAO, CAROLLINE FERNANDA; SOUSA DE OLIVEIRA, KRISHINA RATNA; CORREA-DA-SILVA, FELIPE; DE SA PATRONI, FABIO MALTA; DIAS, MARILIA MEIRA; CONSONNI, SILVIO ROBERTO; MENDES DE MORAES-VIEIRA, PEDRO MANOEL; et al. Dual inhibition of glutaminase and carnitine palmitoyltransferase decreases growth and migration of glutaminase inhibition-resistant triple-negative breast cancer cells. Journal of Biological Chemistry, v. 294, n. 24, p. 9342-9357, JUN 14 2019. (15/15626-8, 14/17820-3, 15/25832-4, 14/18061-9, 15/26059-7, 16/06034-2, 17/06225-5, 13/23510-4, 14/06512-6, 14/15968-3)

QUINTERO, MELISSA; ADAMOSKI, DOUGLAS; DOS REIS, LARISSA MENEZES; RODRIGUES ASCENCAO, CAROLLINE FERNANDA; SOUSA DE OLIVEIRA, KRISHINA RATNA; GONCALVES, KALIANDRA DE ALMEIDA; DIAS, MARILIA MEIRA; CARAZZOLLE, MARCELO FALSARELLA; GOMES DIAS, SANDRA MARTHA. Guanylate-binding protein-1 is a potential new therapeutic target for triple-negative breast cancer. BMC CANCER, v. 17, NOV 7 2017. (14/17820-3, 15/25832-4, 14/18061-9, 12/09452-9, 12/11577-4, 09/53853-5, 13/23510-4, 14/06512-6, 14/15968-3)

DIAS, MARILIA M.; ADAMOSKI, DOUGLAS; DOS REIS, LARISSA M.; ASCENCAO, CAROLLINE F. R.; DE OLIVEIRA, KRISHINA R. S.; PASCHOALINI MAFRA, ANA CAROLINA; DA SILVA BASTOS, ALLINY CRISTINY; QUINTERO, MELISSA; CASSAGO, CAROLINA DE G.; FERREIRA, IGOR M.; et al. GLS2 is protumorigenic in breast cancers. Oncogene, v. 39, n. 3, p. 690-702, JAN 2020. (13/05668-0, 14/18061-9, 12/14298-9, 13/23510-4, 14/17820-3, 14/06512-6, 14/15968-3, 15/25832-4, 12/09452-9, 14/20673-2, 16/06625-0, 12/11577-4, 11/10127-2)

Academic Publications

(References retrieved automatically from State of São Paulo Research Institutions)

ASCENÇÃO, Carolline Fernanda Rodrigues. Understanding the underlying mechanism behind of mTOR control of glutaminase C activity and identification and characterization of glutaminase C post-translational modifications. 2018. Doctoral Thesis - Universidade Estadual de Campinas (UNICAMP). Instituto de Biologia Campinas, SP.