ABSTRACT
In rapidly proliferating cancer cells, glutamine is a major source of energy and building blocks. Increased glutamine uptake and enhanced glutaminolysis are key metabolic features of many cancers. Glutamine is metabolized by glutaminase (GA), which is encoded by two genes: GLS and GLS2. In contrast to isoforms arising from the GLS gene, which clearly act as oncoproteins, the role of GLS2 products in tumorigenesis is far from well understood. While in some cancer types GLS2 is overexpressed and drives cancer development, in some other types it is downregulated and behaves as a tumor suppressor gene. In this review, we describe the essential functions and regulatory mechanisms of human GLS2 and the cellular compartments in which GLS2 has been localized. Furthermore, we present the context-dependent oncogenic and tumor-suppressor properties of GLS2, and delve into the mechanisms underlying these phenomena.
ABSTRACT
Quantitative description of biochemical processes inside living cells and at single-molecule levels remains a challenge at the forefront of modern instrumentation and spectroscopy. This paper demonstrates such single-cell, single-molecule analyses performed to study the mechanism of action of olaparib - an up-to-date, FDA-approved drug for germline-BRCA mutated metastatic breast cancer. We characterized complexes formed with PARPi-FL - fluorescent analog of olaparib in vitro and in cancer cells using the advanced fluorescent-based method: Fluorescence Correlation Spectroscopy (FCS) combined with a length-scale dependent cytoplasmic/nucleoplasmic viscosity model. We determined in vitro olaparib-PARP1 equilibrium constant (6.06 × 108 mol L-1). In the cell nucleus, we distinguished three states of olaparib: freely diffusing drug (24%), olaparib-PARP1 complex (50%), and olaparib-PARP1-RNA complex (26%). We show olaparib accumulation in 3D spheroids, where intracellular concentration is twofold higher than in 2D cells. Moreover, olaparib concentration was tenfold higher (506 nmol L-1vs. 57 nmol L-1) in cervical cancer (BRCA1 high abundance) than in breast cancer cells (BRCA1 low abundance) but with a lower toxic effect. Thus we confirmed that the amount of BRCA1 protein in the cells is a better predictor of the therapeutic effect of olaparib than its penetration into cancer tissue. Our single-molecule and single-cell approach give a new perspective of drug action in living cells. FCS provides a detailed in vivo insight, valuable in drug development and targeting.