Leukemia Cells Resistant to Glutamine Deprivation Express Glutamine Synthetase (GS) Protein

Turk J Haematol. 2021 Apr 22. doi: 10.4274/tjh.galenos.2021.2021.0054. Online ahead of print.

Abstract

Aim: Low glutamine level has been shown in tumor environment for several cancer subtypes. Therefore, it has been suggested that cancer cells rewire their metabolism to adopt low nutrient level for survival and proliferation. Although glutamine is a non-essential amino acid and can be synthesized de novo, many cancer cells including malignant hematopoietic cells have been indicated to be addicted to glutamine. This study aimed to investigate proliferation of leukemia cell lines in glutamine deprived conditions.

Materials and methods: Cell proliferation of K562, NB-4 and HL-60 cells were determined calculating cell numbers in normal vs low glutamine media. Changes in mRNA expressions were investigated using qRT-PCR. GS encoding GLUL gene was knocked out (KO) in HL-60 cells using CRISPR/Cas9 method, protein expression was evaluated with immunoblotting.

Results: Proliferation of all cell lines were decreased in glutamine deprived medium. GS protein expression was increased in glutamine limited medium although mRNA level did not change. Increased protein expression was confirmed with inhibition of new protein synthesis by treating cells with cycloheximide. To further investigate the role of GS protein, GS encoding GLUL gene was knocked out (KO) in HL-60 cells using CRISPR/Cas9 method. GS KO cells less proliferated compared to control cells in glutamine limited medium.

Conclusion: Our results indicates that upregulated GS protein expression is responsible for glutamine addiction of leukemia cell lines. Exploiting the genetic and metabolic mechanisms responsible for GS protein expression could identify new anti-cancer drug targets.

Keywords: Crispr/cas9; glutamine limitation; glutamine synthetase; leukemia.