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NF-YA overexpression protects from glutamine deprivation.

Biochimica et biophysica acta. Molecular cell research (2019-11-11)
Diletta Dolfini, Mario Minuzzo, Sarah Sertic, Roberto Mantovani
ABSTRACT

The heterotrimeric transcription factor NF-Y binds to CCAAT boxes of genes of glutamine metabolism. We set out to study the role of the regulatory NF-YA subunit in this pathway. We produced U2OS and A549 clones stably overexpressing -OE- the two splicing isoforms of NF-YA. NF-YA OE cells show normal growth and colony formation rates, but they become resistant to cell death upon glutamine deprivation. Increased mRNA and protein expression of the key biosynthetic enzyme GLUL in U2OS entails increased production of endogenous glutamine upon deprivation. The use of GLUL inhibitors dampens the NF-YA-mediated effect. NF-YA OE prevents activation of the pro-apoptotic transcription factor CHOP/DDIT3. Elevated basal levels of SERCA1/2, coding for the molecular target of Thapsigargin, correlate with resistance of NF-YA OE cells to the drug. The work represents a proof-of-principle that elevated levels of NF-YA, as found in some tumor types, helps altering cancer metabolic pathways.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
L-Methionine sulfoximine, suitable for cell culture
Supelco
Glufosinate-ammonium, PESTANAL®, analytical standard
Supelco
Glufosinate-ammonium, certified reference material, TraceCERT®, Manufactured by: Sigma-Aldrich Production GmbH, Switzerland
Sigma-Aldrich
Propidium iodide solution
Sigma-Aldrich
Thapsigargin, ≥98% (HPLC), solid film