MM cells exhibit high expression of GLS1 but not GS. (A) GLS1, GAC, KGA, GLUL, and ASNS expression was analyzed by real-time PCR in HMCLs and 697 cells. Gene expression was normalized to the expression of RPL-15. GAC/KGA mRNA was also reported. Means plus SD of 3 experiments with 2 determinations each are shown. (B) GLS1, GAC, and KGA expression in primary CD138⁺ cells, purified from 3 MGUS, 5 SMM, 11 ND-MM, and 10 R-MM patients was evaluated with real-time PCR. Lines represent median values. (C) Western blot of GLS1, GS, and ASNS expression by HMCLs and 697 cells. β-tubulin was used for loading control. (D) GS expression in HMCLs and 697 cells incubated for 19 hours in the presence of 4 mM Gln (+) or in the absence of the amino acid (−). (E) GLS1 and GS expression was evaluated by Western blot in CD138⁺ cells purified from 4 SMM, 7 ND-MM, and 4 R-MM patients; 697 lysate was used as positive control. GAPDH was used for loading control.