Abstract
Preferential expression of the low-activity (dimeric) M2 isoform of pyruvate kinase (PK) over its constitutively active splice variant M1 isoform is considered critical for aerobic glycolysis in cancer cells. However, our results reported here indicate co-expression of PKM1 and PKM2 and their possible physical interaction in cancer cells. We show that knockdown of either PKM1 or PKM2 differentially affects net PK activity, viability, and cellular ATP levels of the lung carcinoma cell lines H1299 and A549. The stable knockdown of PK isoforms in A549 cells significantly reduced the cellular ATP level, whereas in H1299 cells the level of ATP was unaltered. Interestingly, the PKM1/2 knockdown in H1299 cells activated AMP-activated protein kinase (AMPK) signaling and stimulated mitochondrial biogenesis and autophagy to maintain energy homeostasis. In contrast, knocking down either of the PKM isoforms in A549 cells lacking LKB1, a serine/threonine protein kinase upstream of AMPK, failed to activate AMPK and sustain energy homeostasis and resulted in apoptosis. Moreover, in a similar genetic background of silenced PKM1 or PKM2, the knocking down of AMPKα1/2 catalytic subunit in H1299 cells induced apoptosis. Our findings help explain why previous targeting of PKM2 in cancer cells to control tumor growth has not met with the expected success. We suggest that this lack of success is because of AMPK-mediated energy metabolism rewiring, protecting cancer cell viability. On the basis of our observations, we propose an alternative therapeutic strategy of silencing either of the PKM isoforms along with AMPK in tumors.
Highlights
Preferential expression of the low-activity M2 isoform of pyruvate kinase (PK) over its constitutively active splice variant M1 isoform is considered critical for aerobic glycolysis in cancer cells
The phenomenon of co-expression was noticed at RNA level in cultured human cancer cells, using semi-quantitative RTPCR followed by exon-specific restriction digestion of PKM2, a modified technique adopted from David et al [49], to examine the proportion of the expression of the PKM1 and PKM2 isoforms (Fig. 1, A and B)
From the results of two independent liquid chromatography-mass spectrometry (LC-MS) studies, we found nearly 30 interacting proteins of PKM1 (Fig. 2B and supplemental Table S1), which involved the proteins from cytoplasm, mitochondria, and nucleus, as an integral part of diverse cellular machinery of glycolytic pathway, mitochondrial electron transport chain, protein translation, protein folding, DNA replication, and cytoskeletal networks (Fig. 2B)
Summary
Preferential expression of the low-activity (dimeric) M2 isoform of pyruvate kinase (PK) over its constitutively active splice variant M1 isoform is considered critical for aerobic glycolysis in cancer cells. Our findings help explain why previous targeting of PKM2 in cancer cells to control tumor growth has not met with the expected success We suggest that this lack of success is because of AMPK-mediated energy metabolism rewiring, protecting cancer cell viability. In addition to aerobic glycolysis, PKM2 provides multiple benefits to cancer cells by performing the nonmetabolic role of co-transcriptional activation (14 –16), protein kinase function [17, 18], and chromosomal segregation [19]. Supporting such a deep-rooted association with cancer, the M2 isoform of pyruvate kinase has emerged as a potential candidate to target different types of tumors.
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