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The Warburg Effect Mediator Pyruvate Kinase M2 Expression and Regulation in the Retina

View Article: PubMed Central - PubMed

ABSTRACT

The tumor form of pyruvate kinase M2 (PKM2) undergoes tyrosine phosphorylation and gives rise to the Warburg effect. The Warburg effect defines a pro-oncogenic metabolism switch such that cancer cells take up more glucose than normal tissue and favor incomplete oxidation of glucose, even in the presence of oxygen. Retinal photoreceptors are highly metabolic and their energy consumption is equivalent to that of a multiplying tumor cell. In the present study, we found that PKM2 is the predominant isoform in both rod- and cone-dominant retina, and that it undergoes a light-dependent tyrosine phosphorylation. We also discovered that PKM2 phosphorylation is signaled through photobleaching of rhodopsin. Our findings suggest that phosphoinositide 3-kinase activation promotes PKM2 phosphorylation. Light and tyrosine phosphorylation appear to regulate PKM2 to provide a metabolic advantage to photoreceptor cells, thereby promoting cell survival.

No MeSH data available.


Biochemical characterization of PKM2 and PKM1 isoforms on isolated photoreceptor cells from dark- and light-adapted Balb/c mice.Retinal homogenates from dark- and light-adapted Balb/c mice were subjected to OptiPrep™ (8–40%) density gradient centrifugation (A). Fractions of inner segments and intact photoreceptors (B) were collected from the top to the bottom of the gradients. A ten-microliter sample (one-microliter for rhodopsin) was subjected to immunoblot analysis (C) with opsin, α-3 Na/K ATPase, pPKM2, PKM2, and PKM1 antibodies. Full-length blots are presented in the Supplementary Information.
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f5: Biochemical characterization of PKM2 and PKM1 isoforms on isolated photoreceptor cells from dark- and light-adapted Balb/c mice.Retinal homogenates from dark- and light-adapted Balb/c mice were subjected to OptiPrep™ (8–40%) density gradient centrifugation (A). Fractions of inner segments and intact photoreceptors (B) were collected from the top to the bottom of the gradients. A ten-microliter sample (one-microliter for rhodopsin) was subjected to immunoblot analysis (C) with opsin, α-3 Na/K ATPase, pPKM2, PKM2, and PKM1 antibodies. Full-length blots are presented in the Supplementary Information.

Mentions: Biochemical analyses were performed on isolated cellular fragments from mouse retinas on an OptiPrepTM density gradient (8–40%) that yields rod outer segments with large portions of the rod inner segments attached (Fig. 5A,B). Retinas from Balb/c mice and Nrl−/− mice were subjected to density gradient centrifugation, and fractions were collected. We used the most predominant rod outer segment resident marker, rhodopsin, for rods. For cones, M-opsin was used. It has been shown that cells break open and release some portion of the cytoplasm, including soluble proteins, during sample preparation. The breakpoint occurs in the inner segment, while the soluble components are essentially retained within the outer segments19.


The Warburg Effect Mediator Pyruvate Kinase M2 Expression and Regulation in the Retina
Biochemical characterization of PKM2 and PKM1 isoforms on isolated photoreceptor cells from dark- and light-adapted Balb/c mice.Retinal homogenates from dark- and light-adapted Balb/c mice were subjected to OptiPrep™ (8–40%) density gradient centrifugation (A). Fractions of inner segments and intact photoreceptors (B) were collected from the top to the bottom of the gradients. A ten-microliter sample (one-microliter for rhodopsin) was subjected to immunoblot analysis (C) with opsin, α-3 Na/K ATPase, pPKM2, PKM2, and PKM1 antibodies. Full-length blots are presented in the Supplementary Information.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5121888&req=5

f5: Biochemical characterization of PKM2 and PKM1 isoforms on isolated photoreceptor cells from dark- and light-adapted Balb/c mice.Retinal homogenates from dark- and light-adapted Balb/c mice were subjected to OptiPrep™ (8–40%) density gradient centrifugation (A). Fractions of inner segments and intact photoreceptors (B) were collected from the top to the bottom of the gradients. A ten-microliter sample (one-microliter for rhodopsin) was subjected to immunoblot analysis (C) with opsin, α-3 Na/K ATPase, pPKM2, PKM2, and PKM1 antibodies. Full-length blots are presented in the Supplementary Information.
Mentions: Biochemical analyses were performed on isolated cellular fragments from mouse retinas on an OptiPrepTM density gradient (8–40%) that yields rod outer segments with large portions of the rod inner segments attached (Fig. 5A,B). Retinas from Balb/c mice and Nrl−/− mice were subjected to density gradient centrifugation, and fractions were collected. We used the most predominant rod outer segment resident marker, rhodopsin, for rods. For cones, M-opsin was used. It has been shown that cells break open and release some portion of the cytoplasm, including soluble proteins, during sample preparation. The breakpoint occurs in the inner segment, while the soluble components are essentially retained within the outer segments19.

View Article: PubMed Central - PubMed

ABSTRACT

The tumor form of pyruvate kinase M2 (PKM2) undergoes tyrosine phosphorylation and gives rise to the Warburg effect. The Warburg effect defines a pro-oncogenic metabolism switch such that cancer cells take up more glucose than normal tissue and favor incomplete oxidation of glucose, even in the presence of oxygen. Retinal photoreceptors are highly metabolic and their energy consumption is equivalent to that of a multiplying tumor cell. In the present study, we found that PKM2 is the predominant isoform in both rod- and cone-dominant retina, and that it undergoes a light-dependent tyrosine phosphorylation. We also discovered that PKM2 phosphorylation is signaled through photobleaching of rhodopsin. Our findings suggest that phosphoinositide 3-kinase activation promotes PKM2 phosphorylation. Light and tyrosine phosphorylation appear to regulate PKM2 to provide a metabolic advantage to photoreceptor cells, thereby promoting cell survival.

No MeSH data available.