Phosphatases of regenerating liver (PRL-1, PRL-2, and PRL-3, also known as PTP4A1, PTP4A2, and PTP4A3) control magnesium homeostasis through an association with the CNNM magnesium transport regulators. Although high PRL levels have been linked to cancer progression, regulation of their expression is poorly understood. Here we show that modulating intracellular magnesium levels correlates with a rapid change of PRL expression by a mechanism involving its 5′UTR mRNA region. Mutations or CRISPR-Cas9 targeting of the conserved upstream ORF present in the mRNA leader derepress PRL protein synthesis and attenuate the translational response to magnesium levels. Mechanistically, magnesium depletion reduces intracellular ATP but up-regulates PRL protein expression via activation of the AMPK/mTORC2 pathway, which controls cellular energy status. Hence, altered PRL-2 expression leads to metabolic reprogramming of the cells. These findings uncover a magnesium-sensitive mechanism controlling PRL expression, which plays a role in cellular bioenergetics.
The Phosphatase of Regenerative Liver (PRL) enzymes (PRL‐1,‐2,‐3; gene name PTP4A1, PTP4A2, PTP4A3) are members of the protein tyrosine phosphatase family. These enzymes have been identified as key contributors to tumor progression and metastasis in several human cancers, yet the molecular basis of their pro‐oncogenic property is unclear. Using mass spectrometry studies, we identified the CNNM magnesium transporters as key binding partners of PRLs in an evolutionarily conserved complex that regulates the intracellular magnesium concentration. Structural analysis of this complex indicates that the second cystathionine β‐synthase (CBS) domain of CNNM underlines a binding loop unique to the CNNM family that only exists in organisms having PRL orthologs. Mutation of an evolutionarily conserved aspartic acid located in this CBS domain of CNNM3 was able to completely abolish the interaction with PRL‐2 resulting in reduced tumor growth. Also, either the abolishment of complex formation or PRL‐2 knockdown resulted in reduced magnesium transport. Since magnesium is a critical metabolic effector, we confirmed that mitochondrial respiration is altered in cells isolated from PRL2‐null animals and is associated with a lower ATP turnover. This is also consistent with the observed decreased body weight of PRL‐2−/− mice, suggesting that their cellular metabolism is inherently less efficient. As cellular magnesium transport is regulated by intrinsic mechanisms controlling various magnesium transporters and magnesium uptake in cells, we also aimed to examine the involvement of PRLs in the adaptive response related to changes in magnesium availability. Using various cell lines cultured under low magnesium conditions, we observed a decrease in intracellular magnesium levels correlating with a marked increase of PRL‐1 and ‐2 (PRL‐1/2) expression. Although the expression levels of the magnesium transporter CNNM3 remained unchanged after magnesium depletion, we found that the interaction between endogenous PRL‐1/2 and CNNM3 was also markedly increased, indicating a pivotal regulatory role for PRL‐1/2 in the complex to compensate for decreased magnesium availability. On the contrary, upon increasing intracellular magnesium levels by overexpressing the TRPM7 magnesium transporter, we observed a reduction of PRL‐1/2 expression. Consistent with this, using an in vivo reporter mouse model, PRL‐2 tissue expression was upregulated when dietary magnesium was limited. Overall, our findings suggest that PRL phosphatases regulate cellular magnesium levels, essential for cell survival/proliferation, which may explain much of the clinical relationship between PRLs and cancer.Support or Funding InformationThis research is supported by an operating grant from Canadian Institutes of Health Research (CIHR); CIHR MOP 142497.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
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