SUMMARY The regulation and coordination of mitochondrial metabolism with hematopoietic stem cell (HSC) self-renewal and differentiation is not fully understood. Here we report that depletion of PTPMT1, a PTEN-like mitochondrial phosphatase, in inducible or hematopoietic-cell-specific knockout mice resulted in hematopoietic failure due to changes in the cell cycle and a block in the differentiation of HSCs. Surprisingly, the HSC pool was increased by ~40-fold in PTPMT1 knockout mice. Reintroduction of wild-type PTPMT1, but not catalytically deficient PTPMT1 or truncated PTPMT1 lacking mitochondrial localization, restored differentiation capabilities of PTPMT1 knockout HSCs. Further analyses demonstrated that PTPMT1 deficiency altered mitochondrial metabolism and that phosphatidylinositol phosphate substrates of PTPMT1 directly enhanced fatty-acid-induced activation of mitochondrial uncoupling protein 2. Intriguingly, depletion of PTPMT1 from myeloid, T lymphoid, or B lymphoid progenitors did not cause any defects in lineage-specific knockout mice. This study establishes a crucial role of PTPMT1 in the metabolic regulation of HSC function.

The intracellular Ca 2+ ([Ca 2+ ] i ) level of skeletal muscles must be rapidly regulated during the excitation-contraction-relaxation process 1 . However, the signaling components involved in such rapid Ca 2+ movement are not fully understood. Here, we report that mice deficient in the novel phosphatidylinositol phosphate (PIP) phosphatase MIP displayed muscle weakness and fatigue. Muscles isolated from MIP −/− mice produced less contractile force, markedly prolonged relaxation, and exhibited exacerbated fatigue. Further analyses revealed that MIP deficiency resulted in spontaneous Ca 2+ leak from the internal store -the sarcoplasmic reticulum (SR). This was attributed to the decreased metabolism/dephosphorylation and the subsequent accumulation of MIP substrates, especially PI(3,5)P 2 and PI(3,4)P 2 . Furthermore, we found that PI(3,5)P 2 and PI(3,4)P 2 bound to and directly activated the Ca 2+ release channel/ryanodine receptor (RyR1) of the SR. These studies provide the first evidence that finely controlled PIP levels in muscle cells are essential for maintaining Ca 2+ homeostasis and muscle performance.During our systematic genome-wide survey for tyrosine/dual specificity phosphatases (unpublished work), we discovered a novel phosphatase by hidden Markov database mining using the conserved catalytic motif ([V/I][V/I]HCXXGXXR[T/S]) as the bait sequence. Both human (BC035690) and mouse (BC018294) homologies were identified. They share 90% identity in amino acid sequences ( Supplementary Information, Fig. S1). Northern blotting analyses illustrated that this phosphatase was predominantly expressed in skeletal muscle and heart (Fig. 1a). Immunostaining indicates that it is primarily localized in the cytoplasm (data not shown). To verify its phosphatase property, we generated a GST fusion protein and tested its catalytic activity using pNPP (p-Nitrophenyl Phosphate), a widely used non-specific 7Correspondence should be addressed to: C.K.Q. (e-mail: E-mail: cxq6@case.edu). 6 These authors contributed equally to this work. AUTHOR CONTRIBUTIONSJ.S., W.M. Y., M.B., J.A.S., and C.S. conducted the research and summarized the data. C.K.Q., M.B., H.H.V., T.M.N., and C.G. designed the experiments and wrote the manuscript. COMPETING FINANCIAL INTERESTSThe authors declare no competing financial interests. (Fig. 1b). Instead, it dephosphorylated a variety of PIPs, especially PI(3,5) P 2 (Fig. 1c), similar to PTEN and myotubularin and myopathy related (MTMR) phosphatases that also favor PIPs as substrates despite containing tyrosine phosphatase domains 2 . As this new phosphatase is mainly expressed in skeletal muscle and heart, we named it MIP (musclespecific inositol phosphatase). While our gene knockout work on MIP was ongoing, the Mustalin group also identified this phosphatase (FLJ20133) in their comprehensive collection of tyrosine phosphatases from the human genome and listed it as the 14 th member of the MTMR family (MTMR14) based on the homology of its catalytic motif to myotubularin 3 . More recently, ina...

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Mitochondria are involved in a number of cellular processes and are essential for both life and death. As the site of oxidative phosphorylation, these double-membrane organelles provide a highly efficient route for eukaryotic cells to generate ATP from energy-rich molecules. During the mitochondrial energy production process, reactive oxygen species (ROS), such as superoxide (O 2 Ϫ ) and hydrogen peroxide (H 2 O 2 ), are produced as by-products. In fact, mitochondria are the primary source of a majority of cellular ROS (2). Mitochondria also participate in intermediary metabolism. Under normal oxygen tensions, cells catabolize glucose to pyruvate. Pyruvate is then imported into the mitochondria for further catabolism through the Krebs cycle, which transfers electrons to the respiratory chain for ATP synthesis. In low oxygen tension, or hypoxic conditions in which there is a paucity of oxygen as an electron acceptor, cells are surmised to undergo anaerobic glycolysis as a default mode. Pyruvate is then used for low-efficiency energy production in the cytosol by glycolysis. In addition to metabolism and energy production, mitochondria play important roles in the regulation of apoptosis and intracellular Ca 2ϩ homeostasis. Dysfunction in mitochondria results in severe cellular consequences and is linked to a wide range of human diseases (2, 36, 43). The role of mitochondrial activities in early embryonic development and embryonic stem (ES) cell function is not well defined (20,42). The environment of the uterus before placentation is anaerobic (11). To produce ATP in this environment, early embryonic cells, such as ES cells, rely heavily on glycolysis for ATP production (4) and, thus, do not require a large number of mitochondria. ES cells only have a few mitochondria with poorly developed cristae (21). Effective control of mitochondrial mass and function is critical for the prevention of damage by oxidative stress (ROS) in ES cells. However, when these cells are allowed to differentiate, the resulting cells show numerous large mitochondria with distinct cristae. Thus, mitochondria must undergo robust replication/biogenesis during this short period of time. Earlier studies have shown that the mitochondrial genome undergoes significant replication during implantation of blastocysts (41), and once gastrulation occurs, cells replicate their mitochondrial DNA (mtDNA) to match the energy demand of differentiating cells (39). It has also been demonstrated that mitochondrial metabolic rates correlate inversely with the differentiation capacity of ES cells (37). However, exactly how mitochondria coordinate stem cell behavior during embryogenesis is still not well understood.Mitochondria are highly dynamic organelles that undergo continuous fusion and fission. These mitochondrial processes play important roles in mitochondrial biogenesis/replication.

We have recently reported that a novel muscle-specific inositide phosphatase (MIP/MTMR14) plays a critical role in [Ca2+]i homeostasis through dephosphorylation of sn-1-stearoyl-2-arachidonoyl phosphatidylinositol (3,5) bisphosphate (PI(3,5)P2). Loss of function mutations in MIP have been identified in human centronuclear myopathy. We developed a MIP knockout (MIPKO) animal model and found that MIPKO mice were more susceptible to exercise-induced muscle damage, a trademark of muscle functional changes in older subjects. We used wild-type (Wt) mice and MIPKO mice to elucidate the roles of MIP in muscle function during aging. We found MIP mRNA expression, MIP protein levels, and MIP phosphatase activity significantly decreased in old Wt mice. The mature MIPKO mice displayed phenotypes that closely resembled those seen in old Wt mice: i) decreased walking speed, ii) decreased treadmill activity, iii) decreased contractile force, and iv) decreased power generation, classical features of sarcopenia in rodents and humans. Defective Ca2+ homeostasis is also present in mature MIPKO and old Wt mice, suggesting a putative role of MIP in the decline of muscle function during aging. Our studies offer a new avenue for the investigation of MIP roles in skeletal muscle function and as a potential therapeutic target to treat aging sarcopenia.