Mapping the cellular origin and early evolution of leukemia in Down syndrome.

Children with Down syndrome have a 150-fold increased risk of developing myeloid leukemia, but the mechanism of predisposition is unclear. Because Down syndrome leukemogenesis initiates during fetal development, we characterized the cellular and developmental context of preleukemic initiation and leukemic progression using gene editing in human disomic and trisomic fetal hematopoietic cells and xenotransplantation. GATA binding protein 1 (GATA1) mutations caused transient preleukemia when introduced into trisomy 21 long-term hematopoietic stem cells, where a subset of chromosome 21 microRNAs affected predisposition to preleukemia. By contrast, progression to leukemia was independent of trisomy 21 and originated in various stem and progenitor cells through additional mutations in cohesin genes. CD117+/KIT proto-oncogene (KIT) cells mediated the propagation of preleukemia and leukemia, and KIT inhibition targeted preleukemic stem cells.

Chronic AMPK activation via loss of FLCN induces functional beige adipose tissue through PGC-1α/ERRα.

The tumor suppressor folliculin (FLCN) forms a repressor complex with AMP-activated protein kinase (AMPK). Given that AMPK is a master regulator of cellular energy homeostasis, we generated an adipose-specific Flcn (Adipoq-FLCN) knockout mouse model to investigate the role of FLCN in energy metabolism. We show that loss of FLCN results in a complete metabolic reprogramming of adipose tissues, resulting in enhanced oxidative metabolism. Adipoq-FLCN knockout mice exhibit increased energy expenditure and are protected from high-fat diet (HFD)-induced obesity. Importantly, FLCN ablation leads to chronic hyperactivation of AMPK, which in turns induces and activates two key transcriptional regulators of cellular metabolism, proliferator-activated receptor γ (PPARγ) coactivator-1α (PGC-1α) and estrogen-related receptor α (ERRα). Together, the AMPK/PGC-1α/ERRα molecular axis positively modulates the expression of metabolic genes to promote mitochondrial biogenesis and activity. In addition, mitochondrial uncoupling proteins as well as other markers of brown fat are up-regulated in both white and brown FLCN-null adipose tissues, underlying the increased resistance of Adipoq-FLCN knockout mice to cold exposure. These findings identify a key role of FLCN as a negative regulator of mitochondrial function and identify a novel molecular pathway involved in the browning of white adipocytes and the activity of brown fat.

Haploinsufficiency of the ESCRT Component HD-PTP Predisposes to Cancer.

Endosomal sorting complexes required for transport (ESCRT) drive cell surface receptor degradation resulting in attenuation of oncogenic signaling and pointing to a tumor suppressor function. Here, we show that loss of function of an ESCRT protein (HD-PTP encoded by the PTPN23 gene, located on the tumor suppressor gene cluster 3p21.3) drives tumorigenesis in vivo. Indeed, Ptpn23(+/-) loss predisposes mice to sporadic lung adenoma, B cell lymphoma, and promotes Myc-driven lymphoma onset, dissemination, and aggressiveness. Ptpn23(+/-)-derived tumors exhibit an unaltered remaining allele and maintain 50% of HD-PTP expression. Consistent with the role of HD-PTP in attenuation of integrin recycling, cell migration, and invasion, hemizygous Ptpn23(+/-) loss increases integrin ß1-dependent B cell lymphoma survival and dissemination. Finally, we reveal frequent PTPN23 deletion and downregulation in human tumors that correlates with poor survival. Altogether, we establish HD-PTP/PTPN23 as a prominent haploinsufficient tumor suppressor gene preventing tumor progression through control of integrin trafficking.

Interplay of Endosomal pH and Ligand Occupancy in Integrin α5ß1 Ubiquitination, Endocytic Sorting, and Cell Migration.

Membrane trafficking of integrins plays a pivotal role in cell proliferation and migration. How endocytosed integrins are targeted either for recycling or lysosomal delivery is not fully understood. Here, we show that fibronectin (FN) binding to α5ß1 integrin triggers ubiquitination and internalization of the receptor complex. Acidification facilitates FN dissociation from integrin α5ß1 in vitro and in early endosomes, promoting receptor complex deubiquitination by the USP9x and recycling to the cell surface. Depending on residual ligand occupancy of receptors, some α5ß1 integrins remain ubiquitinated and are captured by ESCRT-0/I, containing histidine domain-containing protein tyrosine phosphatase (HD-PTP) and ubiquitin-associated protein 1 (UBAP1), and are directed for lysosomal proteolysis, limiting receptor downstream signaling and cell migration. Thus, HD-PTP or UBAP1 depletion confers a pro-invasive phenotype. Thus, pH-dependent FN-integrin dissociation and deubiquitination of the activated integrin α5ß1 are required for receptor resensitization and cell migration, representing potential targets to modulate tumor invasiveness.

FLCN and AMPK Confer Resistance to Hyperosmotic Stress via Remodeling of Glycogen Stores.

Mechanisms of adaptation to environmental changes in osmolarity are fundamental for cellular and organismal survival. Here we identify a novel osmotic stress resistance pathway in Caenorhabditis elegans (C. elegans), which is dependent on the metabolic master regulator 5'-AMP-activated protein kinase (AMPK) and its negative regulator Folliculin (FLCN). FLCN-1 is the nematode ortholog of the tumor suppressor FLCN, responsible for the Birt-Hogg-Dubé (BHD) tumor syndrome. We show that flcn-1 mutants exhibit increased resistance to hyperosmotic stress via constitutive AMPK-dependent accumulation of glycogen reserves. Upon hyperosmotic stress exposure, glycogen stores are rapidly degraded, leading to a significant accumulation of the organic osmolyte glycerol through transcriptional upregulation of glycerol-3-phosphate dehydrogenase enzymes (gpdh-1 and gpdh-2). Importantly, the hyperosmotic stress resistance in flcn-1 mutant and wild-type animals is strongly suppressed by loss of AMPK, glycogen synthase, glycogen phosphorylase, or simultaneous loss of gpdh-1 and gpdh-2 enzymes. Our studies show for the first time that animals normally exhibit AMPK-dependent glycogen stores, which can be utilized for rapid adaptation to either energy stress or hyperosmotic stress. Importantly, we show that glycogen accumulates in kidneys from mice lacking FLCN and in renal tumors from a BHD patient. Our findings suggest a dual role for glycogen, acting as a reservoir for energy supply and osmolyte production, and both processes might be supporting tumorigenesis.

Pseudophosphatases: methods of analysis and physiological functions.

Protein tyrosine phosphatases (PTPs) are key enzymes in the regulation of cellular homeostasis and signaling pathways. Strikingly, not all PTPs bear enzymatic activity. A considerable fraction of PTPs are enzymatically inactive and are known as pseudophosphatases. Despite the lack of activity they execute pivotal roles in development, cell biology and human disease. The present review is focused on the methods used to identify pseudophosphatases, their targets, and physiological roles. We present a strategy for detailed enzymatic analysis of inactive PTPs, regulation of inactive PTP domains and identification of binding partners. Furthermore, we provide a detailed overview of human pseudophosphatases and discuss their regulation of cellular processes and functions in human pathologies.

Induced apoptosis by mild hyperthermia occurs via telomerase inhibition on the three human myeloid leukemia cell lines: TF-1, K562, and HL-60.

The purpose of this research was to understand the effect of hyperthermia on the telomerase activity in human leukemic cell lines (HL-60, K562, and TF-1). The cells were treated by hyperthermia at the range of 41-44 degrees C for 120 min and incubated for 96 h. Then telomerase activity, cell proliferation, and apoptosis were assessed. The results indicated that hyperthermia significantly induced apoptosis on the cells. The cells exhibited pre-apoptotic pattern at 41 and 42 degrees C at 60-120 min and apoptotic pattern at 43 and 44 degrees C over 30 min after hyperthermia. Telomerase activity (that was assayed immediately after hyperthermia) was stable at 41-42 degrees C for 60 min but decreased to 35-40% at 120 min. However, at severe hyperthermia (43-44 degrees C) telomerase activity was decreased in a time- and dose-dependent manner. Following hyperthermia (41-44 degrees C up to 120 min), the cells were incubated for 96 h. In these conditions, the telomerase activity was decreased by about 60-80% in comparison with that untreated control cells.