ABSTRACT
Janus kinase 2 (JAK2)V617F-activating mutations (JAK2mu) occur in myeloproliferative disorders (MPDs) and myelodysplastic syndromes (MDSs). Cell lines MB-02, MUTZ-8, SET-2 and UKE-1 carry JAK2V617F and derive from patients with MPD/MDS histories. Challenging the consensus that expression of JAK2V617F is the sole precondition for cytokine independence in class I cytokine receptor-positive cells, two of four of the JAK2mu cell lines were growth factor-dependent. These cell lines resembled JAK2wt cells regarding JAK2/STAT5 activation: cytokine deprivation effected dephosphorylation, whereas erythropoetin or granulocyte colony-stimulating factor induced phosphorylation of JAK2 and STAT5. Cytokine independence correlated with low expression and cytokine dependence with high expression of the JAK/STAT pathway inhibitor suppressor of cytokine signaling 2 (SOCS2) suggesting a two-step mechanism for cytokine independence of MPD cells: (i) activation of the oncogene JAK2V617F and (ii) inactivation of the tumor suppressor gene SOCS2. Confirming that SOCS2 operates as a negative JAK2V617F regulator, SOCS2 knockdown induced constitutive STAT5 phosphorylation in JAK2mu cells. CpG island hypermethylation is reported to promote SOCS gene silencing in malignant diseases. Accordingly, in one of two cytokine-independent cell lines and in two of seven MPD patients, we found SOCS2 hypermethylation associated with reduced promoter access to transcription factors. Our results provide solid evidence that SOCS2 epigenetic downregulation might be an important second step in the genesis of cytokine-independent MPD clones.
Subject(s)
Janus Kinase 2/genetics , Janus Kinase 2/metabolism , Myeloproliferative Disorders/genetics , Myeloproliferative Disorders/metabolism , Suppressor of Cytokine Signaling Proteins/genetics , Suppressor of Cytokine Signaling Proteins/metabolism , Apoptosis/physiology , Cell Division/physiology , Cell Line , Chronic Disease , Cytokines/metabolism , Epigenesis, Genetic/physiology , Gene Expression Regulation, Leukemic , Gene Silencing/physiology , Humans , Methylation , Myeloproliferative Disorders/pathology , Phosphorylation , Point Mutation , STAT5 Transcription Factor/metabolism , Signal Transduction/physiologyABSTRACT
A newly developed spectrometer for energy-dispersive micro X-ray fluorescence spectrometry has been designed for the demands of archaeometry. ArtTAX combines the advantages of non-destructive and sensitive multi-elemental analysis at sub-mm resolution with the possibility of working outside the laboratory. The spectrometer consists of an air-cooled, low-power molybdenum tube, new generation polycapillary X-ray optics, a silicon drift detector without the need for liquid-nitrogen cooling, a CCD camera, and three light diodes for sample positioning. The motor-driven measurement head is fixed on a x,y,z-flexible tripod support which can be assembled and dismantled within minutes. The spot size of the primary X-ray beam was determined to be 94 microm for the Cu(Kalpha) energy, the detection limits are in a range of a few tens of microg g(-1) for the medium energy-range in glass. Additional open helium purging in the excitation and detection paths enables the determination of elements down to sodium, thus avoiding vacuum conditions or a size-limiting sample chamber. A selection of qualitative and quantitative results on pigment, metal, glass, and enamel analyses are presented to show the potential of ArtTAX in the field of art and archaeology.
