Home About us Contact
|
Home About us Contact | ||
|
|
||
Monocytic Differentiation (monocytic + differentiation)
Selected AbstractsRegulated expression and intracellular localization of cystatin F in human U937 cellsFEBS JOURNAL, Issue 22 2002Carl-Michael Nathanson Cystatin F is a cysteine peptidase inhibitor recently discovered in haematopoietic cells by cDNA cloning. To further investigate the expression, distribution and properties of the native human inhibitor the promyeloid cell line U937 has been studied. The cells expressed relatively large quantities of cystatin F, which was found both secreted and intracellularly. The intracellular levels were unusually high for a secreted cystatin (, 25% of the cystatin F in 2- or 4-day culture medium). By contrast, U937 cells contained only 3,4% of the related inhibitor, cystatin C. Cystatin F purified from lysates of U937 cells showed three major forms carrying two, one or no carbohydrate chains. Immunocytochemistry demonstrated a marked cytoplasmic cystatin F staining in a granular pattern. Double staining with a marker for endoplasmic reticulum revealed no colocalization for cystatin F. Analysis of the promoter region of the cystatin F gene (CST7) showed that it, like that of the cystatin C gene (CST3), is devoid of typical TATA- and CAAT-box elements. In contrast to the cystatin C promoter, it does not contain multiple Sp1 binding sites, but has a unique site for C/EBP,, possibly explaining the restricted expression of the cystatin F gene. Cells stimulated with all- trans retinoic acid to differentiate them towards a granulocytic pathway, showed a strong (, 18-fold) down-regulation of intracellular cystatin F and almost abolished secreted levels of the inhibitor. Stimulation with tetradecanoyl phorbol acetate, causing monocytic differentiation, also resulted in down-regulation (two fold to threefold) of cystatin F expression, whereas the cystatin C expression was essentially unaltered in both experiments. The results suggest that cystatin F as an intracellular cysteine peptidase inhibitor with readily regulated expression, may be a candidate to control the cysteine peptidase activity known to be essential for antigen presentation in different blood cell lineages. [source] Jun N-terminal kinase pathway enhances signaling of monocytic differentiation of human leukemia cells induced by 1,25-dihydroxyvitamin D3JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 6 2003Qing Wang Abstract Recent studies revealed that the MEK/ERK module of the mitogen-activated protein kinase (MAPK) signaling cascades is up-regulated in the early stages of 1,,25-dihydroxyvitamin D3 (1,25D3)-induced monocytic differentiation of human leukemia cells HL60. In the present study, we investigated whether another MAPK module, the JNK pathway, also participates in this form of differentiation. We found that the dependence on the concentration of the inducer, the vitamin-hormone 1,25D3, in two types of human leukemia cells, HL60 and U937, and the kinetics of monocytic differentiation in HL60 cells, parallel the degree of the activation of the JNK pathway. A blockade of JNK signaling by a stable expression of dominant negative (dn) JNK1 mutant in U937 cells resulted in reduced c-jun phosphorylation, and the differentiation of these cells was markedly decreased. Similarly, inhibition of JNK1 and JNK2 activities by the selective inhibitor SP600125 led to both dose-dependent reduction of c-jun and ATF-2 phosphorylation, and of the differentiation of HL60 cells. In addition, we found that JNK activity is essential for the AP-1 DNA binding induced by 1,25D3 in HL60 and U937 cells. The results indicate that in cultured human leukemia cells, the JNK pathway participates in the induction of monocytic differentiation by 1,25D3, probably by activating the AP-1 transcription factor. © 2003 Wiley-Liss, Inc. [source] Kinase suppressor of RAS (KSR) amplifies the differentiation signal provided by low concentrations 1,25-dihydroxyvitamin D3JOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2004Xuening Wang The activity of kinase suppressor of ras (KSR), a kinase or a molecular scaffold upstream from Raf-1, is involved in the MEK/ERK MAP kinase cascade which can signal cell growth, survival, or differentiation, depending on the cellular context. We provide evidence here that KSR is upregulated in HL60 cells undergoing differentiation induced by low (0.3,3 nM) concentrations of 1,25-dihydroxyvitamin D3 (1,25D3), and an antisense oligo (AS), but not a sense oligo, to KSR inhibits this differentiation. The inhibition of differentiation by AS,KSR oligo was less apparent when the concentration of 1,25D3 was increased, suggesting that at the higher concentrations of 1,25D3 KSR is not essential for the signaling of the differentiated phenotype. The reduced differentiation of HL60 cells exposed to AS,KSR was paralleled by reduced phosphorylation of Raf-1 Ser 259, and of p90RSK, used here as read-out for MAPK cascade activity. Conversely, ectopic expression of Flag-tagged wild type KSR potentiated the differentiation-inducing effects of low concentrations of 1,25D3. Additional data suggest that the kinase activity of KSR is required for these effects, as transfection of a kinase inactive KSR construct did not significantly increase the 1,25D3 -induced differentiation. Enzyme assays performed with KSR immunoprecipitated from 1,25D3 -treated cells showed kinase activity when recombinant Raf-1 was used as the substrate, but not when the 1,25D3 -treated cells were pretreated with AS,KSR oligos. Taken together, these data suggest that KSR participates in signaling of monocytic differentiation by augmenting the strength of the signal transmitted through Raf-1 to downstream targets. J. Cell. Physiol. 198: 333,342, 2004© 2003 Wiley-Liss, Inc. [source] | ||||||||||