Vincristine-resistant erythroleukemia cell line has marked increased sensitivity to hexamethylenebisacetamide-induced differentiation.

Abstract

Hexamethylenebisacetamide (HMBA)-induced murine erythroleukemia (MEL) differentiation is a multistep process. Commitment is the capacity to express terminal cell division and characteristics of the differentiated phenotype even after the cells are removed from culture with inducer. Culture of MEL cell line 745A-DS19 (DS19) with HMBA causes commitment to terminal differentiation after a latent period of about 10-12 hr. Previous studies have shown that during this latent period, HMBA causes a number of metabolic changes, including modulation in expression of certain protooncogenes. We now report the development of a MEL cell line (designated V3-17) derived from DS19 that is resistant to vincristine and is (i) markedly more sensitive to HMBA, (ii) induced to commitment without a detectable latent period, and (iN) resistant to the effects of phorbol ester and dexamethasone, which are potent inhibitors of HMBAmediated DS19 differentiation. We suggest that this V3-17 MEL cell line may express a factor that circumvents HMBAmediated early events, which prepare the cells for commitment to terminal differentiation.Hexamethylenebisacetamide (HMBA)-mediated murine erythroleukemia (MEL) cell terminal differentiation is a multistep process (1, 2). Upon culture of MEL cell line 745A-DS19 (DS19) (3) with HMBA (4), there is a latent period of -10 to 12 hr during which commitment to terminal differentiation cannot be detected. Commitment is defined as the capacity to express characteristics of the erythroid differentiated phenotype, including loss of proliferative capacity, despite removal of the inducer (5, 6). This early, latent period is followed by a period during which an increasing proportion of the population expresses characteristics of terminal differentiation, including loss of proliferative capacity.During the latent period, the inducer initiates a number of metabolic changes that precede irreversible commitment to differentiation. Among these are alterations in membrane permeability, which involve Na', K+, and Ca2+ flux (7-9); changes in cell volume (10); a transient increase in cyclic AMP concentration (11); a prompt increase in membraneassociated protein kinase C activity (PKC); the appearance in the cytosol of a Ca2 + -and phospholipid-independent form of PKC, presumably generated by proteolytic cleavage of membrane-bound PKC (12); and the modulation in expression of a number of genes, including c-myb, c-myc, c-fos, and p53 (13)(14)(15)(16). Upon more prolonged culture with HMBA, DS19 cells become irreversibly committed (5, 6). Morphological and molecular changes occur that are similar to normal erythroid terminal cell differentiation, including the coordinated expression of genes for a'-and 83mj-globin, for the heme synthetic enzymes, and for erythroid-specific membrane proteins, as well as suppression of DNA replication and rRNA synthesis (1,17,18).In the present studies we describe the development of a MEL cell line derived from DS19 that is resistant to inhibition of cell growth by vincristine and is de...

Changes in intracellular localization of calpastatin during calpain activation

“…Murine erythroleukemia (MEL) cells were obtained and cultured as specified previously [11]. Human pheochromocytoma cells (PC12) were purchased from the American Type Culture Collection (A.T.C.C.)…”

cAMP-induced NF-κB (p50/relB) binding to a c-myb intronic enhancer correlates with c-myb up-regulation and inhibition of erythroleukemia cell differentiation

“…Both cAMP-dependent protein kinase (A-kinase) and protein kinase C are necessary for MEL cell dierentiation and ®nely regulated changes in the activity and isozyme pattern of these two protein kinases have been described during dierentiation (Pilz et al, 1992;Schwartz and Rubin, 1985;Melloni et al, 1989Melloni et al, , 1990. Paradoxically, MEL cell dierentiation is inhibited when the cells are treated with cAMPanalogues or phorbol esters which activate A-kinase or protein kinase C, respectively (Yamasaki et al, 1977;Mignotte et al, 1990;Sherman et al, 1986Sherman et al, , 1988.…”

A 6 kDa protein homologous to the N‐terminus of the HMG1 protein promoting stimulation of murine erythroleukemia cell differentiation

“…N23 and C44 MEL cell clones [9] were cultured in a-MEM containing 10% fetal calf serum as previously described [10]. N23 MEL cells (105 cells/ml) were induced to differentiate by addition of 5 mM HMBA.…”

“…oc-PKC was isolated from MEL cells [9] and assayed as previously specified [14]. c~-PKC stimulating activity of different protein preparations was evaluated as specified elsewhere [3].…”