cytochalasin-b and Leukemia--Myelogenous--Chronic--BCR-ABL-Positive

BRIT1 (BRCT-repeat inhibitor of hTERT expression), also known as microcephalin (MCPH1), is a crucial gene in the complex cellular machine that is devoted to DNA repair and acts as a regulator of both the intra-S and G2/M checkpoints. The most important role of BRIT1/MCPH1 in the regulation of cell cycle progression appears to be the G2/M checkpoint. The K562 and peripheral blood cells of chronic myeloid leukemia (CML) patients at diagnosis were found to downregulate BRIT1/MCPH1. However, we could not find any correlation between bcr/abl activity and the BRIT1/MCPH1 level. In order to study the genomic instability of CML cells, we evaluated the ability of these cells to arrest mitotic division after exposure to hydroxyurea, a known genotoxic agent. We showed that CML cells continue to proliferate without the activation of the G2/M cell cycle checkpoint arrest or of the apoptotic mechanism. This behavior may predispose the cells to accumulate genomic defects. In conclusion, we found that CML cells have a low BRIT1/MCPH1 level and show a defective G2/M arrest, confirming that these cells have a constitutive genomic instability.

Topics: Actins; Adult; Aged; Cell Cycle Proteins; Cell Proliferation; Cells, Cultured; Cytochalasin B; Cytokinesis; Cytoskeletal Proteins; Female; G2 Phase Cell Cycle Checkpoints; Gene Expression Regulation, Neoplastic; Genomic Instability; Humans; Hydroxyurea; K562 Cells; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Leukocytes; Male; Middle Aged; Mutagens; Neoplasm Proteins; Nerve Tissue Proteins; RNA, Messenger

Lectin-receptors on leukocyte and endothelial surfaces are becoming more important in the light of increasing evidence which implicates lectin-carbohydrate interactions in diverse physiological phenomena. This study reports the identification of a major 118 kDa granulocyte surface protein, (Protein 1 a) which binds the lectin wheat germ agglutinin (WGA), and is distinctly different from reported WGA binding granulocyte membrane proteins. Protein 1 a has been isolated from the Triton-soluble and Triton-insoluble lysates of normal individuals and patients with Chronic Myeloid Leukemia (CML) using a combination of differential solubilization, lectin affinity, ion exchange chromatography and HPLC. The protein from the detergent lysates of both normal and CML granulocytes has similar pI values, lectin affinities, and hydrophobicity. However, its solubility in Triton is different in the two cell types. In 71% of CML cases examined, Protein 1 a exhibits decreased Triton solubility suggesting its increased association with the cytoskeleton (CSK). Stimulation of normal granulocytes with WGA leads to the translocation of the soluble form of Protein 1 a to the Triton-insoluble fraction. This cytoskeletal recruitment of Protein 1 a is sustained only under conditions of excess WGA and occupied receptor. The CSK disruptive agent dihydrocytochalasin B (H2CB) releases the insoluble form of the receptor into the Triton-soluble fraction. Investigation of a CSK-involving process such as ligand internalization revealed that CML granulocytes exhibit slower kinetics of internalization of fluorescent WGA molecules.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Antigens, CD; Concanavalin A; Cytochalasin B; Cytoskeleton; Endocytosis; Female; Granulocytes; Humans; In Vitro Techniques; Kinetics; Leukemia, Myelogenous, Chronic, BCR-ABL Positive; Leukosialin; Male; Polyethylene Glycols; Precipitin Tests; Receptors, Cell Surface; Receptors, Mitogen; Sialoglycoproteins; Solubility; Time Factors; Wheat Germ Agglutinins