Mutated D4-guanine diphosphate–dissociation inhibitor is found in human leukemic cells and promotes leukemic cell invasion

• Published in: Experimental hematology Vol. 36; no. 1; pp. 37 - 50
• Main Authors: Nakata, Yuji, Kondoh, Kensuke, Fukushima, Sachiko, Hashiguchi, Akinori, Du, Wenlin, Hayashi, Mutsumi, Fujimoto, Jun-ichiroh, Hata, Jun-ichi, Yamada, Taketo
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier Inc 2008
• Subjects: Advanced Basic Science; Amino Acid Sequence; Amino Acid Substitution; Animals; Burkitt Lymphoma - pathology; Cell Adhesion; Cell Line, Tumor - metabolism; Cell Movement; Child; Conserved Sequence; Female; Genes, Dominant; Guanine Nucleotide Dissociation Inhibitors - genetics; Guanine Nucleotide Dissociation Inhibitors - physiology; Guanosine Diphosphate - metabolism; Hematology, Oncology and Palliative Medicine; Humans; Leukemia, T-Cell - pathology; Mice; Mice, SCID; Molecular Sequence Data; Mutation, Missense; Neoplasm Invasiveness - physiopathology; Neoplasm Proteins - genetics; Neoplasm Proteins - physiology; Paresis - etiology; Precursor Cell Lymphoblastic Leukemia-Lymphoma - pathology; rho GTP-Binding Proteins - metabolism; rho Guanine Nucleotide Dissociation Inhibitor beta; rho-Specific Guanine Nucleotide Dissociation Inhibitors; Sequence Alignment; Tumor Suppressor Proteins - genetics; Tumor Suppressor Proteins - physiology
• ISSN: 0301-472X; 1873-2399
• DOI: 10.1016/j.exphem.2007.08.023

Objective Rho GTPase may be involved in human cancer invasion via the augmentation of cell motility and adhesion. We report on two point mutations of the D4-guanine diphosphate (GDP)–dissociation inhibitor (GDI) gene, one of the Rho-GDIs, which were found in a human leukemic cell line, Reh, and the mutated D4-GDI functions as an accelerator of leukemic cell invasion. Material and Methods We investigated the altered activity of GDP dissociation by mutated (mt) D4-GDI and the functions of this mt and wild-type (wt) D4-GDI in invasion. The mice inoculated with wt or mt D4-GDI vector–transfected Raji cells were observed and examined pathologically. Adhesiveness and cell motility of wt or mt D4-GDI vector–transfected Raji cells were examined. Finally, it was examined whether Rho activation was changed by mutation of D4-GDI under the condition of Rho-GDI knockdown. Results Two point mutations of the D4-GDI gene were found in Reh cells. The region of mutations is conserved among members of the Rho-GDI family at the amino acid level. D4-GDI with two mutations (V68L and V69A) functioned in a dominant negative manner in the inhibition of GDP dissociation from Rho. Severe combined immune-deficient mice inoculated with Raji cells developed hemiparalysis. The Raji cells were present in bone marrow and peripheral blood, and hepatic invasion was observed in 20% of the mice. Mice inoculated with wt D4-GDI vector–transfected Raji cells (wt D4) showed later paralysis and none developed hepatic invasion. Mice inoculated with mt D4-GDI–transfected Raji cells (mt D4) showed a 5-day reduction in the time to paraplegia and death. In addition, hepatic invasion was evident in 80% of mice transplanted with mt D4 cells. There were no differences in growth rates and amounts of guanine triphosphate (GTP)–bound Rho, cdc42, or Rac among all clones, however, GTP-bound Rho in mt D4 clone with short hairpin RNA (shRNA) vector for Rho-GDI knockdown was increased compared with wt D4 clone with shRNA vector for Rho-GDI knockdown. The mt D4 cells showed an augmentation of adhesiveness and cell motility. On the other hand, wt D4 cells showed a decreased ability of cell motility. Conclusion These results suggest the mutated D4-GDI functions as a dominant negative molecule against the wt D4-GDI and accelerates invasion via regulation of cytoskeletal machinery.