Cloning and expression of the cDNA encoding rat granulocyte colony-stimulating factor

Granulocyte colony-stimulating factor (G-CSF) acts on precursor hematopoietic cells to control the production and maintenance of neutrophils. Recombinant G-CSF (re-G-CSF)is used clinically to treat patients with neutropenia and has greatly reduced the infection risk associated with bone marrow transplantation. Cyclic hematopoiesis, a stem cell defect characterized by severe recurrent neutropenia, occurs in man and grey collie dogs, and can be treated by administration of re-G-CSF. Availability of the rat G-CSF cDNA would benefit the use of rats as models of gene therapy for the treatment of cyclic hematopoiesis. In preliminary rat experiments, retroviral-mediated expression of canine G-CSF caused neutralizing antibody formation which precluded long-term increases in neutrophil counts. To overcome this problem we cloned the rat G-CSF cDNA from RNA isolated from skin fibroblasts. The rat G-CSF sequence shared a high degree of identity in both the coding and non-coding regions with both the murine G-CSF (85%) and human G-CSF (74%). The signal peptides of murine and human G-CSF both contained 30 amino acids (aa), whereas the deduced signal sequence for rat G-CSF possessed 21 aa. A retrovirus encoding the rat G-CSF cDNA synthesized bioactive G-CSF from transduced vascular smooth muscle cells.

Keywords

Gene therapy, Neutropenia, Nucleotide sequence, complementary dna, granulocyte colony stimulating factor, neutralizing antibody, recombinant granulocyte colony stimulating factor, animal cell, animal experiment, animal model, bone marrow transplantation, dog, gene therapy, graft infection, hematopoiesis, hematopoietic cell, human, human cell, molecular cloning, neutropenia, neutrophil, nonhuman, priority journal, rat, retrovirus, stem cell, vascular smooth muscle, Amino Acid Sequence, Animals, Base Sequence, Cloning, Molecular, DNA, Complementary, Genetic Vectors, Granulocyte Colony-Stimulating Factor, Mice, Molecular Sequence Data, Rats, Sequence Analysis, DNA, Sequence Homology, Animalia, Canis familiaris, Murinae, unidentified retrovirus