CELLULAR CONTROL OF HEMOPOIETIC GROWTH FACTOR PRODUCTION

Project: Research project

Description

DESCRIPTION: The availability of
recombinant colony-stimulating factors (CSFs) for clinical use has
presented medical science with its first opportunity to stimulate blood
cell formation and host defense in man. Potential benefits of CSF
administration include treating primary hematologic diseases, enhancing
host defense against infection, and preventing chemotherapy-induced
cytopenias. Elucidating the role colony-stimulating factors play in normal
hematopoiesis, inflammation and leukemia cell growth requires a thorough
understanding of the mechanisms by which cells control their production of
CSFs. Studies on the inducible expression of GM-CSF in T cells and the
constitutive expression of GM-CSF in HTLV infected T cell lines have
identified a critical region of the GM-CSF promoter required for its
mitogen-inducible activity and its responsiveness to the HTLV-I and -II tax
proteins. In this grant we propose to investigate the DNA-protein
interactions in this region to understand the role that expression of
sequence-specific DNA-binding proteins play in the normal and abnormal
production of hematopoietic growth factors. We will identify the precise
nucleotide sequence recognized by GM-CSF promoter DNA-binding proteins by
mutating specific nucleotides in the promoter and analyzing their effect on
the binding of transcription factors using gel shift and DNase I
footprinting assays. The proteins binding specifically to the GM-CSF
promoter sequences will be characterized using Southwestern blotting. UV
cross-linking and biochemical manipulations designed to identify
post-transcriptional protein modifications. We will use these techniques
to determine the presence and activity of these factors in nuclear extracts
prepared from stimulated and unstimulated mature T cell lines and from a
variety of immature or transformed cell lines. T cell DNA-binding proteins
will be purified using DNA affinity chromatography and amino acid
sequencing will enable us to design oligonucleotides for cloning
DNA-binding factors from a T cell cDNA library. We will obtain polyclonal
antisera for screening our lambda gtll T cell cDNA expression library and
will also use radiolabelled DNA recognition site probes to isolate
transcription factor cDNA clones. The studies proposed will provide
valuable insights into the transcriptional mechanisms controlling
hematopoiesis.
StatusFinished
Effective start/end date9/1/903/31/01

Funding

  • National Institutes of Health: $218,795.00
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health: $169,620.00
  • National Institutes of Health: $251,738.00
  • National Institutes of Health: $183,458.00
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health

Fingerprint

Colony-Stimulating Factors
Intercellular Signaling Peptides and Proteins
Myeloid Leukemia
T-Lymphocytes
Acute Myeloid Leukemia
DNA-Binding Proteins
Gene Library
DNA
Southwestern Blotting
Cell Line
Human T-lymphotropic virus 2
TCF Transcription Factors
Core Binding Factor Alpha 2 Subunit
Transformed Cell Line
Human T-lymphotropic virus 1
Deoxyribonucleases
Hematologic Diseases
Organized Financing
Transcription Factors
Affinity Chromatography

Keywords

  • Medicine(all)