Cloning and expression of unique murine macrophage colony-stimulating factor transcripts

Area of Science:

• Hematopoiesis and Cellular Signaling
• Molecular Biology and Genetics
• Radiation Biology

Background:

• Cocultivation of irradiated murine bone marrow stromal cells with hematopoietic progenitor cells induces factor-independent sublines.
• Evidence suggests macrophage colony-stimulating factor (M-CSF) or a related protein (leukemogenic stromal factor, LSF) may mediate this emergence.

Purpose of the Study:

• To isolate and characterize M-CSF-related factors involved in hematopoietic cell line development.
• To investigate the role of specific M-CSF transcripts in bone marrow stromal cell response to irradiation.

Main Methods:

• Isolation of molecular clones from a D2XRII cDNA library using a mouse M-CSF genetic probe.
• Analysis of cDNA clones for deletions within the M-CSF coding region.
• Detection and quantification of specific RNA transcripts using Northern blot and reverse transcription-polymerase chain reaction (RT-PCR).
• Assessment of transcript levels following gamma irradiation.

Main Results:

• Two cDNA clones (60.8.2 and 6452) with an 885-bp deletion in the M-CSF coding region were identified.
• Three specific transcripts (4.8, 3.4, and 1.8 kb) containing the deletion were detected.
• Levels of 4.8-kb and 3.4-kb deletion-containing transcripts, along with other M-CSF transcripts, increased 1.4- to 2.2-fold after gamma irradiation.
• Deletion-specific transcripts were found in multiple mouse bone marrow stromal cell lines and normal tissues.

Conclusions:

• Murine bone marrow stromal cells express a broader spectrum of M-CSF transcripts than previously recognized.
• The increased accumulation of specific M-CSF transcripts post-irradiation supports their role in stromal cell response to ionizing radiation.