Calmodulin mediates DNA repair pathways involving H2AX in response to low-dose radiation exposure of RAW 264.7 macrophages.
Understanding the molecular mechanisms that modulate macrophage radioresistance is necessary for the development of effective radiation therapies, as tumor-associated macrophages promote both angiogenesis and matrix remodeling that, in turn, enhance tumor metastasis. In this respect, we have identified a dose-dependent increase in the abundance (i.e., expression level) of the calcium regulatory protein calmodulin (CaM) in RAW 264.7 macrophages upon irradiation. At low doses of irradiation there are minimal changes in the abundance of other cellular proteins detected using mass spectrometry, indicating that increases in CaM levels are part of a specific radiation-dependent cellular response. CaM overexpression results in increased macrophage survival following radiation exposure, acting to diminish the sensitivity to low-dose radiation exposures. Following macrophage irradiation, increases in CaM abundance also result in an increase in the number of phosphorylated histone H2AX foci, associated with DNA repair, with no change in the extent of double-stranded DNA damage. In comparison, when nuclear factor kappaB (NFkappaB)-dependent pathways are inhibited, through the expression of a dominant-negative IkappaB construct, there is no significant increase in phosphorylated histone H2AX foci upon irradiation. These results indicate that the molecular basis for the up-regulation of histone H2AX-mediated DNA repair pathways is not the result of nonspecific NFkappaB-dependent pathways or a specific threshold of DNA damage. Rather, increases in CaM abundance act to minimize the low-dose hypersensitivity to radiation by enhancing macrophage radioresistance through processes that include the up-regulation of DNA repair pathways involving histone H2AX phosphorylation.
Smallwood
Heather S
HS
Washington State University Tri-Cities, Richland, Washington 99352, USA. Heather.Smallwood@StJude.org
Lopez-Ferrer
Daniel
D
Eberlein
P Elis
PE
Watson
David J
DJ
Squier
Thomas C
TC
D000818:Animals; D002147:Calmodulin / Q000378:metabolism*; D002460:Cell Line; D004249:DNA Damage; D004260:DNA Repair; D004307:Dose-Response Relationship, Radiation; D006657:Histones / Q000378:metabolism*; D008264:Macrophages / Q000378:metabolism / Q000528:radiation effects*; D051379:Mice
D002147:Calmodulin; D006657:Histones