Molecular analysis of the bison phagocyte NADPH oxidase: cloning and sequencing of five NADPH oxidase cDNAs.
During the host defense process, neutrophils migrate into infected tissues where they become activated, resulting in the assembly of a superoxide anion-generating complex known as the NADPH oxidase. Despite the importance of this system in animal host defense, almost nothing is known about the NADPH oxidase in neutrophils from wild ruminant species. In the present studies, we provide a molecular analysis of the bison leukocyte NADPH oxidase. Using reverse transcriptase-polymerase chain reaction and rapid amplification of cDNA ends, we cloned and sequenced the full-length cDNAs for five bison NADPH oxidase components: p22(phox), p40(phox), p47(phox) and p67(phox), and gp91(phox). When compared to other species, the deduced amino acid sequences of the bison homologs were most similar to those of bovine. Interestingly, a bison p40(phox) alternative splice product was isolated, which was similar to that observed for human p40(phox) in that the cDNAs contained sequence from intron 8. Consistent with the high degree of similarity between bison and bovine amino acid sequences, immunoblot analysis showed that the bison homologs migrated similarly to their bovine counterparts. Overall, these studies show that the bison and bovine NADPH oxidase genes are highly conserved between these two species, despite their divergence from a common ancestor over 1 million years ago.
Gauss
Katherine A
KA
Department of Veterinary Molecular Biology and Center for Bison and Wildlife Health, Montana State University, Bozeman, MT 59717, USA.
Bunger
Peggy L
PL
Siemsen
Daniel W
DW
Young
Catherine J
CJ
Nelson-Overton
Laura
L
Prigge
Justin R
JR
Swain
Steve D
SD
Quinn
Mark T
MT
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D016164:Bison / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry*; D008562:Membrane Glycoproteins / Q000235:genetics; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D009252:NADPH Dehydrogenase / Q000235:genetics; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000096:biosynthesis / Q000235:genetics*; D009504:Neutrophils / Q000201:enzymology; D010586:Phagocytes / Q000201:enzymology*; D010750:Phosphoproteins / Q000235:genetics; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA
D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D010750:Phosphoproteins; C085072:neutrophil cytosol factor 40K; C079789:neutrophil cytosol factor 67K; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; C086986:CYBA protein, human; C060505:neutrophil cytosolic factor 1; D009252:NADPH Dehydrogenase