Acetyl CoA Carboxylase in Avian Eimeria
P.C. Allen, K.B. Miska, M.C. Jenkins, R.F. Fetterer.
USDA/ARS, Beltsville, Maryland, USA. pallen@anri.barc.usda.gov
Acetyl coA carboxylase, catalyzes the first committed
step in the biosynthesis of fatty acids. In some apicomplexan parasites such as
Toxoplasma gondii and Plasmodium sp., a multi-domain form of this enzyme (ACC 1)
has been identified as a nuclear-encoded protein that is transported to and
apparently functions in the apicoplast organelle found in zoite stages of
parasites life cycles. The unique genetic origin of this enzyme and its apparent
requirement for parasite development suggests its potential as a target for
control of apicomplexan parasites. We have therefore undertaken the
characterization of ACC1 in avian Eimeria. BLAST searches of both nucleotide and
protein data files from T. gondii ACC1 (tACC1) precursor against the Sanger
Institute Eimeria tenella genome data base found a putative homolog Eimeria ACC1
(eACC1) gene to be contained in Contig 5754, and the carbamyl phosphate synthase,
(5’end), biotin binding, and carboxyl transferase (3’ end) domains were
identified both through bioinformatics as well as RT-PCR. The low genetic
complexity of the central portion of the eACC1 gene made its complete sequencing
difficult. A 224 nt sequence that is 73 nt upstream from the first exon common
to tACC1 and eACC1 was identified as a possible signal sequence for eACC1.
However, no tripartite leader sequence, such as is found in tACC1 precursor that
allows transport of the protein into an apicoplast, was defined. These
observations, along with the lack of evidence for an Eimerian apicoplast in the
ultrastructure literature suggest major differences between tACC1 and eACC1.
Expression of eACC1 protein was documented through detection on polyacrylamide
gels of a biotin-containing protein band of about 250 kd. This band appears most
intense in merozoite extracts as compared to extracts of oocysts and sporozoites.
Determinations of differential gene and protein expression of eACC1 among stages
of development are ongoing through use of RT-PCR and Western blots of extracts.