Multiple Splice Variants Encode a Novel Adenylyl Cyclase of Possible Plastid Origin Expressed in the Sexual Stage of the Malaria Parasite Plasmodium falciparum

Abstract

We report the characterization of an unusual adenylyl cyclase gene from Plasmodium falciparum, here designated PfAC␣. The level of mRNA expression is maximum during development of gametocytes (the sexual blood stage of the parasite life cycle). The gene is highly interrupted by 22 introns, and reverse transcriptase-PCR analysis revealed that there are multiple mRNA splice variants. One intron has three alternative 3-splice sites that confer the potential to encode distinct forms of the enzyme using alternative start codons. Deduced amino acid sequences predict membrane-spanning regions, the number of which can vary between two and six depending on the splice variant. Expression of a synthetic form of two of these variants in Xenopus oocytes and in Dictyostelium adenylyl cyclase-deficient mutants, confirms that PfAC␣ is a functional adenylyl cyclase. These results identify a novel mechanism in P. falciparum for the generation of multiple isoforms of a key, membranebound signaling molecule from a single genomic copy. Comparisons of the catalytic domains of PfAC␣ and a second putative P. falciparum adenylyl cyclase (PfAC␤) with those from other species reveal an unexpected similarity with adenylyl cyclases from certain prokaryotes including the cyanobacteria (blue green algae). In addition, the presence of an unusual active site substitution in a position that determines substrate specificity, also characteristic of these prokaryotic forms of the enzyme, further suggests a plastid origin for the Plasmodium cyclases.

Conservation of functional domain structure in bicarbonate-regulated ?soluble? adenylyl cyclases in bacteria and eukaryotes

“…1a). A putative sAC-like cyclase, with tandemly arranged catalytic domains, was recently detected within a hypothetical open reading frame from the malaria parasite Plasmodium falciparum (Muhia et al 2003), but its complete domain organization is not yet known. We also found a number of interesting sAC-like sequences in the genome of the thermophilic bacterium, C. aurantiacus.…”

Molecular Details of cAMP Generation in Mammalian Cells: A Tale of Two Systems

“…20,42 Bicarbonate-regulated sAC-like cyclases are also found in Anabaena PCC 7120, Stigmatella aurantiaca, Mycobacterium tuberculosis, 43 and the thermophilic bacterium Chloroflexus aurantiacus, 44 and sAC-like proteins were cloned and expressed from sea urchin 45 and Plasmodium falciparum, the parasite responsible for the most lethal form of malaria. 46 Database searches identify sAC-like ACs in the genomes of all mammals studied (human, chimpanzee, dog, cow, rabbit, mouse, and rat), as well as in chicken, ciona intestinalis, and insects (mosquito and honey bee). The human, chimpanzee, dog, and cow genomes predict a second locus encoding a sAC-like gene, but its activity or physiological relevance is unknown.…”

Niche metabolism in parasitic protozoa

“…In many eukaryotes, including some microbial parasites (e.g. Muhia et al 2003;Loftus et al 2005b), cohorts of highly related proteins with subtly different biochemical properties can be generated either by alternatively splicing different exons from a single gene, or initiating the transcription of a single gene at alternative start sites. Trypanosomatid parasites are unique, however, in that they fail to regulate gene expression at the level of transcript initiation (Clayton 2002)-there are no individual transcription promoters for protein-coding genes transcribed by RNA polymerase II-and the coding regions of individual genes are, with only four possible exceptions, not interrupted by intervening non-coding sequences (introns).…”