Mara Jeffress completed her Ph.D. in Molecular & Cellular Biology in 2004 at the University of Washington in Seattle in the laboratory of Dr. Stanley Fields. Her doctoral thesis work focused on discovery of novel mechanisms of malaria drug resistance, specifically, P. falciparum resistance to the drug mefloquine (Larium). Please find her dissertation abstract here:

Identification of putative mefloquine resistance genes from Plasmodium falciparum (1999-2004, Laboratory of Dr. Stanley Fields. Department of Genome Sciences, University of Washington)

Mefloquine (MQ) is an effective antimalarial drug; however, resistant strains of the human malarial pathogen, Plasmodium falciparum, are beginning to arise. The yeast S. cerevisiae is sensitive to mefloquine, enabling a screen for P. falciparum genes involved in resistance. Yeast were transformed with a P. falciparum expression library, followed by selection on mefloquine plates and sequencing of plasmids that conferred resistance. I characterized the four genes that exhibited the strongest mefloquine resistant phenotype in yeast. All four (PFD0090c, PFI0195c, PF10_0372 and PF14_0649) are uncharacterized P. falciparum genes from distinct chromosomes (4, 9, 10 and 14, respectively). The mefloquine-resistant phenotype was dependent on induction of the P. falciparum gene and independent of vector context.

PFI0195c, which likely encodes a small GTPase activator (GAP), also conferred resistance to cycloheximide in yeast. Immunolocalization showed that the encoded protein colocalized with the yeast GTPase Ypt6 to the Golgi complex, which is consistent with potential GAP function. Overexpression of YPT6 causes MQ resistance and overexpression of PFI0195c cannot cause MQ resistance when YPT6 is deleted. These data suggest that PFI0195c mediates MQ resistance by acting through Ypt6 in yeast, and potentially through the Ypt6 homolog, PfRab6, in P. falciparum.

The other three candidate proteins localized to the cytoplasm and plasma membrane (PF14_0649), nuclear envelope/ER (PF10_0372) and Golgi (PFD0090c) of yeast. PF10_0372 interacted with six yeast proteins Gtt1, Gtt3, Mga2, Bap3, Iks1 and Prm3 in the two-hybrid assay. The interactions among these six proteins and their links to yeast drug resistance, cellular detoxification, vesicular transport and cell membrane integrity genes are discussed in my dissertation. Additionally, overexpression of PF10_0372 cannot mediate MQ resistance when the GTT1 gene, encoding glutathione-S-transferase, is deleted, suggesting that PF10_0372 affects glutathione detoxification of mefloquine.

Analysis of mefloquine-resistant P. falciparum strains and the mefloquine-sensitive strain, W2, by sequencing and semi-quantitative RT-PCR, identified no relevant mutation in the resistant strains. However, PFI0195c was upregulated in two out of three resistant strains and PF14_0649 was upregulated in all resistant strains tested, suggesting a role in MQ resistance that should be examined by overexpression and MQ testing in P. falciparum parasites. Pub Med Publication