Surface glycoprotein PSA (GP46) expression during short- and long-term culture of Leishmania chagasi

Abstract

The mRNAs encoding promastigote surface antigen (PSA) of Leishmania chagasi have previously been shown to increase about 30-fold as in vitro cultured parasites progress from logarithmic to stationary phase, growth phases that are, respectively associated with parasites having low and high infectivity to mammals. Experiments reported here establish by western blot analysis that PSA proteins of 44 and 66 kDa also increase about 30-fold as parasite cultures reach stationary phase. Serial passage of parasite cultures resulted in a progressive reduction in PSA protein and RNA abundance to levels less than 3% that of cultures newly-initiated with parasites derived from a parasitized rodent. Loss of PSA mRNA abundance in serially passaged cells was not due to reduced PSA gene transcription rates, as determined by nuclear run-on assays. Neither was the loss associated with a marked decrease in PSA mRNA stability. Analysis of PSA RNA stability in the presence of actinomycin D, an inhibitor of transcription elongation, failed to detect a difference in fully processed cytosolic PSA mRNA stability regardless of the number of times a culture was passaged or the growth phase of the culture. Based on the lack of detectable difference in (cytosolic) mature PSA mRNA stability during promastigote development, the data indirectly suggest that the regulated expression of PSA in cells from low-passage cultures and the loss of PSA expression in high-passage cultures may be mediated by nuclear events that occur after transcription of the PSA genes and before arrival of the mature mRNAs in the cytoplasm.