Analysis of the conserved cysteine periodicity of Paramecium variable surface antigens.

The major surface antigens expressed by free-living and parasitic protozoa commonly contain repeating cysteine motifs. Despite the common occurrence of these repeats their functional significance remains largely unexplored. In this paper we investigate the conserved cysteine repeats within the variable surface antigens of Paramecium tetraurelia. We show that deletion of 2 entire repeating units or portions of repeats near the N-terminus does not prevent expression of the A51 variable surface antigen. Alteration of a single cysteine to serine residue also has no effect on A51 expression. In contrast, deletions near the C-terminus of the protein have identified a small segment within the repeats that is required for expression on the surface. The required region contains a number of conserved amino acid residues, yet site-directed mutagenesis of two residues (serine and threonine to alanine) did not prevent expression. These studies demonstrate the feasibility of using deletion analysis to identify regions critical for the expression of cysteine-rich surface antigens. The relationship of these results to the structure and expression of cysteine-rich surface proteins in other protozoa is discussed.

Evidence for transcriptional self-regulation of variable surface antigens in Paramecium tetraurelia.

Variable surface antigens are commonly found on free-living and parasitic protozoa, yet the regulation of antigen expression and switching is not fully understood in any system. A cell line of Paramecium tetraurelia stock 51 can express at least 11 different antigens yet only one type is found on the surface at any one time. Previous studies have shown that mutually exclusive expression of Paramecium surface antigens can be overcome if two antigen genes contain the same 5' coding region. In this article we utilize a gene chimera containing portions of A51 and B51 to analyze the effect of a frameshift mutation on transcription and steady-state mRNA levels. We show that a frameshift mutation near the 3' end prevents expression of the protein on the cell surface and reduces the rate of transcription of the corresponding gene. The difference in transcription is not the result of differences in plasmid copy number. We propose that expression of the antigen on the cell surface is part of a self-regulatory pathway that influences transcription of the corresponding gene. A model incorporating the previous and current data is presented.