Control of variant surface glycoprotein gene-expression sites in Trypanosoma brucei

• Published in: The EMBO journal Vol. 18; no. 17; pp. 4846 - 4855
• Main Authors: Chaves, I, Rudenko, G, Dirks-Mulder, A, Cross, M, Borst, P
• Format: Journal Article
• Language: English
• Published: Chichester, UK John Wiley & Sons, Ltd 01.09.1999
• Subjects: allelic exclusion; Animals; Anti-Bacterial Agents - pharmacology; antibiotics; antigenic variation; Cinnamates; Cloning, Molecular; drug resistance; Drug Resistance - genetics; expression site switching; Fluorescent Antibody Technique; g418; gene expression; Gene Expression Regulation; Genes, Protozoan; Genetic Markers; hygromycin B; Hygromycin B - analogs & derivatives; Hygromycin B - pharmacology; kinases; mono-allelic expression; reporter genes; Telomere - physiology; telomeric silencing; Time Factors; transcription (genetics); Transcription, Genetic; Trypanosoma brucei; Trypanosoma brucei brucei - cytology; Trypanosoma brucei brucei - genetics; variant surface glycoproteins; Variant Surface Glycoproteins, Trypanosoma - genetics; Variant Surface Glycoproteins, Trypanosoma - metabolism
• ISSN: 0261-4189; 1460-2075; 1460-2075
• DOI: 10.1093/emboj/18.17.4846

Trypanosoma brucei has 20 similar telomeric‐expression sites for variant surface glycoprotein genes. Expression sites appear to be controlled at the level of transcription initiation, resulting in only one site being active at any time. Switching between expression sites occurs at a low rate. To analyse the switching mechanism, we used trypanosomes with two expression sites tagged with two different drug‐resistance genes and selected these on agarose plates containing both drugs. Double‐resistant clones arose at a low frequency of 10−7 per cell, but these behaved as if they rapidly switched between the two tagged expression sites and lost double resistance in the absence of selection. Using in situ hybridization we found that only 10% of the double‐resistant cells had two fluorescent spots corresponding to transcribed expression sites. Our results suggest that: (i) a double expressor is not a stable intermediate in expression site switching; (ii) expression sites are not independently switched on and off; and (iii) expression sites can be in a ‘pre‐active’ silent state from which they can be readily activated.