SELECTION FOR ACTIVATION OF A NEW VARIANT SURFACE GLYCOPROTEIN GENE-EXPRESSION SITE IN TRYPANOSOMA-BRUCEI CAN RESULT IN DELETION OF THE OLDONE

The African trypanosome Trypanosoma brucei expresses the active varian t surface glycoprotein (VSG) gene in a telomeric VSG gene expression s ite. We have generated trypanosomes with a neomycin resistance gene in serted behind an active VSG gene expression site promoter, and a hygro mycin resistance gene behind a silent one. By alternating drug selecti on, we could select for trypanosomes that had switched between the two marked VSG gene expression sites, Surprisingly, trypanosomes that had activated a new VSG gene expression site had often lost the old one. Using polymerase chain reaction (PCR), we screened large numbers of sw itched trypanosomes and found that sequences lost invariably included the drug marker near the promoter, as well as the telomeric VSG gene m any tens of kilobases away. We postulate that stable activation of a n ew expression site requires silencing of the old one. If silencing doe s not occur at a sufficient rate by normal switch-off, stable activati on of the new site can only occur if the old site is lost in random de letion events. The fact that we pick up these normally infrequent dele tions, indicates that inactivation of the old VSG expression site coul d be rate limiting during switching in our strain of T. brucei. (C) 19 98 Elsevier Science B.V. All rights reserved.