MCM-BP is required for repression of life-cycle specific genes transcribed by RNA polymerase I in the mammalian infectious form of Trypanosoma brucei

• Published in: PloS one Vol. 8; no. 2; p. e57001
• Main Authors: Kim, Hee-Sook, Park, Sung Hee, Günzl, Arthur, Cross, George A M
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 25-02-2013; Public Library of Science (PLoS)
• Subjects: Allelic exclusion; Amino Acid Sequence; Animals; Antigens; Biology; Chromatin; Chromosomes; Deoxyribonucleic acid; Developmental biology; Dihydrofolate reductase; DNA; DNA damage; DNA helicase; DNA-directed RNA polymerase; Gene expression; Gene loci; Gene Silencing; Genes; Genetic engineering; Genomes; Genomics; Glycoproteins; Humans; Immunity; Infectious diseases; Insects; Insertional mutagenesis; Laboratories; Mass spectrometry; Mass spectroscopy; Molecular Sequence Data; Nuclear Proteins - chemistry; Nuclear Proteins - physiology; Parasites; Parasitology; Protein binding; Proteins; Protozoa; Pseudogenes; Ribonucleic acid; RNA; RNA polymerase; RNA Polymerase I - metabolism; RNA-mediated interference; Sequence Homology, Amino Acid; Telomerase; Transcription, Genetic; Transposons; Trypanosoma brucei; Trypanosoma brucei brucei - isolation & purification; Trypanosoma brucei brucei - physiology; Trypanosome; Trypanosomiasis - genetics; Trypanosomiasis - parasitology; Variant surface glycoprotein; Viability; Yeast; New York; United States > US; Connecticut
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0057001

Trypanosoma brucei variant surface glycoprotein (VSG) expression is a classic example of allelic exclusion. While the genome of T. brucei contains >2,000 VSG genes and VSG pseudogenes, only one allele is expressed at the surface of each infectious trypanosome and the others are repressed. Along with recombinatorial VSG switching, allelic exclusion provides a major host evasion mechanism for trypanosomes, a phenomenon known as antigenic variation. To extend our understanding of how trypanosomes escape host immunity by differential expression of VSGs, we attempted to identify genes that contribute to VSG silencing, by performing a loss-of-silencing screen in T. brucei using a transposon-mediated random insertional mutagenesis. One identified gene, which we initially named LOS1, encodes a T. brucei MCM-Binding Protein (TbMCM-BP). Here we show that TbMCM-BP is essential for viability of infectious bloodstream-form (BF) trypanosome and is required for proper cell-cycle progression. Tandem affinity purification of TbMCM-BP followed by mass spectrometry identified four subunits (MCM4-MCM7) of the T. brucei MCM complex, a replicative helicase, and MCM8, a subunit that is uniquely co-purified with TbMCM-BP. TbMCM-BP is required not only for repression of subtelomeric VSGs but also for silencing of life-cycle specific, insect-stage genes, procyclin and procyclin-associated genes (PAGs), that are normally repressed in BF trypanosomes and are transcribed by RNA polymerase I. Our study uncovers a functional link between chromosome maintenance and RNA pol I-mediated gene silencing in T. brucei.