Nuclear Phosphatidylinositol 5-Phosphatase Is Essential for Allelic Exclusion of Variant Surface Glycoprotein Genes in Trypanosomes

Abstract

Allelic exclusion of variant surface glycoprotein (VSG) genes is essential for African trypanosomes to evade the host antibody response by antigenic variation. The mechanisms by which this parasite expresses only one of its ∼2,000 VSG genes at a time are unknown. We show that nuclear phosphatidylinositol 5-phosphatase (PIP5Pase) interacts with repressor activator protein 1 (RAP1) in a multiprotein complex and functions in the control of VSG allelic exclusion. RAP1 binds PIP5Pase substrate phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P3], and catalytic mutation of PIP5Pase that inhibits PI(3,4,5)P3 dephosphorylation results in simultaneous transcription of VSGs from all telomeric expression sites (ESs) and from silent subtelomeric VSG arrays. PIP5Pase and RAP1 bind to telomeric ESs, especially at 70-bp repeats and telomeres, and their binding is altered by PIP5Pase inactivation or knockdown, implying changes in ES chromatin organization. Our data suggest a model whereby PIP5Pase controls PI(3,4,5)P3 binding by RAP1 and, thus, RAP1 silencing of telomeric and subtelomeric VSG genes. Hence, allelic exclusion of VSG genes may entail control of nuclear phosphoinositides.

Phosphoinositide signaling and regulation in Trypanosoma brucei: Specialized functions in a protozoan pathogen

“…This set of synthesis, cleavage, and modifying enzymes (hereafter referred as PIP/IP-related proteins) produces at least 11 different PIP and IP metabolites ( Fig 1C), some of which have been detected in T. brucei via immunofluorescence or mass spectrometry methods [7,14,15] or predicted to exist based on in vitro enzymatic studies [6,13,18]. T. brucei PIP and IP kinases and phosphatases with different specificities are distributed in distinct subcellular locations, e.g., plasma membrane, endosomes, and nucleus [5,7,9,10,12] (Table 1). The subcellular distribution of PIPs, IPs, and related proteins in T. brucei indicates that they function as a regulatory system in addition to their role in the synthesis of membrane or glycoconjugate structures.…”

“…How such a system regulates silencing and switching of VSG genes is yet unclear, but it might involve the control of PIPs subcellular fluxes and levels. T. brucei expresses a nuclear phosphatidylinositol phosphate 5-phosphatase (PIP5Pase) enzyme that also controls silencing of telomeric and subtelomeric VSG genes [5]. PIP5Pase associates with repressor-activator protein 1 (RAP1) within a 0.9 megadalton protein complex, which also includes protein kinases, phosphatases, chromatin regulatory proteins, and nuclear pore proteins [5,7].…”

“…T. brucei expresses a nuclear phosphatidylinositol phosphate 5-phosphatase (PIP5Pase) enzyme that also controls silencing of telomeric and subtelomeric VSG genes [5]. PIP5Pase associates with repressor-activator protein 1 (RAP1) within a 0.9 megadalton protein complex, which also includes protein kinases, phosphatases, chromatin regulatory proteins, and nuclear pore proteins [5,7]. PIP5-Pase regulation of VSG silencing revolves around the control of phosphatidylinositol 3,4,5-triphosphate (PIP3) levels.…”

“…They also have elaborate mechanisms of gene regulation that control the expression of genes involved in host immune evasion during infection. The control of developmental changes and immune evasion mechanisms entails a complex network of signaling and regulatory processes that includes phosphatidylinositol (PI) phosphates (PIP, also called phosphoinositides) and inositol phosphates (IP) [2][3][4][5][6][7][8]. PIPs and IPs are ubiquitous in eukaryotes and consist of a subset of molecules containing mono or poly phosphorylated inositol ( Fig 1A).…”

“…PIPs and IPs interact with proteins or RNA and regulate numerous cellular functions in eukaryotes. As detailed below, these metabolites and related enzymes function as a regulatory system with essential roles in T. brucei metabolism and development [6], trafficking and organelle biogenesis [9][10][11], Ca 2+ signaling [12], and immune evasion mechanisms [5,7].…”