Selective nuclear export of mRNAs is promoted by DRBD18 in Trypanosoma brucei

Abstract

Kinetoplastids, including Trypanosoma brucei, control gene expression primarily at the posttranscriptional level. Nuclear mRNA export is an important, but understudied, step in this process. The general heterodimeric export factors, Mex67/Mtr2, function in the export of mRNAs and tRNAs in T. brucei, but RNA binding proteins (RBPs) that regulate export processes by controlling the dynamics of Mex67/Mtr2 ribonucleoprotein formation or transport have not been identified. Here, we report that DRBD18, an essential and abundant T. brucei RBP, associates with Mex67/Mtr2 in vivo, likely through its direct interaction with Mtr2. DRBD18 downregulation results in partial accumulation of poly(A)+ mRNA in the nucleus, but has no effect on the localization of intron‐containing or mature tRNAs. Comprehensive analysis of transcriptomes from whole‐cell and cytosol in DRBD18 knockdown parasites demonstrates that depletion of DRBD18 leads to impairment of nuclear export of a subset of mRNAs. CLIP experiments reveal the association of DRBD18 with several of these mRNAs. Moreover, DRBD18 knockdown leads to a partial accumulation of the Mex67/Mtr2 export receptors in the nucleus. Taken together, the current study supports a model in which DRBD18 regulates the selective nuclear export of mRNAs by promoting the mobilization of export competent mRNPs to the cytosol through the nuclear pore complex.

Proteomics Uncovers Novel Components of an Interactive Protein Network Supporting RNA Export in Trypanosomes

“…Interestingly, we also observed RBSR2 that is associated with RRM1, RBSR1, and other RNA-binding proteins that were also identified, such as UBP-1, UBP-2, and ALBA3 ( 121 ). Additionally, DRBD18, identified in TcSub2 and TcMex67 isolates, is involved in mRNA export in association with Mex67/Mtr2 by regulating the mobilization of mRNPs through the nuclear pore complex ( 122 ).…”

The Trypanosoma brucei RNA-binding protein DRBD18 ensures correct mRNA trans splicing and polyadenylation patterns

“…Various lines of evidence indicate that DRBD18 is involved in the export of a subset of mRNAs from the nucleus. DRBD18 interacts directly with MTR2, and coimmunoprecipitates MEX67 ( Mishra et al 2021 ). Moreover, in procyclic forms, depletion of DRBD18 by RNAi caused partial retention of MTR2 and MEX67, and some poly(A) + mRNAs, in the nucleus ( Mishra et al 2021 ).…”

“…DRBD18 interacts directly with MTR2, and coimmunoprecipitates MEX67 ( Mishra et al 2021 ). Moreover, in procyclic forms, depletion of DRBD18 by RNAi caused partial retention of MTR2 and MEX67, and some poly(A) + mRNAs, in the nucleus ( Mishra et al 2021 ). Levels of various mRNAs were affected, but the effects were different depending on whether cytoplasmic or whole-cell mRNAs were analyzed ( Mishra et al 2021 ).…”

“…One of the mRNAs that increased significantly in abundance after DRBD18 depletion in procyclic forms was that encoding RBP10 ( Lott et al 2015 ; Mishra et al 2021 ). During studies of RBP10 regulation, we unexpectedly discovered that DRBD18 depletion led to accumulation of truncated versions of the RBP10 mRNA.…”

Control of Variant Surface Glycoprotein Expression by CFB2 in Trypanosoma brucei and Quantitative Proteomic Connections to Translation and Cytokinesis

“…MKT1 (Tb927.6.4770) is recruited to mRNAs by sequence-specific RNA binding proteins and stabilizes the bound mRNA, while PBP1 (Tb927.8.4540) interacts with MKT1 as well as with poly(A) binding protein 2 ( 13 , 14 ); both MKT1 and PBP1 were recently found to bind VSG mRNA through CFB2 ( 6 ). DRBD18 also registered >35% increased G 2 M-phase accumulation, but this protein is known to promote nuclear mRNA export ( 15 ), so it was not investigated further here.…”